Le cancer de l'ovaire représente la première cause de décès par cancer gynécologique. La survie globale des patientes à 5 ans est inférieure à 30%. Ce sombre pronostic s'explique à la fois par la découverte tardive de la maladie et par le développement d'une chimiorésistance. L'ascite est un fluide exsudatif qui est fréquemment accumulé dans la cavité péritonéale au cours de la progression des cancers de l'ovaire. Ce « microenvironnement tumoral » particulier contribue à la dissémination des cellules cancéreuses et à leurs implantations péritonéales.L'objectif global du travail de thèse a été, d'une part d'évaluer l'influence de l'ascite sur le comportement des cellules cancéreuses ovariennes et d'autre part, d'étudier les mécanismes de résistance à la perte d'ancrage des cellules cancéreuses ovariennes.Nous avons ainsi démontré que l'ascite induit une transition épithélio-mésenchymateuse partielle et que les modifications des comportements cellulaires observées sont dépendantes des intégrines alpha-v.Deux ligands de ces intégrines, la vitronectine et la fibronectine, ont été purifiés selon un protocole original permettant la caractérisation des deux protéines à partir d'une même ascite. Ces protéines ascitiques ont des propriétés différentes selon leur origine, donc selon les patientes dont elles sont issues, et influencent le comportement adhésif des cellules avec un degré variable. L'importance de la signalisation dépendante des intégrines alpha-v et des voies MAP Kinases a également été démontrée dans l'établissement d'une résistance des sphéroïdes tumoraux à l'anoïkis.En perspective, approfondir les connaissances des processus cellulaires et moléculaires conduisant à la dissémination intrapéritonéale et à l'émergence de chimiorésistance ainsi que déterminer le rôle potentiel de protéines ascitiques dans ces processus pourraient permettre la découverte de nouvelles cibles thérapeutiques. / Ovarian cancer is the most lethal gynaecological malignant disease, mainly due to late diagnosis and to acquired chemoresistance. An exudative fluid named ascites is frequently accumulates within the abdominal cavity during ovarian cancer progression. This unique tumor microenvironment contributes to cancer cell dissemination and peritoneal metastasis.The aim of the study was to evaluate the influence of ascites on cancer cell behaviors and to better understand the mechanisms of ovarian cancer cell resistance to the loss of anchorage.We demonstrate that ascites induces a partial epithelial–mesenchymal transition and that the modifications of cell behaviors observed are dependent of alpha-v integrins. A combined purification protocol has been established in order to purify vitronectin and Fibronectin, both ligands of these integrins, from a single pathological sample. These purified ascitic proteins have different molecular features according to the patients and impact the adhesive cell behavior with various degrees.Moreover we showed the importance of alpha-v integrins and MAP Kinases signalling pathways in the anoikis resistance of ovarian cancer spheroids.Our prospects are i) to increase the knowledge of the cellular and molecular processes leading to the intraperitoneal dissemination and to the emergence of chemoresistance and also ii) to determine the potential role of ascitics proteins in these processes. We will expect that these investigations couldlead to the discovery of new therapeutic targets.

Thèse d'exercice : Médecine. Médecine générale : Paris 12 : 2007. / Titre provenant de l'écran-titre. 67 f. ill. Bibliogr. f. 57-66.

Thesis (M.S.)--West Virginia University, 2003. / Title from document title page. Document formatted into pages; contains vii, 88 p. : ill. (some col.). Includes abstract. Includes bibliographical references.

Orientador: Denise Fecchio / Resumo: O presente trabalho objetivou avaliar o envolvimento das prostaglandinas no crescimento tumoral, influxo inflamatório e secreação de citocinas durante a evolução do Tumor Ascítico de Ehrlich (TAE). Para tanto, camundongos foram inoculados com 1 x 103 células tumorais (ip) e tratados com indometacina (1mg/Kg,1x/dia,ip) ou com diluente (0,1 ml,1x/dia,ip). Decorridos 1, 3, 6, 10 e 13 dias os animais foram sacrificados e avaliados quanto ao influxo inflamatório diferencial, secreção de TNF-a, IL1-a, IL-2, IL- 4, IL-6, IL-10 e IL-13 e níveis de PGE2 no lavado peritoneal.. Dois grupos controle adicionais foram constituídos de animais não portadores de TAE tratados com indometacina ou diluente, seguindo o mesmo protocolo. Os resultados obtidos demonstraram que o implante do TAE induz produção de PGE2 durante toda sua evolução; aumento do número de células neoplásicas a partir do 10o dia e diminuição do influxo de células mesoteliais no 10º dia e de basófilos no 10º e 13º dia pós implante neoplásico. Em relação as citocinas o TAE induziu produção de IL-6 no 10º e 13º dia e de IL 2 no 13º dia, não alterando de modo significativo o perfil das outras citocinas estudadas. O tratamento de animais portadores de TAE com indometacina, foi eficaz em inibir o crescimento tumoral e a síntese de PGE2 a partir do 10o dia de crescimento neopásico, e promoveu aumento significativo no influxo de neutrófilos segmentados e de células nucleadas, apenas em tempos iniciais da evolução tumoral. Ainda, o tratamento com indometacina promoveu síntese de IL-13 e inibição significativa de IL-6 no 13o dia de crescimento tumoral, não alterando as outras citocinas analisadas. No grupo não portador de tumor tratado com indometacina observamos aumento no influxo de neutrófilos segmentados no 1º dia... (Resumo completo, clicar acesso eletrônico abaixo). / Abstract: The aim of the present study was investigate the prostaglandin involvement during the growth of Ehrlich Ascites Tumor (EAT), using as parameters: tumoral growth, inflammatory influx and cytokine profile. Mice were inoculated with 1 x 103 tumor cells (ip) and treated with indomehacin (1mg/Kg,1x/day,ip) or diluent (0,1ml,1x/day,ip). After 1, 3, 6, 10 and 13 days the animals were sacrificed and evaluated in relation to inflammatory influx, secretion of TNF -a , IL1-a, IL-2, IL-4, IL-6, IL-10 and IL-13 and PGE2 level, in peritoneal cavity. Two groups no bearing EAT were treated with indomethacina or diluent as control ,following the same protocol. The results demonstrated that EAT implant induces PGE2 production during all evolution; increases tumoral cells number from the 10th day and decreases the mesotelial cells on 10th day and basophils cells on 10th and 13rd day.The cytokine profile showed EAT induces production of IL 6 from 10th day and of IL 2 on 13rd day, the other studied cytokines were not affected in a significant way. The indomethacin treatment of EAT bearing mice inhibited the tumoral growth and PGE2 synthesis from the 10th day and promoted significant increase on the neutrophils influx and total inflammatory cells, just in initial times of the tumoral evolution. Indomethacin treatment also promoted IL-13 synthesis and significant inhibition of IL-6 on 13rd day of EAT growth, but did not altered the others cytokines. The indomethacin treatment of animals do not bearing EAT increases neutrophils influx on the 1st day, lymphocytes on the 3rd day, eosinophils on 10th day; and no detected alteration was detected on cytokine profile Taken together, the results suggest that EAT growth is modulate by PGE2 and the inhibition of the tumoral growth could be partly related with suppression of IL-6 and liberation of IL-13. / Mestre

Ovarian cancer is the fifth leading cause of all cancer deaths in women and the most lethal gynecologic cancer in the United States. During metastasis, cancer cells exfoliate from the primary tumor and aggregate to form spheroids, enhancing their survival within the peritoneal cavity during dissemination to a secondary outgrowth site. The inability of removal of these aggregates by traditional surgical interventions may contribute to the high recurrence and mortality rate of ovarian cancer diagnosed at late stages. Obesity, particularly abdominal obesity, has been shown to increase ovarian cancer risk and decrease survival. The recruitment of stromal vascular fraction (SVF) present in adipose tissue represents a growth and proliferation advantage to ovarian tumors, and endogenous sphingolipids like sphingosine-1-phosphate are increased in ovarian cancer patients. These conditions, combined with the physiological conditions within malignant ascites (hypoxia and low glucose), represent a physiological environment that can impact the metabolic responses of ovarian cancer spheroids. Here, we investigated the metabolic adaptations of serous ovarian cancer cells across the metastatic cycle and in conditions that mimic those of the peritoneal cavity and malignant ascites. We first investigated the different in metabolic responses between adherent monolayers and 3D spheroids. We confirmed that spheroids have a reduced metabolic rate and drug response that is affected by the incorporation of obese SVF into aggregates. To investigate these changes in the next stages of the metastatic cycle, we used time trials to observe how adherence of spheroids to a secondary site changes metabolic response and substrate utilization in physiological conditions. Adhesion of spheroids showed changes in energy metabolism and substrate utilization, switching from mainly glutamine oxidation to glucose oxidation that could support successful outgrowth. Spheroids also were resilient to culture conditions, even non-permissive conditions such as those found in the peritoneal cavity. Finally, we utilized human malignant ascites from ovarian cancer patients as a further investigation into conditions that imitate in vivo characteristics that could affect spheroid metabolism. Exposure to malignant ascites reduced spheroid viability as well as basal respiration and ATP synthesis. However, spare respiratory capacity was increased, and human spheroids changed their substrate utilization in response to ascites. Taken together, these studies provide an identification of metabolic switches across different stages of ovarian cancer metastasis that contribute to their survival, which represents an emerging target for prevention and treatment for individuals with ovarian cancer. / Doctor of Philosophy / Ovarian cancer is the deadliest reproductive cancer in women, and most women who are diagnosed will die from the disease because of its high recurrence rate and because it is typically detected at late stages. When ovarian cancer metastasizes, cells or cell clusters from the original tumor aggregate together to form balls of cells called spheroids, which move through the abdominal cavity to other sites to grow additional tumors. These spheroids are thought to contribute to recurrence of this cancer, since they cannot be removed by surgery. As these spheroids move through the abdominal cavity, they are exposed to an environment that has a low amount of oxygen and glucose. These spheroids may also be exposed to bioactive lipids and cells from the adipose tissue called stromal vascular fraction, both of which are related to obesity and may help cancer spheroids survive. The survival of these spheroids is in part related to how their metabolism functions, which may help them make energy and the building blocks needed to continue growing and form successful secondary tumors. Identifying how these spheroids change their metabolism at different points during the disease may help identify points that can be targeted to prevent changes in metabolism that could support their growth. This dissertation identified metabolic changes that occur in spheroids, in conditions that are similar to those spheroids would be exposed to in an abdominal cavity. First, we compared single layers of cells to spheroids and found that spheroids had a lower metabolic rate and lower drug response, which may help them survive in the abdominal cavity. Next, we allowed the spheroids to lay down and grow out, like they would when they found a new location during metastasis, to see how their metabolism changed and what substances they used to make energy in conditions that mimicked the abdominal cavity. As spheroids adhered, they changed their energy metabolism and switched the substances they used to make energy, all while continuing to survive and grow out even in conditions that were not supportive. These switches could help them grow out and successfully metastasize. Lastly, we used ascites fluid from human ovarian cancer patients and treated spheroids with this to see how their metabolism changed in response. While some aspects of metabolism and survival was reduced, their ability to increase their metabolism when stressed increased and human spheroids used nutrients to make energy differently. Overall, we show that across the stages of metastasis, ovarian cancer spheroids can change their metabolism in response to their environment. Identifying these metabolic switches helps us understand how successful metastasis happens, and can inform future targets to slow or prevent metastasis, prolonging the life of women who have been diagnosed with ovarian cancer.

Vid cytologisk bedömning av celler i exsudat kan olika metoder användas vid preparering. Vid konventionell utstryksmetod stryks provet ut manuellt på objektglas medan vätskebaserad ThinPrepmetod preparerar provet i en automatiserad process. Studiens syfte var att jämföra konventionell utstryksmetod och vätskebaserad ThinPrepmetod med tillsats av ättiksyra vid preparering av etanolfixerade exsudat. I studien användes totalt 34 prover av pleuravätska och ascites. Proverna preparerades med konventionell utstryksmetod och två varianter av ThinPrepmetod, ThinPrepmetod 1 och ThinPrepmetod 2. Preparaten bedömdes av två cytodiagnostiker utifrån fem kriterier; cellförekomst, förekomst av inflammatoriska celler, cellmorfologi, bakgrundsmaterial och förekomst av atypiska eller maligna celler. Resultaten visade på att ThinPrepmetod 2 gav likvärdiga, och i vissa avseenden bättre, resultat än konventionell utstryksmetod. ThinPrepmetoden har generellt fler fördelar jämfört med konventionell utstryksmetod, framförallt innebär den en tidsbesparing för de som bedömer preparaten vilket också gynnar patienter. En mer omfattande studie rekommenderas för att konfirmera resultaten i denna studie. / Preparation of effusions for cytological evaluation can be performed with different methods. Conventional smears are performed by spreading the sample directly onto a slide while liquid-based ThinPrep method prepares the sample in an automated process. The aim of this study was to compare conventional smears with liquid-based ThinPrep preparation, with additional acetic acid, of effusions fixated with ethanol. A total of 34 samples of pleural effusions and ascites were included. The samples were prepared with conventional smears and two versions of the ThinPrep method, ThinPrep method 1 and ThinPrep method 2. The preparations were examined by two cytotechnologists based on five criteria; occurrence of cells, occurrence of inflammatory cells, cell morphology, background material and the presence of atypical or malignant cells. The results showed that ThinPrep method 2 obtained equivalent and, in some aspects, even better results compared to conventional smears. In general, the ThinPrep method has several advantages compared to the conventional smears. In particular, it is timesaving when examining the slides, which also benefits the patients. A more comprehensive study is recommended to confirm the results of this study.

Cirrhosis is characterized by the progressive development of portal hypertension. Portal hypertension, associated with sodium retention, leads to the formation of ascites. Ascites may be difficult to treat and, thereby, become "refractory". To date, the mortality of patients with refractory ascites remains high. In order to offer patients adequate treatment, it is important to identify complications that may develop and the factors of good and poor prognosis affecting their survival. One can retain five specific complications of cirrhosis with ascites: hepatorenal syndrome, spontaneous bacterial peritonitis, paracentesis induced circulatory dysfunction, hepatic hydrothorax and dilutional hyponatremia. This thesis reported an update of pathogenesis, complications and treatment of refractory ascites. Our original clinical research has, meanwhile, focused on the identification of different prognostic factors in refractory ascites in direct relation to these five complications. We have focused on the effect of the administration of beta blockers in patients with refractory ascites: this treatment is widely prescribed for prevention of gastrointestinal bleeding. It is associated in these patients with higher mortality and a high incidence of paracentesis induced circulatory dysfunction. We specified that the severe hyponatremia, leading to withhold diuretic therapy, is a more accurate prognostic factor than the MELDNa score in refractory ascites. With regard to the hepatorenal syndrome of slow onset (type 2), we emphasized the high frequency of this syndrome in refractory ascites. We showed that there was an association between the level of portal hypertension and the incidence of spontaneous bacterial peritonitis. Furthermore, we demonstrated that the presence of bacterial DNA in ascites of outpatients suffering from refractory ascites was particularly low. The detection of this DNA is not a precursor to infection of ascites. In conclusion, with a systematic analysis of various prognostic factors related to complications of ascites in cirrhotic patients, this work gives a better understanding of their chances of survival. This will allow a more adequate stratification in the decision trees for a cure of the disease such as liver transplantation. / Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

No description available.

En vanlig orsak till otillfredsställande cytologiska preparat är kontamination av blod. På Cytologilaboratoriet, Länssjukhuset Ryhov tvättas blodiga gynekologiska preparat i Cytolyt® solution och ättiksyra för att reducera kontaminationer. Syftet med studien var att studera den potentiella påverkan tvätt med Cytolyt® solution och ättiksyra har på mesotelcellers morfologi. I studien ingick 60 exsudat av pleuravätska och ascites som analyserades med konventionell cytologisk metod. Utstryk utfördes och analyserades utan tvätt (kontroller) och efter tvätt i olika tider varpå preparaten bedömdes med avseende på mesotelcellers morfologi. Av kontrollerna hade 53% en välbevarad cytoplasma men endast 5.8% av de tvättade proverna. Cirka 87% av kontrollerna och 46% av de tvättade proverna hade en välbevarad och distinkt cellkärna. Morfologiska förändringar hos mesotelceller kunde påvisas efter tvätt i Cytolyt® solution och ättiksyra men förändringarna kunde inte kopplas till eller misstolkas som patologiska förändringar. / A common reason for unsatisfactory cytological smears is contamination of blood. Blood contaminated gynaecological samples are washed with Cytolyt® solution and acetic acid at the Cytological Laboratory, County Hospital Ryhov to reduce contaminations. The objective of this study was to determine the potential effect of Cytolyt® solution and acetic acid on the morphology of mesothelial cells. In this study 60 samples of pleural effusions and ascites was analysed with conventional cytological method. Cytological smears were prepared without wash (controls) and with wash for different durations. The morphology of mesothelial cells in the samples were then examined. Out of the controls 53% had a wellpreserved cytoplasm while this distribution decreased to 5.8% amongst the washed smears. About 87% of the controls and 46% of the washed smears had a preserved and distinct nucleus. Changes in morphology could be seen in mesothelial cells after wash with Cytolyt® solution and acetic acid. However, the changes could not be associated with disease. Washing exudates with Cytolyt® solution and acetic acid can alter morphological changes in mesothelial cells although the changes seen cannot be mistaken for pathological changes.

Le syndrome néphrotique (SN) est causé par une altération glomérulaire, responsable d’une excrétion urinaire anormale de protéines plasmatiques, compliquée d’hypoalbuminémie. Le SN est toujours associé à une rétention rénale de Na + qui conduit à la génération d'ascite et / ou d'œdèmes. La pathogénie de la rétention de Na + et de la constitution d’œdèmes n’est pas entièrement élucidée. Dans notre étude, nous avons évalué le rôle possible des espèces réactives de l'oxygène (ROS) dans cette pathogénie. Notre étude dans le modèle de rat aminonucléoside puromycine (PAN) de SN fournit des éléments de preuve d'un rôle critique des ROS dans les troubles hydro-électrolytiques associés au SN. Dans le rein, l'endocytose de l'albumine anormalement filtrée dans le néphron distal induit un stress oxydatif qui est responsable de l’augmentation de la Na, K-ATPase. Dans le péritoine, le SN est associé à une augmentation marquée de la perméabilité à l'eau et à une diminution du coefficient de réflexion des protéines de la barrière péritonéale. Ces modifications, déclenchées par le stress oxydatif et l'activation subséquente de NF-kB, comptent pour environ deux tiers du volume de l'ascite. Enfin, nous avons confirmé que le stress oxydatif participe à la sécrétion de l'angiotensine-aldostérone et est nécessaire à l’apparition de l'hyperaldostéronémie observée chez les rats PAN-néphrotiques. / Nephrotic Syndrome (NS) is a nonspecific kidney disorder defined by abnormal urinary excretion of plasma proteins and hypoalbuminemia. NS is always associated with a renal retention of Na+ leading to the generation of ascites and/or edema. The pathogenesis of Na+ retention and edema is not fully elucidated. In our studies we evaluated the possible role of reactive oxygen species (ROS) in this pathogenesis. Our studies in the puromycin aminonucleoside (PAN) rat model of NS provided pieces of evidence for a critical role of ROS in the hydro-electrolytic disorders associated with NS. In the kidney, endocytosis of abnormally filtered albumin in the distal nephron induces an oxidative stress which is responsible for the up-regulation of Na,K-ATPase. In the peritoneum, NS is associated with a marked increase in water permeability and a decrease in the reflection coefficient to proteins of the peritoneal barrier. These changes, which are triggered by oxidative stress and subsequent activation of NF-kB, account for approximately two-third of the volume of ascites. Finally, we confirmed that oxidative stress participates in the angiotensin-stimulated secretion of aldosterone and is required for the hyperaldosteronemia observed in PAN-nephrotic rats.