Dissertationen zum Thema „Instestine“

Les cytokines favorisent la défense et l'homéostasie de l'intestin. Parmi les cytokines clés de l'intestin, l'interleukine-22 (IL-22) est produite par les cellules immunitaires et cible principalement les cellules épithéliales. l'IL-22 protège l'intestin contre les agents pathogènes en induisant l'expression de peptides antimicrobiens et en favorisant la réparation des tissus. Une dysrégulation de cette cytokine peut conduire à des maladies inflammatoire de l'intestin et au cancer. Au cours du développement, le tube digestif post-embryonnaire est colonisé par des micro-organismes commensaux qui favorisent la maturation de l'intestin et de son système immunitaire. Cependant, il est encore incertain si et comment l'IL-22 joue un rôle dans le développement et la maturation de l’intestin au cours de cette période critique. Le poisson-zèbre nous permet de visualiser et de manipuler les processus physiologiques in vivo dès le début du développement en raison de son développement externe et de sa transparence.Au cours de ma thèse de doctorat, j'ai cherché à décrypter la fonction de l'IL-22 dans le développement de l'intestin. Mes données montrent que l'il22 est exprimée dans les cellules épithéliales intestinales des larves avant que les lymphocytes exprimant l'IL-22 n'apparaissent dans l'intestin. J'ai identifié les cellules entéroendocrines (EEC), un sous-type de cellules épithéliales intestinales, comme la principale source d'il22 chez les larves. De plus, j'ai révélé la conservation de la voie de signalisation de l'IL-22, sa régulation transcriptionnelle par la détection des microbes et sa fonction antibactérienne dans l'intestin. Mes données les plus récentes suggèrent que l'expression d’il22 peut également être induite par l'activation de Trpa1, un récepteur connu pour reconnaître les métabolites du tryptophane chez le poissons-zèbre et la souris. Enfin, j'ai découvert un nouveau rôle de l'IL-22 dans la modulation de la motilité intestinale chez les poissons-zèbres. Mécaniquement, une dysbiose a été observée chez les il22 mutants, ainsi que des défauts potentiels dans la production de métabolites dérivés des bactéries dans l'intestin. De plus, un dysfonctionnement a été constaté dans la fonction des EEC, caractérisé par une expression hormonale dysrégulée. Il a été intéressant de constater que le mutant exprime des niveaux plus bas de 5-HT, une hormone importante régulant la motilité intestinale chez les mammifères et les poissons-zèbres. Nous avons démontré que l'administration de celle-ci est capable de rétablir la motilité intestinale, ce qui signifie que 5-HT est suffisante pour restaurer une motilité intestinale correcte. Enfin, j'ai découvert la conservation du défaut de motilité intestinale chez les souris en bas âge, ce qui suggère la conservation de cette nouvelle fonction de l'IL-22 chez les mammifères tôt au cours du développement. Dans l'ensemble, ce projet contribue à une meilleure compréhension de la manière dont les cytokines orchestrent le développement et la maturation de l'intestin au cours des premières étapes de la vie
Cytokines promote gut defense and homeostasis. Among key gut cytokines, interleukin-22 (IL-22) is produced by immune cells and primarily targets epithelial cells. IL-22 protects the gut from pathogens by inducing anti-microbial peptides expression and promoting tissue repair. Dysregulation of this cytokine can lead to inflammatory bowel disease and cancer. During development, the post-embryonic gut is colonized by commensal microorganisms that promote maturation of the gut and its immune system. Nevertheless, whether and how cytokines such as IL-22 play a role in gut organ development and maturation during this critical time window remains unclear. The zebrafish allows us to visualize and manipulate physiological processes in vivo since early development due to its external development and transparency.During my PhD, I wanted to decipher the function of il22 in the developing gut. My data show that il22 is expressed in larval gut epithelial cells before il22-expressing lymphocytes appear in the gut. I identified enteroendocrine cells (EECs), a gut epithelial cell subtype, as the main source of il22 in larvae. Furthermore, I revealed conservation of the IL-22 signaling pathway, its transcriptional regulation by microbe sensing, and its antibacterial function in the gut. My latest data suggest that il22 expression can also be induced by the activation of Trpa1, a receptor known to recognize tryptophan metabolites in zebrafish and mice. Finally, I found a novel role of IL-22 in modulating gut motility in zebrafish. Mechanistically, dysbiosis was observed in il22-/-, along with potential defects in the production of bacteria-derived metabolites in the gut. Surprisingly, we found that the microbiota from wild-type larvae was able to restore the gut motility impairment of il22-/-, highlighting the important role of the microbiota in il22-mediated regulation of this process. Furthermore, an impairment was found in EECs function, which are known to be sensitive and respond to microbial cues. The dysregulation of EECs was characterized by abnormal hormone expression, specifically reduced levels of serotonin (5-HT), a critical hormone regulating gut motility in both zebrafish and mammals. External administration of 5-HT successfully rescued the gut motility phenotype of il22-/-, indicating that 5-HT is sufficient to restore proper gut motility. Finally, I found conservation of the gut motility defect in early life mice, suggesting the conservation of this novel IL-22 function in mammals. Altogether, this project contributes to a better understanding of how cytokines orchestrate gut development and maturation during the early stages of vertebrate life

Les cellules mésenchymateuses sont des cellules non-hématopoïétiques, non-épithéliales et non-endothéliales présentes dans tous les organes. Elles contribuent à la structure des organes en synthétisant la matrice extracellulaire (MEC). En plus de leur rôle dans la synthèse de la MEC, un nombre croissant d’études au cours de ces vingt dernières années s’accordent sur le fait que les cellules mésenchymateuses (CM) jouent un rôle essentiel dans l’homéostasie de l’intestin. Cependant, le manque de marqueurs et d’outils spécifiques pour étudier ces cellules ont empêché une meilleure compréhension de leurs fonctions. La plupart des CMs intestinales expriment podoplanin (aussi connu sous le nom gp38), un marqueur des CMs des ganglions lymphatiques. En utilisant de nouveaux marqueurs et modèles murins pour étudier les CMs qui expriment le récepteur de la lymphotoxin-ß (LTßR), un récepteur impliqué dans l’interaction avec les cellules du système immunitaire, nous avons identifié différentes sous-population de CMs gp38+ avec des fonctions spécifiques dans l’homéostasie et l’immunité intestinale. En traçant génétiquement les CMs gp38+LTßR+ nous avons identifié un lignage spécifique de fibroblastes sous-épithéliaux qui expriment PDGFRα, appelé par la suite LTßR-SF. Nous avons montré que les ce lignage possède une fonction unique dans les premières semaines après la naissance. En effet, les LTßR-SFs se développent au moment du sevrage des souris de manière indépendante de la colonisation microbienne où ils promeuvent la maturation de la barrière intestinale, y compris la différentiation des cellules caliciformes et des cellules de Paneth qui protège l’intestin des bactéries, ainsi que le développement des cellules dendritiques CD103+CD11b+, qui régulent la tolérance intestinale. Les LTßR-SFs expriment un set de gènes essentiels pour l’homéostasie de l’épithélium et la régulation du système immunitaire tels que bmp4, sfrp3, dll1, vcam, aldh1a3 et cox1. Enfin, nous avons montré que la voie de signalisation de PDGFRα est nécessaire pour l’acquisition de ces fonctions par les LTßR-SFs. En conclusion, nos résultats montrent qu’une sous-population de CMs gp38+ joue un rôle essentiel dans le développement de la barrière intestinale au moment du sevrage en coordonnant la maturation de l’épithélium et du système immunitaire via la voie de signalisation de PDGFRα
Mesenchymal cells (MCs) are non-immune, non-epithelial and non-endothelial cells present in all organs. They contribute to the organ’s structure by producing extracellular matrix (ECM). In addition to their role as ECM-producing cells, increasing evidence suggests that they play an active role in intestinal homeostasis. However, the lack of specific markers and tools to investigate MCs hindered a deeper understanding of their function(s). In the intestine, a major fraction of MCs expresses podoplanin (also known as gp38) a marker for MCs in lymphoid organs. Using new markers and reporter mice to study MCs that express the Lymphotoxin-ß Receptor (LTßR), a receptor involved in the crosstalk with immune cells, we identified distinct subsets of gp38+ MCs with specific functions in intestinal homeostasis and immunity. Inducible lineage tracing of gp38+LTßR+ MCs identifies a specific lineage of PDGFRα+ mesenchymal cells located closely to epithelial cells. We show that the lineage of PDGFRα+ subepithelial fibroblasts derived from LTßR+ progenitors (LTßR-SF) has a unique function in the first few weeks after birth. Indeed, this specific lineage develops around weaning independently of microbial colonization, and promotes maturation of the epithelial barrier, including differentiation of Paneth and goblet cells, involved in bacterial protection, and development of CD103+CD11b+ dendritic cells, a specific subset of immune cells involved in intestinal tolerance. LTßR-SF overexpress a set of genes essential for epithelial homeostasis and immune regulation, including bmp4, sfrp3, dll1, vcam, aldh1a3 and cox1. We further show that PDGFRα signaling in LTßR-SF is required to imprint both epithelial and immune function. Taken together, our results show that a subset of gp38+ MCs derived from LTßR+ progenitors plays an essential role in intestinal barrier development at weaning, by coordinating immune and epithelial maturation through PDGFRα signaling

SOARES, Fábio Alves. Análise da freqüência de anorretocele em mulheres adultas com evacuação obstruída, comparando com a paridade e idade, utilizando cinedefecografia e eletromanometria anorretal. 2006. 66 f. Dissertação (Mestrado em Cirurgia) - Universidade Federal do Ceará. Faculdade de Medicina, Fortaleza, 2006.
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The aim of this study is to analyse the frequence of anorectocele in adult women with obstructed defecation accordind to parity and age by means of cinedefaecography and anorectal eletromanometry. Forty-five adult women complaining of obstructed defecation were evaluated, with mean age of 46.3 years (23-72) and mean SCCC-C score of 13.3 points (6-23). Fifteen (33.3%) patients were nulliparous, seven (15,6%) primiparous and 23 (51,1%) multiparous, with mean parity per patient of 2.8 (0-11), considering only vaginal deliveries. Eighteen (60%) had a history of episiotomy, fourteen (46,7%) delivered macrossomic children and two (6,7%) had history of forceps-assisted delivery. Anal hipertony was verified in fourteen (31,1%) patients, while anal hipotony was present in eight (17,8%). Anismus was identified in thirteen (28,9%) patients. Anorecoceles were demonstrated in 34 (75,6%) patients, with mean size (TAR) of 24,8 mm (0-64). Thirty-six (80%) patients presented excessive perineal descent (DPM), rectal mucosal prolapse (PM) in 17 (37,8%) and rectoanal intussusception (IRA) in twelve (26,7%). There were no correlations between anorectocele and anal hipertony (p = 0,7171), anismus (p = 0,4666), IRA (p= 0,6991), PM (p = 0,2279), parity comparing nulliparous and multiparous patients (p = 1,000), episiotomy (p = 1,0000), forceps assistance (p = 1,0000) and delivery of macrossomic children (p = 1,0000). There were also no correlation between TAR and PMR (p =0,0883), PVM (p = 0,7327), parity (p = 0,4987) or age (p = 0,8603). There were correlations between anorectocele and DPM (p = 0,0275), score of SCCC-C (p =0,0082) and anal hipotony (p = 0,0141). In conclusion, anorectocele frequence is high and doesn’t correlate to parity, age but correlates to anal hipotony, DPM andconstipation.
O objetivo é avaliar a freqüência e o tamanho de anorretocele em mulheres adultas com evacuação obstruída, correlacionando-os com paridade, idade e parâmetros clínicos, utilizando cinedefecografia e eletromanometria. Foram avaliadas 45 mulheres adultas, com idade média de 46,3 anos (23-73) e sintomas de evacuação obstruída, com escore médio de 13,3 (6-23) pontos, segundo o Sistema de Classificação da Cleveland Clinic para Constipação (SCCC-C). Os parâmetros avaliados foram idade, dados obstétricos, escore do, SCCC-C, dados manométricos e achados de cinedefecografia.. Quinze (33,3%) pacientes eram nulíparas, 7 (15,6%) primíparas e 23 (51,1%) multíparas, com média de 2,8 (0-11) partos vaginais por paciente. Dezoito (60,0%) pacientes haviam sido submetidas a parto vaginal com episiotomia, sendo verificado feto macrossômico em 14 (46,7%) e aplicação de fórcipe em duas (6,7%) . Foi observada hipertonia esfincteriana em 14 (31,1%) e hipotonia em 8 (17,8%) pacientes. Foi identificado anismus em 13 (28,9%) pacientes. Foram demonstradas anorretoceles em 34 (75,6%) pacientes com tamanho (TAR) médio de 24,8 mm (0 - 64). Foram verificados descenso perineal móvel acentuado (DPM) em 36 (80%) pacientes, prolapso mucoso (PM) em 17 (37,8%) e intussuscepção reto-anal (IRA) em doze (26,7%). Não houve correlação entre anorretocele e hipertonia esfincteriana (p = 0,7171), anismus (p = 0,4666), IRA (p = 0,6991), PM (p= 0,2279), paridade comparando-se nulíparas e multíparas (p =1,000), episiotomia (p = 1,0000), uso de fórcipe (p = 1,0000), parto de feto macrossômico (p = 1,0000). Não houve correlação entre TAR e PMR (p = 0,0883), PVM (p = 0,7327), paridade (p = 0,4987) e idade (p = 0,8603). Houve correlação entre anorretocele e DPM (p = 0,0275), escore de constipação do SCCC-C (p = 0,0082) e hipotonia esfincteriana (p = 0,0141). Conclui-se que a freqüência de anorretocele foi elevada, não se correlacionou com paridade e idade, associando-se com hipotonia esfincteriana, DPM e constipação.

Introducción. Actualmente la intervención de Hartmann es una alternativa válida en el tratamiento de patologías de colon izquierdo o recto en pacientes ASA IV, con peritonitis fecaloidea, desnutridos, inmunodeprimidos o con inestabilidad hemodinámica. Se emplea en casos de riesgo de dehiscencia anastomótica, de recidiva tumoral o de incontinencia anal. Sin embargo, los factores relacionados con la decisión de reconstruir el tránsito intestinal no están tan bien establecidos. El objetivo principal del estudio consiste en determinar si existen factores que pueden predecir la reconstrucción intestinal tras una intervención de Hartmann. El análisis de las complicaciones de estas intervenciones y la determinación de los factores predictivos de ausencia de reconstrucción del tránsito intestinal pueden ayudar a realizar una precisa selección de los pacientes y ofrecer información preoperatoria individualizada sobre los resultados más probables de la intervención. Material y métodos. Estudio observacional, retrospectivo y longitudinal que incluye todos los pacientes sometidos a intervención de Hartmann desde Enero de 1999 hasta Diciembre 2014 en un hospital terciario universitario. No se excluyó ningún paciente candidato a una posible reconstrucción del tránsito intestinal. Las variables recogidas fueron clasificadas en 1) específicas del paciente: edad, sexo, IMC, riesgo ASA, índice de Charlson, incontinencia anal; 2) específicas de la enfermedad: tipo de patología (benigna vs maligna), diagnóstico principal, estadio tumoral, grado de contaminación peritoneal, 3) específicas del tratamiento: indicación de procedimiento de Hartmann, transfusión perioperatoria y postoperatoria, procedimiento quirúrgico principal, tipo de cirugía (programada vs urgente), tipo de cirujano (general vs colorrectal), longitud del muñón rectal, clasificación Clavien- Dindo, reingresos, causas de no reversión de la intervención de Hartmann. Se realizó un estudio descriptivo de la muestra. Se evaluó la asociación estadística entre las variables cualitativas con la prueba de 2 o prueba exacta de Fisher. Se compararon grupos mediante los tests U de Mann-Whitney o Kruskal-Wallis en función del tipo de variables. Se realizó un análisis univariante y multivariante mediante una regresión logística binaria. Además se realizó un árbol de clasificación y regresión. Por último, se elaboraron las curvas COR de cada modelo y se compararon entre sí mediante la prueba de DeLong. Resultados. Se incluyeron 533 pacientes consecutivos a los que se realizó un procedimiento de Hartmann. 110 (20,6%) pacientes fueron intervenidos para la reconstrucción intestinal. La edad media fue de 71,7 años. En el análisis multivariante se obtuvieron como factores independientes predictivos de presentar mayor probabilidad de reconstrucción del tránsito la edad inferior a 69 años, el grado ASA I ó II, la indicación de intervención de Hartmann por dehiscencia anastomótica y la altura del muñón rectal por encima del promontorio sacro o a la altura del mismo. Sin embargo, los factores independientes predictivos de una menor probabilidad de reconstrucción fueron la incontinencia anal, estadio IV, recibir transfusión postoperatoria o realizar intervención de Hartmann de forma programada. Del árbol de clasificación se deduce que un paciente de edad inferior a 69 años, que presente baja comorbilidad, con un muñón rectal en el promontorio o por encima de éste y que no haya requerido transfusión tendrá un 85% de probabilidad de reconstrucción intestinal. Discusión. La identificación de factores predictivos de restauración de la continuidad intestinal puede servir para aconsejar a los pacientes y ayudar a los cirujanos a escoger la mejor opción, tanto antes de la realización de una intervención de Hartmann, como en el momento de indicar su reconstrucción. Conclusión. Edad, riesgo ASA, indicación de procedimiento de Hartmann, longitud del muñón rectal, incontinencia anal, estadio tumoral, transfusión postoperatoria y cirugía programada pueden predecir la reconstrucción de la intervención de Hartmann.
Introduction. Hartmann´s procedure is still a valid alternative in the treatment of pathologies of the left colon or rectum in patients having an ASA score IV, feculent peritonitis, malnutrition, immunosuppression or with hemodynamic instability. Hartmann´s procedure is mainly indicated in cases of high risk of anastomotic leakage, local tumor recurrence or anal incontinence. However, the factors related to the decision to restore intestinal continuity are not well established. The main objective of this study was to determine predictive factors of Hartmann´s reversal. The analysis of the morbidity of those interventions and the identification of predictive factors of intestinal transit reconstruction could ease an accurate selection of patients and give individualized preoperative counselling providing information on the most likely outcomes of the intervention. Material and methods. A retrospective observational study in which all consecutive patients that underwent Hartmann´s procedure from January 1999 to December 2014 in a tertiary University Hospital were included. No patient with a possible intestinal continuity restoration was excluded. The data collected were classified into 1) patient-specific: age, sex, body mass index, ASA score, Charlson index, anal incontinence; 2) disease-specific: type of disorder (benign vs malignant), main diagnosis, tumor stage, degree of peritoneal contamination, 3) treatment-specific: period of years of surgery, indication of Hartmann´s procedure, perioperative and postoperative transfusion, main surgical procedure, type of surgery (elective vs urgent), type of surgeon (general vs colorrectal), length of the rectal stump, Clavien-Dindo classification, readmission rate, causes of nonreversal Hartmann´s procedure. A descriptive analysis was performed. The 2 test or the Fisher exact test were used for categorical variables. Comparisons between groups were made using Mann-Whitney U test or Kruskal-Wallis test for continuous variables, where appropriate. Univariate and multivariate binary logistic regression model were used. Further, a classification and regression tree was performed. Finally, COR curves of each model were elaborated and compared with the DeLong test. Results. A total of 533 consecutive patients underwent Hartmann’s procedure. 110 (20,8%) patients underwent Hartmann´s reversal procedure. Mean age was 71,7 years. Multivariate analysis showed that the independent predictors of higher probability of intestinal transit reversal were age lower than 69 years, ASA grade I or II, indication of HP for anastomotic leak and the rectal stump above or at the sacral promontory. However, the independent factors related to a reduced probability of intestinal reconstruction following HP were anal incontinence, stage IV, postoperative transfusion or elective Hartmann's intervention. From the classification tree it is deduced that a patient below 69 years of age who presents low comorbidity, with a rectal stump at or above the promontory and that did not require perioperative transfusion would have 85% of probability of intestinal transit reconstruction. Discussion. Identification of predictive factors of intestinal continuity restoration may help surgeons to inform the patient and to choose the better option, both before performing a Hartmann procedure, and at the time of indicating reconstruction of intestinal continuity. Conclusion. Age, ASA, indication of Hartmann’s procedure, length of rectal stump, anal incontinence, tumor stage, postoperative transfusion and elective surgery can predict Hartmann’s reversal.

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O Bacillus sphaericus (Bsp) é uma bactéria entomopatógena eficiente para o controle de Culex quinquefasciatus, um importante vetor da filariose e arboviroses. O fator larvicida do Bsp é a toxina binária (Bin) e a sua ação em C. quinquefasciatus depende da ligação ao receptor Cqm1. A ausência deste receptor no epitélio intestinal é o principal mecanismo de resistência à toxina Bin. Larvas resistentes a esta toxina, são susceptíveis ao Bsp IAB59 que, além da Bin, produz as toxinas Cry48Aa e Cry49Aa. O principal objetivo deste estudo foi caracterizar os efeitos de toxinas do Bsp nas células do epitélio intestinal de C. quinquefasciatus, utilizando como modelos larvas de uma colônia susceptível a todas as toxinas estudadas (S), de uma colônia resistente à toxina Bin (R2362) e de uma colônia resistente à Bin e Cry48Aa/Cry49Aa (RIAB59). Na primeira etapa, larvas não tratadas das colônias S e R2362 disseccionadas 30 min, 4, 6 e 48 h após a muda para o 4° estádio foram fixadas e processadas para microscopia eletrônica de transmissão (MET). A avaliação morfológica do epitélio intestinal mostrou que células de larvas R2362, ao final do 4° estádio, são caracterizadas por um intenso acúmulo de inclusões lipídicas, sugerindo que a ausência da a-glicosidase Cqm1 pode estar envolvida com alterações no metabolismo. Para caracterizar os efeitos causados pelas toxinas no epitélio intestinal, as larvas foram disseccionadas 1 e 6 h após o tratamento e processadas para MET. A avaliação ultra-estrutural da ação da toxina Bin nas células do epitélio intestinal mostrou que os principais efeitos em larvas S foram a vacuolização citoplasmática e destruição de microvilosidades. Estes foram observados exclusivamente em células que possuem o receptor Cqm1, demonstrando que esta molécula é essencial para mediar a ação da toxina Bin. Em células de larvas das colônias S e R2362, susceptíveis à Cry48Aa/Cry49Aa, o principal efeito destas toxinas foi a vacuolização mitocondrial, e este parece estar associado à Cry48Aa que possui estrutura de 3 domínios típica de toxinas da família Cry. Efeitos similares aos da toxina Bin também foram observados e parecem resultantes da ação da Cry49Aa que possui homologia com toxinas do tipo binária. Os dados mostram que a Cry48Aa/Cry49Aa possui uma ação complexa nas células e seu sítio de ligação é diferente do Cqm1. Combinações da toxina Bin com as toxinas Cry11Aa e Cyt1Aa do Bti também provocaram alterações em células de larvas R2362, desprovidas do receptor Cqm1. A combinação Bin/Cry11Aa causou efeitos similares aos induzidos pela toxina Cry48Aa/Cry49Aa, e sugerem que as toxinas de 3 domínios, Cry11Aa e Cry48Aa, podem mediar os efeitos das toxinas Bin e Cry49Aa, respectivamente, nos modelos estudados. A combinação Bin/Cyt1Aa provocou efeitos drásticos como a perda precoce de microvilosidades e a lise celular, característica da toxina Cyt1Aa que apresenta ação citolítica. Os resultados deste trabalho contribuem para o entendimento do modo de ação de toxinas do Bsp, bem como do efeito sinergístico de suas toxinas com aquelas do Bti

Les invalidations individuelles et simultanées des protéines de mise en faisceau de l’actine, villine, I-fimbrine et espine, n’abolissent pas la formation de microvillosités intestinales. Leur formation en absence de protéines de mise en faisceau est discutée. Quelle est alors la fonction de ces protéines ? L’étude du rôle de la villine démontre que son activité de fragmentation augmente la dynamique d’un mouvement dépendant de l’actine. La I-fimbrine apparaît comme un acteur central de la polarité et de la stabilité du réseau apical des entérocytes. Enfin, un modèle est envisagé selon lequel la villine agirait comme un facteur déstabilisant du cytosquelette de la microvillosité compensé en partie ou en totalité par le rôle stabilisateur de la I-fimbrine. Ces deux protéines seraient des protagonistes permettant d’une part le maintien de la microvillosité en équilibre dynamique en conditions normales, et d’autre part son remodelage rapide en réponse à des situations de stress
The individual and the simultaneous invalidations of the genes encoding for the bundling proteins, villin, I-fimbrin ans espin do not abolish the formation of intestinal microvilli in mice. The possibilities of formation of this structure in the absence of bundling proteins is discussed. What is then the role of these proteins in the enterocytes ? We demonstrate that villin actin severing property enhances an actin-based movement. We also report polarity and stability defects of the apical network of the enterocytes lacking I-fimbrin. A model in which villin acts as a destabilising factor of the cytoskeleton of the microvilli that would be compensated by the stabilising effect of I-fimbrin is proposed. These two proteins would be two protagonists, on the one hand maintaining the microvilli in a dynamic equilibrium in normal conditions, and on the other hand allowing a fast remodelling of the structure in response to stresses

La maladie de Crohn (MC) est une affection inflammatoire chronique du tube digestif dont l’étiologie est multifactorielle. Les lésions intestinales des patients atteints de MC sont anormalement colonisées par des souches pathogènes d’Escherichia coli appartenant au pathovar AIEC pour «Adherent-Invasive Escherichia coli ». Ces souches sont capables d’adhérer et d’envahir les cellules épithéliales intestinales, et ont la capacité de survivre et de se multiplier en macrophages en induisant une synthèse intense de cytokines pro-inflammatoires. Les AIEC pourraient ainsi être impliquées dans l’induction et/ou l’entretien de l’état inflammatoire de la muqueuse intestinale.L’objectif de ce travail est d’identifier les déterminants bactériens des AIEC qui vont intervenir dans les étapes précoces de l’implantation des AIEC au niveau intestinal et de définir quel est le rôle des facteurs micro-environnementaux de l’hôte dans cette implantation.Nous montrons que l’AIEC LF82 possède une activité mucinolytique qui est portée par le gène vat-AIEC et qui favorise la traversée du mucus et la colonisation intestinale. Nous avons retrouvé ce gène chez 42% des souches AIEC isolées de patients atteints de MC, et chez 97% des souches AIEC appartenant au phylogroupe B2. Par ailleurs, nous avons montré que les sels biliaires augmentent l’expression de cette mucinase.Nous avons ensuite étudié l’influence des sels biliaires sur l’expression globale des gènes de la souche LF82. Les sels biliaires modifient profondément le métabolisme de la souche, induisant une diminution globale des voies de biosynthèse (protéines, lipides) et une augmentation des voies de dégradation (alcools, acides carboxyliques, polyamines, …). L’étude du catabolisme de l’éthanolamine et du propanediol indique que les AIEC pourraient se servir de ces substrats pour s’implanter au sein de la flore iléale. De plus, les analyses transcriptomiques révèlent que les sels biliaires augmentent l’expression de gènes codant des facteurs de virulence comme l’invasine IbeA, les systèmes de sécrétion de type VI et la yersiniabactine. Nous montrons également qu’ils favorisent la formation de biofilm chez les souches AIEC.Ces données indiquent que les sels biliaires constituent un signal permettant à la souche AIEC LF82 de mettre en place différentes voies métaboliques et déterminants bactériens nécessaires à son implantation au niveau intestinal.Mots-clé : Escherichia coli, maladie de Crohn, mucines, serine protéase, mucinase, AIEC
The etiology of Crohn's disease (CD) involves disorders in host genetic factors and intestinal microbiota. Ileal mucosa of CD patients is often abnormally colonized by adherent-invasive Escherichia coli (AIEC). These strains isolated from the intestinal mucosa of CD patients are able to adhere to intestinal epithelial cells (IECs). This adhesion to IECs promotes the invasion of cells by the bacteria. Furthermore, the invasive ability of AIEC strains allows bacteria to translocate across the human intestinal epithelium, move into the deep tissues and activate immune cells continuously upon arrival. Thus AIEC could be involved in the inflammatory state of the intestinal mucous membrane. The aim of this study was to identify components of AIEC virulence, which might favor their implantation in the gut of CD patients and to define the role of several chemical factors from the ileal environment. Here, we reported a protease called Vat-AIEC from AIEC which favors the penetration of AIEC through the mucus layer and enhances gut colonization. The screening of E. coli strains isolated from CD patients revealed a preferential vat-AIEC association with AIEC strains belonging to the B2 phylogroup. Besides, Vat-AIEC transcription was increased with bile salts from the ileum environment. Then a global RNA sequencing (RNA-seq) of E. coli LF82 has been used to observe the impact of bile salts on the expression of bacterial genes. The results demonstrate the explosive effect of bile salts with a dysregulation of about 40% of the genome, with a global upregulation of genes involved in degradation and downregulation of those implicated in several biosynthesis. Our results show that LF82 use ethanolamine as a nitrogen source and propane diol as a carbon source, which can favor their colonization in the gut compared to the other bacteria. We also studied virulence genes expression in the presence of bile salts. They increase the expression of several virulence factors like the IbeA invasion, the type 6 secretion systems and the yersiniabactin. Furthermore, we noticed an increased expression of genes implicated in biofilm formation. These results improve the understanding of the global regulatory network in the presence of bile salts and thus of AIEC implantation in the human gut of CD patients

Na presente dissertação objetivou-se avaliar a reatividade vascular e a repercussão do pré-tratamento com sinvastatina (SINV, 20mg/kg, p.o.), sob esse parâmetro, em artéria mesentérica superior (AMS) e pulmonar (AP) de ratos Wistar submetidos a 45 minutos de isquemia e 2 horas de reperfusão intestinal (iIR). Nas artérias avaliou-se a função dependente e independente do endotélio e a contração induzida pela fenilefrina agonista seletivo a1-adrenérgico, em preparações com e sem endotélio e na presença ou ausência de inibidores da NOS e da iNOS (L-NAME e 1400W, respectivamente). Pode-se concluir que a iIR modificou a reatividade vascular em AP, porém não em AMS. Na artéria pulmonar, a iIR causou disfunção endotelial por meio de ativação da iNOS, a qual levou à diminuição da resposta vasodilatadora dependente do endotélio e à hiporreatividade contrátil. Além disso, confirmando a segunda hipótese, uma única dose de sinvastatina antes da iIR previne as alterações de reatividade vascular, o que demonstra o efeito benéfico desse fármaco nessa doença.
This work aimed to evaluate the vascular reactivity and the effect of pre-treatment with simvastatin (SIMV, 20mg/kg, p.o.), under this parameter, in the superior mesenteric artery (SMA) and pulmonary artery (PA) of rats subjected to 45 minutes of ischemia and 2 hours of reperfusion (iIR). In these arteries, it was evaluated the endothelium-dependent and -independent function and the contraction induced by phenylephrine the selective a1-adrenergic agonist, in preparations with and without endothelium and at the presence or absence of non-selective NOS and of iNOS inhibitors (L-NAME and 1400W, respectively). It can be concluded that the iIR modified vascular reactivity in PA, but not in SMA. In the pulmonary artery, the iIR caused endothelial dysfunction via activation of iNOS, which led to decreased endothelium-dependent vasodilator response and to contractile hyporeactivity. Moreover, confirming the second hypothesis, a single dose of simvastatin prior to iIR prevents changes in vascular reactivity, which shows the beneficial effects of this drug in this disease.