Introduction: les progrès thérapeutiques pour la prise en charge du carcinome urothélial de la vessie, ou tumeur de vessie infiltrant le muscle (TVIM), sont faibles depuis deux décennies, notamment sur le plan de la chimiothérapie, tant dans les formes localisées que métastatiques. L'identification de nouveaux biomarqueurs pouvant améliorer la prise en charge et le pronostic des patients apparait indispensable, notamment pour prédire la réponse aux traitements systémiques. Notre travail a cherché : (i) à identifier des facteurs biologiques prédictifs de rechute après les chimiothérapies adjuvantes (CA) utilisées actuellement, (ii) à déterminer si les altérations moléculaires, cibles de nouvelles thérapies, identifiées dans la tumeur vésicale sont conservées dans les métastases.Objectifs:(i) Rechercher une association entre la rechute après CA et l'expression de six biomarqueurs, ERCC1, Emmprin, Survivin, MDR1, p53 et topoisomérase IIα, dans la tumeur vésicale et/ou une métastase ganglionnaire(ii) Etudier l'hétérogénéité des mutations de FGFR3 dans les TVIM localement avancéesMatériels et Méthodes: Une cohorte rétrospective de 226 patients traités par cystectomie et CA, et une collection tumorale d'échantillons représentatifs de la tumeur primitive et d'une métastase ganglionnaire ont été constituées. L'expression des six biomarqueurs a été déterminée en immunohistochimie sur ces échantillons. Le statut mutationnel de FGFR3 a été déterminé en SNaPshot sur 84 paires tumeur primitive/métastase ganglionnaire.Résultats: Les survies observées dans la cohorte étaient proches de celles rapportées dans la littérature pour des patients de mêmes stades. La collection tumorale étudiée semble donc représentative des TVIM traitées en France. Aucun des six biomarqueurs n'était retrouvé comme prédictif de rechute après traitement. Une mutation de FGFR3 était retrouvée dans 4 (4,7%) des 84 échantillons appariés sans aucune discordance entre tumeur primitive et métastase ganglionnaire.Conclusions: L'expression de ces six biomarqueurs étudiés en immunohistochimie n'est pas associée à un risque de rechute après CA. Le statut mutationnel de FGFR3 est conservé au cours de la dissémination métastatique, et ses mutations activatrices sont présentes dans près de 5% des TVIM localement avancées. Leur conservation dans le processus métastatique renforce l'intérêt de l'évaluation de nouvelles thérapies ciblant le récepteur FGFR3 muté. Enfin, il est possible de rechercher cette mutation sur la tumeur vésicale ou une métastase avec la même pertinence. / Introduction: In muscle invasive urothelial carcinoma of the bladder (MI UCB), few progresses have been made in the last two decades, particularly in chemotherapy, both in localized and metastatic setting. The identification of new biomarkers that can improve the management and prognosis of patients appears essential. Such biomarkers will be clinically relevant if they can predict the response to systemic treatments. In this thesis, we studied biomarkers in two ways: (i) to identify predictive factors of relapse after cisplatin-based adjuvant chemotherapy (AC), (ii) to confirm that the molecular alterations targeted by new therapies and identified in bladder tumor are present in metastases.Objectives:(i) To examine the relationship between relapse after AC and expression of six biomarkers, ERCC1, Emmprin, Survivin, MDR1 p53 and topoisomerase IIα, in bladder tumor and / or lymph node metastases.(ii) To study the heterogeneity of FGFR3 mutations in locally advanced MI UCBMaterials and Methods: A retrospective cohort of 226 patients treated with cystectomy and AC, and tumor collection of representative samples of the primary tumor and lymph node metastases were constituted. The expression of six biomarkers was determined by immunohistochemistry on these samples and determined by morphometry. FGFR3 mutation status was determined by SNaPshot in 84 paired primary tumors and positive lymph nodes.Results: Survivals observed in the cohort were similar to those reported in the literature for patients of same stages, with tumor collection being representative of the MI UCB treated in France. None of the six biomarkers was found as an independent predictive factor of relapse after AC. FGFR3 mutation was found in 4 (4.7%) of the 84 paired samples with complete concordance between primary tumor and lymph node metastases.Conclusions: The expression of the six studied biomarkers by immunohistochemistry is not associated with risk of relapse following AC. FGFR3 mutation status is maintained during metastases process, and FGFR3 activating mutations are present in about 5% of locally advanced MI UCB. FGFR3 mutation remains a promising therapeutic target in a low subset of patients, and the evaluation of new therapies targeting the mutated FGFR3 receptor are needed. Finally, search for these mutations either in bladder tumor or in metastasis is relevant.

Le potentiel invasif et métastatique d'une tumeur est contrôlé par l'angiogenèse via le VEGF (Vascular Endothelial Growth Factor) et les altérations des systèmes impliqués dans l'adhérence cellulaire via les cadhérines. Dans ce travail, nous montrons que le L-165041, agoniste du récepteur nucléaire PPARβ (Peroxisome Proliferator-Activated Receptor) régule l'expression du VEGF dans des cellules dérivées de cancer du col de l'utérus dont le facteur étiologique est un papillomavirus (HPV). Cependant, cet effet est observé uniquement dans des cellules HPV18 indépendamment de l'oncoprotéïne virale E6 et de sa cible p53. De façon intéressante, à la diflérence de ce que nous observons dans des cellules de cancer vésical, le L- 165041 ne contrôle pas la transcription mais stabilise les messagers des isoformes du VEGF (VEGF121, VEGF165, VEGF189) indépendamment du récepteur PPARβ. La stabilisation des ARNm VEGF nécessite la phosphorylation de la p38 MAPK qui conduit, seule ou suite à une cascade de phosphorylations, au déplacement de la protéine HuR du noyau vers le cytoplasme et in fine à la stabilisation des ARNm VEGF. D'autre part, l'agoniste du PPARβ interviendrait dans la« transition épithélio-mésenchymateuse » de cellules urothéliales malignes, puisqu'une augmentation de l'expression de la cadhérine E (marqueur épithélial) et une diminution de l'expression de la cadhérine N (marqueur mésenchymateux) sont observées selon le phénotype cellulaire. Elucider le mécanisme moléculaire d'action des agonistes de PPARβ est d'un intérêt crucial puisque certains ligands sont en cours d'évaluation clinique pour le traitement de maladies métaboliques (dyslipidémies diabète de tvpe II). / Tumor invasive and metastatic potential is controlled by angiogenesis through VEGF (Vascular Endothelial Growth Factor) up-regulation and altered cadherin-mediated cellular adhesion as well. In this work, we show that the L-165041 nuclear receptor PPARβ (Peroxisome Proliferator-Activated Receptor) agonist regulates VEGF expression in cervical cancer derived cells harboring a Human Papilloma Virus (HPV). However, this effect is only observed in HPVl8 positive cells through E6 viral oncoprotein and p53 independent mechanisms. Interestingly, unlike our data obtained in bladder cancer cells, L-165041 does not control VEGF transcription but stabilizes VEGF isoforms mRNA (VEGF121, VEGFl65, VEGFl89) and acts through PPARβ-independent mechanisms. This mRNA stabilization requires p38 MAPK phosphorylation leading to the nuclear/cytoplasmic shuttling ofHuR protein and consequently to the VEGF mRNA stabilization. Furthennore, the PPARβ agonist would be involved in epithelial to mesenchymal transition since it increases E-cadherin expression and decreases N-cadherin one according to the cellular phenotype. To elucidate the molecular mechanism of PPARβ agonists action is crucial because some of ligands are currently in clinical trials for the treatment of metabolic diseases (dyslipidemia, type II diabetes).

L’infection des voies urinaires est l'une des infections bactériennes les plus courantes avec des coûts de soins de santé très élevés. On estime que 50% des femmes connaîtront une infection urinaire au cours de leur vie, ceci de manière récurrente chez la moitié d’entre elles. Le développement de thérapies efficaces a été limité par le manque de connaissance concernant la mise en place de la réponse immune adaptative lors de cette infection. Dans cette étude, nous avons démontré qu'une réponse adaptative est générée lors de l'infection urinaire, cependant celle-ci n’a pas d’action protectrice. Afin de comprendre les mécanismes aboutissant à ce phénomène, nous avons cherché à caractériser les cellules immunitaires présentes dans la vessie. Des tests d’absorption bactérienne ont montré que ces macrophages phagocytent la majorité des bactéries au début de l'infection. Pour évaluer l’influence de ces cellules sur la mise en place de la réponse immune adaptative, nous avons déplété les macrophages et évalué la clairance bactérienne lors d’une deuxième infection. En comparaison avec les animaux non traités, les souris déplétées présentaient une réduction de la charge bactérienne conséquente lors de la seconde infection, cette clairance dépendant de la réponse immune adaptative. Pour comprendre ce mécanisme d'inhibition par les macrophages, nous avons évalué le microenvironnement vésical et la phagocytose au début de l'infection chez les souris déplétées, et chez les souris non traitées. Bien que nous n’ayons pas observé de différences dans la production de cytokines, l'absorption bactérienne par les cellules dendritiques s’avère deux fois plus importante chez les animaux déplétés. Ces données suggèrent que l'absorption bactérienne par les macrophages tissulaires est néfaste pour la mise en place de la réponse adaptative, ouvrant de nouvelles options thérapeutiques. Nous avons également évalué le rôle des lymphocytes T dans ce processus en déplétant ces cellules au cours de l'infection primaire ou avant la deuxième infection. Ainsi, nous avons observé que les lymphocytes T sont nécessaires dans la réponse adaptative, mais ne sont cependant pas indispensables à la clairance bactérienne lors d'une réinfection. De plus, l'infection des souris Batf3-/-, déplétées en cellules dendritiques spécialisées dans la présentation croisée, a montré que ces souris contrôlent une seconde infection aussi bien que les souris contrôle. Ces résultats suggérent que la présence lymphocytes T CD8+ n’est pas nécessaire pour lutter contre l’infection urinaire. Notre étude révèle un mécanisme par lequel le système immunitaire est compromis lors de l'infection urinaire, offrant un point de départ intéressant pour une recherche plus approfondie sur le rôle du système immunitaire adaptatif dans ce contexte, élément fondamental dans le développement de nouvelles thérapies. / Urinary tract infection (UTI) is one of the most common bacterial infections with exorbitant health care costs. It is estimated that 50% of women will experience a UTI during their lifetime and approximately half will suffer recurrent infections. Infected women are treated with antibiotics, however, antibiotic resistance is increasing, raising the need for new therapeutic options. Development of efficient therapies has been impeded by the lack of knowledge of events leading to adaptive immunity. In this study, we demonstrated that an adaptive immune response is generated during UTI, however this response does not confer protective immunity. To begin to understand why the response induced during UTI was not effective, we delineated the immune cell compartment of the bladder and identified macrophages as the most populous immune cell. We evaluated bacterial acquisition in the bladder observing that macrophages phagocytize the majority of the bacteria early in infection. To evaluate the impact of macrophages on the generation of adaptive immunity, we depleted bladder resident macrophages and evaluated bacterial clearance during a challenge infection. Interestingly, mice depleted of resident macrophages, prior to primary infection, exhibited a nearly 2-log reduction in bacterial burden following secondary challenge compared to untreated animals. This improvement in clearance was dependent on the adaptive immune system. To shed light on the mechanism of macrophage inhibition, we evaluated the bladder microenvironment and bacterial acquisition early in infection in macrophage-depleted and control-treated mice. While we did not observe differences in the cytokine microenvironment, bacterial uptake by dendritic cells was increased nearly 2-fold in macrophage-depleted animals. These data suggest that bacterial uptake by tissue macrophages negatively impacts the development of adaptive immunity, revealing a novel target for enhancing host responses to bacterial infection of the bladder. We also evaluated the role of T cells during UTI by depleting these cells during the course of the infection or just prior to challenge infection. We observed that T cells were necessary to mount an adaptive immune response to UTI, however, they were dispensable for bacterial killing during challenge infection. Additionally, infection of Batf3-/- mice, lacking cross-presenting dendritic cells, suggested that CD8+ T cells are dispensable for the response against UTI as these mice cleared a challenge infection as well as wildtype mice. Our study has revealed a mechanism by which the immune system is compromised during UTI, providing an interesting start point for further investigation of the role of the adaptive immune system during UTI, which will be fundamental for the development of new therapies to efficiently treat infection.

Urofacial syndrome (UFS; MIM# 236730) is a rare autosomal recessive condition characterised by urinary bladder and bowel voiding dysfunction with a pathognomonic abnormality of facial movement with expression. UFS can be caused by biallelic putative loss-of-function mutations in HPSE2, which encodes heparanase 2. Failure to discover HPSE2 mutations in all cases of UFS suggests genetic heterogeneity. The urinary tract features of UFS overlap those seen in the spectrum of non-syndromic non-neurogenic voiding dysfunction and vesicoureteric reflux (VUR). This overlap suggests there may be some aspects of pathogenesis in common. The project aimed to define the genotypic and phenotypic spectrum associated with mutations in HPSE2 by Sanger sequencing and multiplex ligation-dependent probe amplification (MPLA) in newly referred cases of UFS and making comparison to a review of mutations and phenotypes seen in the literature. This work discovered five further families with HPSE2 associated UFS increasing known mutations whilst, reinforced that this is an under-recognised condition and emphasised the previously under-reported feature of facial weakness. The failure to discover HPSE2 mutations in all cases referred provided further evidence of genetic heterogeneity. The project also aimed to discover further genes associated with UFS. Autozygosity mapping and whole exome sequencing was carried out in cases of UFS without mutations in HPSE2. This led to the recognition that UFS is also caused by biallelic putative loss-of-function mutations in LRIG2 encoding the leucine-rich repeats and immunoglobulin-like domains 2 (LRIG2) protein in three families. Failure to identify LRIG2 mutations in all HPSE2 negative families suggests further genetic heterogeneity. To address the question of whether the pathogenesis of UFS overlaps more common conditions with a similar spectrum of urinary tract abnormalities I aimed to examine whether pathogenic variants in HPSE2 and LRIG2 were seen in these phenotypes. Unexpectedly this led to the discovering of further families affected by UFS but failed to show an association of variants in UFS genes with non-syndromic urinary tract abnormalities. However, variants of potential interest were discovered. As part of work toward understanding the pathogenesis of UFS and designing a model to test the pathogenesis of sequence variants expression studies in a Xenopus tropicalis hpse2 knock-down model of UFS were carried out. The knock-down model provided valuable insight in to the likely pathogenesis of UFS with evidence pointing towards a congenital peripheral neuropathy with failure of correct nerve path finding. Understanding the pathogenesis of UFS has the potential to direct further research in to therapeutic intervention.

Orientadores: Carlos Arturo Levi D'Ancona, Cássio Luíz Zanettini Riccetto / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T23:13:06Z (GMT). No. of bitstreams: 1 Kos_RodolfoSilva_M.pdf: 2065885 bytes, checksum: 76b4bdc123fa42204d321a899cd184f6 (MD5) Previous issue date: 2015 / Resumo: Mielomeningocele (MMC) é a principal causa de bexiga neurogênica na infância, descrita por disfunção na inervação vésico-esfincteriana e da musculatura perineal, interferindo no funcionamento correto do trato urinário inferior e alterando o processo de micção normal. O tratamento urológico conservador é orientado com base nos resultados de todos os exames relacionados à função vesical e deve-se levar em consideração os sintomas apresentados pelo paciente. A disfunção vesical sem o adequado tratamento pode inibir o indivíduo socialmente e contribuir para influências negativas sobre a qualidade de vida de seus portadores, definida por sensação subjetiva de felicidade e satisfação pessoal do indivíduo. Pela imaturidade da criança com MMC as responsabilidades relacionadas ao tratamento urológico são atribuídas ao cuidador, sendo possível observar aumento da sobrecarga global neste. O objetivo deste trabalho foi avaliar a influência do tratamento urológico conservador na qualidade de vida de crianças com MMC e na sobrecarga de seus cuidadores. Foram estudadas 24 crianças com MMC, divididos em grupos de acordo o tratamento urológico em que a criança estava atualmente de cateterismo intermitente limpo ou uso contínuo de fraldas. Os procedimentos da avaliação contaram com ficha para caracterização da amostra, Teste do Absorvente, Diário Miccional, além dos questionários Burden Interview feito com o cuidador, e AUQUEI e Qualiveen, feitos com a criança. Os resultados obtidos demonstraram que não houve diferença significativa com relação à qualidade de vida da criança quando comparadas entre os grupos de tratamentos urológicos diferentes, entretanto, os cuidadores do grupo que realizava o cateterismo intermitente limpo apresentaram maior sobrecarga global, sendo afetados pelo tratamento urológico com mais ênfase que as próprias crianças / Abstract: Myelomeningocele (MMC) is the leading cause of neurogenic bladder in children, described by dysfunction in vesical-sphincter innervation and the perineal muscles, interfering in the proper function of the lower urinary tract and altering the normal urination process. The conservative urological treatment is targeted based on the results of all tests related to bladder function and should take into account the symptoms presented by the patient. A bladder dysfunction without proper treatment can inhibit the individual socially and contribute to adverse effects on the quality of life of their patients, defined by subjective feeling of happiness and satisfaction of the individual. Because of the immaturity of the children with MMC, the responsibilities related to urological treatment are assigned to the caregiver and are recording increasing global burden on them. The objective of this study was to evaluate the influence of conservative urological treatment on quality of life of children with MMC and overload on their caregivers. 24 children with MMC were studied, divided into groups according to the current urological treatment of clean intermittent catheterization or continuous use of diapers. The evaluation procedures relied on file for sample characterization, Pad Test, Voiding Diary, and the questionnaires Burden Interview done with the caregiver, and AUQUEI and Qualiveen, made with the child. The results showed no significant difference with regard to the quality of a child's life when compared between different urological treatment groups, however, the group of caregivers who performed clean intermittent catheterization showed higher overall burden, being affected by the distinct urological treatment with more emphasis that children themselves / Mestrado / Fisiopatologia Cirúrgica / Mestre em Ciências

Orientadores: Ubirajara Ferreira, Wagner José Fávaro / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-27T12:56:11Z (GMT). No. of bitstreams: 1 Stopiglia_RafaelMamprin_D.pdf: 6417556 bytes, checksum: 52b7e5f09e9aa811c8597b3dbd50aae3 (MD5) Previous issue date: 2015 / Resumo: Em diferentes tipos de cânceres uma população muito pequena de células tem sido reconhecida como células-tronco, por apresentarem capacidade de auto-renovação e diferenciação. Para o seu mapeamento, nos tumores uroteliais, diversos marcadores de superfície celular são utilizados para caracterizar tanto células-tronco normais (CTU) quanto cancerosas (CTC), destacando-se entre eles os antígenos de superfície CD44, CD117 e CD133 e o transportador de membrana do tipo ATP Binding Cassette (ABCG2). Com relação a sinalização dos receptores toll-like (TLRs) no câncer, ainda é uma questão controvertida, pois dados conflitantes os apontam como reguladores negativos ou positivos da carcinogênese. Assim, os objetivos principais do presente estudo foram caracterizar e comparar os perfis das potenciais células-tronco uroteliais e correlacioná-los com a expressão e sinalização dos TLRs 2 e 4 no câncer da bexiga urinária. Para isso, foram selecionados tecidos de bexiga humana sem lesões uroteliais e outras com tumores não-músculo invasivos e músculo invasivos e separadas 30 (trinta) amostras obtidas de homens na faixa etária de 50 a 80 anos (média de 61 anos). Dez dessas amostras de bexiga foram provenientes de necropsia de pacientes sem diagnóstico de lesão urotelial ou doença urológica. As outras 20 amostras vesicais foram obtidas de pacientes submetidos à ressecção transuretral (RTU) e cistectomia radical. Em seguida, divididos em 3 grupos com 10 amostras cada e assim denominados: Grupo Normal, Grupo Câncer Não-Músculo Invasivo e Grupo Câncer Músculo Invasivo de bexiga. E por fim, as amostras foram processadas e submetidas às análises histopatológicas e imunohistoquímicas. Os resultados observados são os seguintes: As imunorreatividades para CD44 e CD133 foram significativamente intensas no grupo câncer músculo invasivo quando relacionadas aos demais grupos. O biomarcador ABCG2 apresentou intensa imunorreatividade nos grupos não-músculo e músculo invasivos, enquanto que no grupo normal a imunorreatividade para esse biomarcador foi ausente. Fraca imunorreatividade para o biomarcador CD117 foi verificada tanto no grupo normal quanto nos grupos não-músculo e músculo invasivos. Assim, as potenciais CTC apresentaram positividade para CD44/CD133/ABCG2 e ocorreram somente nos grupos tumores não-músculo e músculo invasivos. Com relação à caracterização das células-tronco uroteliais normais (CTU), estas foram positivas para CD44/CD133/CD117 e ocorreram com maior frequência no grupo normal em relação aos grupos não-músculo e músculo invasivos. Portanto, os 4 receptores analisados foram reconhecidos como possíveis identificadores de células tronco normais e cancerosas na bexiga. Com relação a análise dos receptores toll-like, as imunorreatividades para TLR2 e TLR4 foram significativamente reduzidas no grupo de pacientes com câncer não-músculo e músculo invasivos, sendo que a imunorreatividade para TLR2 foi ausente neste último. Assim, verificamos que no desenvolvimento dos tumores vesicais a partir de células tronco cancerosas, essas puderam ser identificadas com os marcadores propostos neste estudo e também que a expressão dos receptores TLR 2 e 4 nesta população celular foi mínima ou ausente, atuando, provavelmente, como receptores negativos na carcinogênese urotelial. Como os TLR2 e 4 provavelmente atuaram como reguladores negativos da carcinogênese urotelial pode-se concluir que possivelmente a ocorrência das CTC foi sensível ao decréscimo das imunorreatividades para os TLR2 e TLR4 / Abstract: In different cancers a very small population of cells has been recognized as stem cells , since they have the capacity for self - renewal and differentiation. For their mapping in urothelial tumors, several cell surface markers are used to characterize both normal stem cells ( CTU ) and cancer ( CTC ) , foremost among them the surface antigens CD44 , CD117 and CD133 and membrane transporter ATP Binding Cassette type ( ABCG2 ) . Relationship with the signaling of toll-like receptors ( TLRs ) in cancer is still a controversial issue because conflicting data indicate them as negative or positive regulators of carcinogenesis . Thus, the main objectives of this study were to characterize and compare the profiles of urothelial stem cells and correlates them with the expression and signaling of TLRs in cancer of the urinary bladder . Tissues of human bladder urothelial lesions and no other separate tumors with invasive and invasive non- muscle and muscle were selected thirty (30 ) samples of men aged 50-80 years (mean 61 years). Ten of these samples were derived from autopsy bladder of patients without a diagnosis of urothelial injury or urologic disease. The other 20 bladder samples were obtained from patients undergoing transurethral resection (TURP ) and radical cystectomy. After this, divided into 3 groups with 10 samples each and named as follows: Normal Group , Group Non- Muscle Invasive Cancer Group and Muscle Invasive Bladder Cancer . Then, the samples were processed and subjected to histological and immunohistochemical analysis. The observed results are as follows: The imunorreatividades for CD44 and CD133 were significantly intense in group Muscle Invasive Cancer as related to the other groups. The ABCG2 biomarker groups showed intense immunoreactivity in Non- Muscle Invasive and muscle, while in the Normal group immunoreactivity was absent for that biomarker . Weak immunoreactivity for CD117 biomarker was observed both in the Normal group and the groups Non-Muscle Invasive and Muscle. Thus, CTC were positive for CD44/CD133/ABCG2 and occurred only in tumors groups Non- Muscle Invasive and Muscle. Regarding the characterization of normal urothelial stem cells ( CTU ), these were positive for CD44/CD133/CD117 and occurred more frequently in the Normal group in relation to groups and Non-Muscle Invasive Muscle. Therefore, the four receptors were analyzed for possible identifiers recognized as normal stem cells and bladder cancer . Regarding the analysis of toll-like receptors, TLR2 and TLR4 to imunorreatividades were significantly reduced in patients with Cancer and Non- Muscle Invasive Muscle, and the TLR2 immunoreactivity was absent in the latter . Thus, we see that the development of bladder tumors from stem cells cancerous, these cells could be identified with markers and also proposed that the expression of TLR receptors 2 e 4 this cell population was minimal or absent, acting probably as receptors negative in urothelial carcinogenesis. As TLR2 and 4 probably acted as negative regulators of urothelial carcinogenesis can be concluded that most likely the occurrence of the CTC was sensitive to the decrease in imunorreactivities for TLR2 and TLR4 / Doutorado / Fisiopatologia Cirúrgica / Doutor em Ciências

Harnblase. Weiterhin bestärkt die beobachtete Kopplung die Hypothese, dass das Myofibroblastennetzwerk afferente Signale Verstärken und modulieren kann. Die Verbesserung des Verständnisses der Harnblasenfunktion ist von grundlegendem Interesse für die Behandlung von Miktionsstörungen. Eine Störung von mechanischer Stimulierbarkeit und Kopplung der suburothelialen Myofibroblasten kann eine Störung der Sensorik der Harnblase plausibel erklären und stellt einen neuen Therapieansatz dar.:1 Abkürzungsverzeichnis 2 Einführung 2.1 Hintergrund 2.2 Anatomische und physiologische Grundlagen der Harnblasenfunktion 2.2.1 Aufbau der Harnblasenwand 2.2.2 Terminologie und Klassifikation interstitieller Zellen der Harnblase 2.2.3 Phänotypisierung der suburothelialen Myofibroblasten 2.2.4 Funktionsweise der Myofibroblasten 2.2.5 Innervation des unteren Harntraktes und neuronale Kontrolle der Harnblasenfunktion 2.2.6 Lokale sensorische Netzwerke der Harnblase 3 Aufgabenstellung 4 Material und Methoden 4.1 Gewinnung und Zellkultur suburothelialer Myofibroblasten 4.2 Lösungen und Chemikalien 4.3 Calcium imaging 4.4 Datenanalyse 4.4.1 Erzeugung der FI-Ratio Datensätze 4.4.2 Automatische Fluoreszenz-Signal-Analyse 4.5 Aufbau und Anordnung der Experimente 4.5.1 Spontanaktivität 4.5.2 Mechanische Stimulation mittels Glasmikropipette 4.5.3 Mechanische Stimulation durch Scherstress 4.5.4 Hypoosmolare Stimulation 5 Ergebnisse 5.1 Spontane Kalziumaktivität humaner suburothelialer Myofibroblasten 5.2 Mechanische Stimulierbarkeit durch Druck mittels Glasmikropipette 5.2.1 Intrazelluläre Ausbreitung mechanisch induzierter Kalziumsignale 5.2.2 Interzelluläre Ausbreitung mechanisch induzierter Kalziumsignale in kultivierten humanen suburothelialen Myofibroblasten 5.3 Mechanische Stimulierbarkeit durch Scherstress 5.4 Mechanische Stimulierbarkeit durch osmotischen Stress 6 Diskussion 6.1 Beobachtete Kalziumsignale suburothelialer Myofibroblasten 6.2 Bedeutung der mechanischen Stimulierbarkeit 6.3 Identifikation der stretch-activated channels 6.4 Bedeutung der interzellulären Kopplung 6.5 Konzept zur Rolle der suburothelialen Myofibroblasten 6.6 Methodischer und experimenteller Ansatz 6.6.1 Selbst entwickelte Fluoresence Analysis Software 6.6.2 Methoden der mechanischen Stimulierbarkeit 6.7 Pathophysiologische Aspekte 7 Zusammenfassung der Arbeit 8 Literaturverzeichnis 9 Anlagen 9.1 Selbstständigkeitserklärung 9.2 Lebenslauf 9.3 Publikationen 9.4 Danksagung

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No description available.

Because the urinary bladder stores and releases urine, its normal function includes filling and emptying, accompanied by distension and relaxation. It is known that chronic distension compromises blood flow. Recent studies of the rabbit bladder vasculature have described specializations of that vasculature that appear to enhance blood flow in the bladder wall during distension. The present report describes the location, orientation, and structure of an elastic sheath surrounding the vesicular arteries, which may represent one of these specializations. The location, vasculature, and structure of an accessory elastic sheath surrounding the vesicular arteries of the rabbit bladder is described using light and electron microscopy, India ink injection, and vascular corrosion casting. The common iliac arteries of rabbits were cannulated to permit perfusion of the distal vasculature including the urinary bladder. After the bladder vasculature was visually cleared of blood by perfusion with buffered saline, one of the following procedures was used: 1) for light or electron microscopy, the bladder was perfuse-fixed with buffered 2% glutaraldehyde; 2) the bladder vasculature was filled with India ink for vessel tracing; or 3) corrosion casts of the bladder vasculature were prepared by infusion of a Mercox resin mixture. Casts, cleaned of tissue with KOH, and water and formic acid rinses, are dried, and mounted for routine scanning electron microscopy. The presence of an accessory sheath surrounding the main vesicular arteries and some of their branches in the basal two thirds of the urinary bladder was observed on India ink injected specimens and confirmed by micrographs and vascular corrosion casts. The sheath consists of elastic and collagenous fibers and is separated from the tunica media of the arteries by a loose connective tissue layer of variable width. The sheath is circumscribed by a layer of fine blood vessels. The vesicular arteries undulate within the sheath to an extent which is dependent upon the degree of distension of the bladder. This sheath likely represents a specialization which permits the bladder vasculature to accommodate expansion and contraction of the wall during normal filling and emptying. Undulations or coiling of the vesicular arteries within the loose connective tissue core of the sheath increase with bladder contraction, and apparently the sheath simply holds the artery in position during such coiling. The sheath, may represent a modification of the external elastic lamina found in some muscular arteries.