Dissertations / Theses on the topic 'Endometrium'

Lynch syndrome (LS), an autosomal dominant inherited cancer susceptibility syndrome, also known as hereditary non-polyposis colon cancer (HNPCC), is caused by a germline mutation in one of several DNA mismatch repair (MMR) genes. LS is the most common presentation of hereditary colorectal cancer (CRC), accounting for about 2-5% of all CRC cases. More recently, it is found that a similar number of endometrial cancers is also due to one of the MMR gene mutations. There has been significant progress in LS-related CRC in terms of molecular pathogenesis, risks, genetic basis, and cancer prevention. In contrast, the advance about LS-related endometrial cancer (EC) is very much limited. In this commentary, we summarize the main clinicopathologic features of LS-related EC and propose universal screening for LS in individuals with endometrial cancer.

El càncer d’endometri és la neoplàsia ginecològica més freqüent en els països desenvolupats. Malgrat que la majoria dels carcinomes són curables amb un tractament adequat, al voltant del 20% dels tumors es comporten de forma agressiva i suposen un repte terapèutic. És per aquest motiu que sorgeix la necessitat d’identificar nous paràmetres per tal de seleccionar pacients amb risc de recidiva o metàstasi. La cèl•lula eucariota presenta canvis en el seu metabolisme com a resposta coordinada a diferents situacions fisiològiques i patològiques, entre elles el càncer. L’anàlisi del metaboloma, mitjançant la metabolòmica, pot permetre identificar metabòlits diferencials que representen el producte final de les vies de senyalització que estan alterades en el càncer. Per aquest motiu hem volgut realitzar una anàlisi metabolòmica del càncer d’endometri. A més a més, per tal de validar els resultats i traslladar-los a la pràctica clínica s’han avaluat els nivells d’expressió dels metabòlits diferencials més significatius en arrays de teixit (TMAs). En primer lloc, els resultats han mostrat que el procés de carcinogènesi del càncer d’endometri defineix un perfil metabolòmic específic. Els resultats suggereixen que la via dels endocannabinoides pot estar implicada en la gènesi i progressió del carcinoma endometrioide. Així mateix, l’alteració del metabolisme de les purines pot estar implicada en fenòmens d’invasió miometrial en el càncer d’endometri. En segon lloc, l’estudi metabolòmic ha mostrat un perfil diferencial entre carcinomes endometrioides i serosos. A més a més, ha permès identificar dues molècules, ADI1 i BCAT1, que poden estar implicades en la gènesi de les neoplàsies endometrioides i en la progressió tumoral. Així mateix, aquests compostos poden ajudar a establir el diagnòstic diferencial entre aquests dos subtipus histològics amb pronòstics molt diferents. Finalment, partint de la premissa que l’angiogènesi és un mecanisme essencial per al creixement, invasió i disseminació tumoral i que els carcinomes d’endometri amb flux sanguini intratumoral disminuït tenen pitjor pronòstic, s’ha analitzat el perfil metabolòmic del càncer d’endometri en funció del seu flux sanguini. El resultats mostren un perfil metabolòmic específic dels tumors segons el seu flux sanguini i permeten identificar Resolvina D i fosfolípids específics diferencials entre tumors d’alt i baix flux sanguini. Aquestes molècules poden estar implicades en l’angiogènesi i progressió tumoral en el càncer d’endometri.
El cáncer de endometrio es la neoplasia ginecológica más frecuente en los países desarrollados. A pesar de que la mayoría de los carcinomas son curables con un tratamiento adecuado, alrededor del 20% de los tumores se comportan de forma agresiva y suponen un reto terapéutico. Por este motivo surge la necesidad de identificar nuevos parámetros que permitan seleccionar pacientes con riesgo de recidiva o metástasis. La célula eucariota presenta cambios en su metabolismo como respuesta coordinada a diferentes situaciones fisiológicas y patológicas, entre ellas el cáncer. El análisis del metaboloma, mediante la metabolómica, puede ayudar a identificar metabolitos diferenciales que representan el producto final de las vías de señalización que están alteradas en el cáncer. Por esta razón hemos querido realizar un análisis metabolómico del cáncer de endometrio. Además, para la validación de los resultados y su translación a la práctica clínica se han evaluado los niveles de expresión de los metabolitos diferenciales más significativos en arrays de tejido (TMAs). En primer lugar, los resultados han mostrado que el proceso de carcinogénesis del cáncer de endometrio define un perfil metabolómico específico. Los resultados sugieren que la vía de los endocannabinoides puede estar implicada en la génesis y progresión del carcinoma endometrioide. Además, la alteración del metabolismo de las purinas puede estar implicada en fenómenos de invasión miometrial en el cáncer de endometrio. En segundo lugar, el estudio metabolómico ha mostrado un perfil diferencial entre carcinomas endometrioides y serosos. Además ha permitido identificar dos moléculas, ADI1 i BCAT1, que pueden estar implicadas en la génesis de las neoplasias endometrioides así como en la progresión tumoral. Asimismo, estos dos compuestos pueden ser útiles en el diagnóstico diferencial de estos dos subtipos histológicos con pronósticos tan distintos. Finalmente, partiendo de la base que la angiogénesis es un mecanismo esencial para el crecimiento, invasión y diseminación tumoral y que los carcinomas de endometrio con flujo sanguíneo intratumoral disminuido tienen peor pronóstico, se ha analizado el perfil metabolómico del cáncer de endometrio en función de su flujo sanguíneo. Los resultados muestran un perfil metabolómico específico de los tumores según su flujo sanguíneo y permiten identificar Resolvina D i fosfolípidos específicos diferenciales entre tumores de alto y bajo flujo sanguíneo. Estas moléculas pueden estar implicadas en la angiogénesis y progresión tumoral en el cáncer de endometrio.
Endometrial cancer is the most frequent gynecological malignancy in developed countries. Although most carcinomas are curable with adequate treatment, about 20% of tumors behave aggressively and pose a therapeutic challenge. For this reason, there is a need to identify new parameters that allow the selection of a patient with risk of recurrence or metastasis. The eukaryotic cell presents changes in its metabolism as a coordinated response to different physiological and pathological situations, including cancer. The analysis of the metabolome, through metabolomics, can help identify differential metabolites that represent the final product of the signaling pathways that are altered in cancer. For this reason we wanted to perform a metabolomic analysis of endometrial cancer. In addition, for the validation of the results and their translation to clinical practice, the expression levels of the most significant differential metabolites have been evaluated using tissue arrays (TMAs). First, the results have shown that the process of carcinogenesis of endometrial cancer defines a specific metabolomic profile. The results suggest that the endocannabinoid pathway may be involved in the genesis and progression of endometrioid carcinoma. In addition, the alteration of the purine metabolism may be involved in myometrial invasion phenomens in endometrial cancer. Second, the metabolomic study has shown a differential profile between endometrioid and serous carcinomas and has allowed the identification of two molecules, ADI1 and BCAT1, which may be involved in the genesis of endometrioid neoplasms as well as in tumor progression. Likewise, these two compounds can be useful in the differential diagnosis of these two histological subtypes with such different prognoses. Finally, starting from the basis that angiogenesis is an essential mechanism for tumor growth, invasion and dissemination and that endometrial carcinomas with decreased intratumoral blood flow have a worse prognosis, the metabolomic profile of endometrial carcinoma has been analysed according to its blood flow. The results show a specific metabolomic profile of the tumors according to their blood flow and allow identifying Resolvin D and specific phospholipids differentials between high and low blood flow tumors. These molecules may be involved in angiogenesis and tumor progression in endometrial cancer.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Pólipos endometriais são neoformações resultantes da hipertrofia focal da camada basal do endométrio associada a um hiperestímulo hormonal. Sua etiopatogenia ainda não está bem estabelecida, não havendo consenso sobre sua história natural, seu real significado como entidade patológica e sua relação com a neoplasia endometrial. Os pólipos endometriais são a principal indicação de histeroscopia cirúrgica, sem que haja, no entanto, protocolo estabelecido para seu melhor manejo. Apresentar um panorama sobre esta condição pode auxiliar na busca em traçar indicadores para seu tratamento adequado. Avaliar os dados histeroscópicos e o perfil imuno-histoquímico dos pólipos endometriais de mulheres acompanhadas no Setor de Endoscopia Ginecológica da Faculdade de Medicina de Botucatu - UNESP. Trata-se de estudo clínico de avaliação prospectiva e transversal com amostra de conveniência de 82 mulheres, submetidas à polipectomia histeroscópica cirúrgica, e 20 mulheres, submetidas à cirurgia oncológica devido à neoplasia endometrial, entre o período de janeiro de 2010 a dezembro de 2011. Foram analisados parâmetros clínicos, ultrassonográficos, histeroscópicos, histopatológicos e imuno-histoquímicos dos pólipos endometriais e do câncer de endométrio. Para estudo estatístico das variáveis quantitativas considerou-se o teste t de Student para amostras independentes, e o teste não paramétrico de Mann-Whitney. Para as variáveis qualitativas utilizou-se o teste de Goodman para contrastes entre e dentro de populações multinomiais, tendo sido adotado o nível de 5% de significância. Na comparação entre biópsias endometriais ambulatoriais, orientada e dirigida, a sensibilidade da biópsia dirigida no diagnóstico dos pólipos endometriais variou de 35,3% a 36,8%, quando realizada no ápice e na base da lesão...
Endometrial polyps are focal neoformation resulting from hypertrophy of the basal layer of the endometrium associated with a hormonal hyperstimulation. Its pathogenesis is not well established, there is no consensus about its natural history, its real significance as a disease and its relationship to endometrial neoplasia. The endometrial polyps are the main indication of hysteroscopic surgery, without a defined protocol for its better management. Presenting an overview of this condition can aid in tracing indicators for its proper treatment. To evaluate hysteroscopic features and the immunohistochemical profile of endometrial polyps in women followed by the Gynecological Endoscopy Sector from Botucatu Medical School - UNESP. This is a clinical study with a prospective and cross-sectional analysis with convenience sample of 82 women who underwent surgical hysteroscopic polypectomy and 20 women undergoing oncological surgery due to endometrial cancer, between January 2010 to December 2011. We analyzed the clinical, sonographic, hysteroscopic, histological and immunohistochemical parameters of endometrial polyps and the cancer of endometrium. For statistical analysis of quantitative variables we considered the Student's t test for independent samples and nonparametric Mann-Whitney test. For qualitative variables we used the Goodman test for contrasts between and within multinomial populations, being adopted the 5% level of significance. In comparison of outpatient endometrial biopsies, guided and directed, the sensitivity of directed biopsy in the diagnosis of endometrial polyps ranged from 35.3% to 36.8% when performed at the apex and base of the lesion, while for the guided biopsy was 29.2%. Specificity was 33.3%, 50% and 60% respectively for each type of biopsy. The positive predictive values were 75%, 77.8% and... (Complete abstract click electronic access below)

Orientador: Jorge Nahás Neto
Coorientador: Eliana Aguiar Petri Nahás
Banca: Nilton José Leite
Banca: Reginaldo Guedes Coelho Lopes
Banca: Waldir Pereira Modotte
Banca: Ricardo Bassif Lasmar
Resumo: Pólipos endometriais são neoformações resultantes da hipertrofia focal da camada basal do endométrio associada a um hiperestímulo hormonal. Sua etiopatogenia ainda não está bem estabelecida, não havendo consenso sobre sua história natural, seu real significado como entidade patológica e sua relação com a neoplasia endometrial. Os pólipos endometriais são a principal indicação de histeroscopia cirúrgica, sem que haja, no entanto, protocolo estabelecido para seu melhor manejo. Apresentar um panorama sobre esta condição pode auxiliar na busca em traçar indicadores para seu tratamento adequado. Avaliar os dados histeroscópicos e o perfil imuno-histoquímico dos pólipos endometriais de mulheres acompanhadas no Setor de Endoscopia Ginecológica da Faculdade de Medicina de Botucatu - UNESP. Trata-se de estudo clínico de avaliação prospectiva e transversal com amostra de conveniência de 82 mulheres, submetidas à polipectomia histeroscópica cirúrgica, e 20 mulheres, submetidas à cirurgia oncológica devido à neoplasia endometrial, entre o período de janeiro de 2010 a dezembro de 2011. Foram analisados parâmetros clínicos, ultrassonográficos, histeroscópicos, histopatológicos e imuno-histoquímicos dos pólipos endometriais e do câncer de endométrio. Para estudo estatístico das variáveis quantitativas considerou-se o teste t de Student para amostras independentes, e o teste não paramétrico de Mann-Whitney. Para as variáveis qualitativas utilizou-se o teste de Goodman para contrastes entre e dentro de populações multinomiais, tendo sido adotado o nível de 5% de significância. Na comparação entre biópsias endometriais ambulatoriais, orientada e dirigida, a sensibilidade da biópsia dirigida no diagnóstico dos pólipos endometriais variou de 35,3% a 36,8%, quando realizada no ápice e na base da lesão... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Endometrial polyps are focal neoformation resulting from hypertrophy of the basal layer of the endometrium associated with a hormonal hyperstimulation. Its pathogenesis is not well established, there is no consensus about its natural history, its real significance as a disease and its relationship to endometrial neoplasia. The endometrial polyps are the main indication of hysteroscopic surgery, without a defined protocol for its better management. Presenting an overview of this condition can aid in tracing indicators for its proper treatment. To evaluate hysteroscopic features and the immunohistochemical profile of endometrial polyps in women followed by the Gynecological Endoscopy Sector from Botucatu Medical School - UNESP. This is a clinical study with a prospective and cross-sectional analysis with convenience sample of 82 women who underwent surgical hysteroscopic polypectomy and 20 women undergoing oncological surgery due to endometrial cancer, between January 2010 to December 2011. We analyzed the clinical, sonographic, hysteroscopic, histological and immunohistochemical parameters of endometrial polyps and the cancer of endometrium. For statistical analysis of quantitative variables we considered the Student's t test for independent samples and nonparametric Mann-Whitney test. For qualitative variables we used the Goodman test for contrasts between and within multinomial populations, being adopted the 5% level of significance. In comparison of outpatient endometrial biopsies, guided and directed, the sensitivity of directed biopsy in the diagnosis of endometrial polyps ranged from 35.3% to 36.8% when performed at the apex and base of the lesion, while for the guided biopsy was 29.2%. Specificity was 33.3%, 50% and 60% respectively for each type of biopsy. The positive predictive values were 75%, 77.8% and... (Complete abstract click electronic access below)
Doutor

The TGF-β and the PI3K/Akt/mTROC1 pathways are two important regulators of the homeostasis of endometrial cellular cells. Alterations in the elements of these two pathways have been associated with endometrial carcinogenesis. On the one hand, the PI3K/Akt pathway is one of the main regulators of cell proliferation in the endometrium, this pathway is altered in a large number of endometrial tumors, both due to gain of function alterations and also because a loss of its main inhibitor, Pten. On the other hand, the TGF-/Smad pathway is also capable of regulating cell proliferation in a large number of tissues. Its role in cell regulation is controversial cause it can act as a tumor suppressor or as a powerful inducer of EMT and metastasis, depending on the cellular context on which it acts. That is why this thesis analyzes the effect of TGF- in the same cell type (endometrial epithelial cells) but in two different contexts, one in a well-polarized three-dimensional culture and another in a two-dimensional culture with poor cellular polarization. In recent times, many interaction mechanisms have been described between these two pathways that attempt to explain the coordinated effects which occurs between the PI3K/Akt and the TGF-β pathway that regulate processes such as proliferation and cell survival. In this thesis, the effects of cellular polarity and alterations in the PI3K/Akt and the TGF-/Smad pathways have been studied in terms of proliferation and survival on endometrial epithelial cells, as well as its role in the development and progression of endometrial cancer. To achieve these objectives, genetically modified murine models and three-dimensional endometrial cell cultures have been used. In the case of three-dimensional cultures of normal endometrial cells, TGF- is capable to induce apoptotic cell death. However, in Smad3 or Pten deficient cultures, there is a total inhibition of the apoptotic cell death induced by TGF-β and a marked increase in cell proliferation. These results support a tumor suppressor role of TGF-β when acting on endometrial cells, and that molecular alterations that affect its signaling can result in the acquisition of resistance to apoptosis and an increased cell proliferation, two of the fundamental characteristics of oncogenic processes. At a molecular level, the possible interactions between PTEN and Smads have been studied. In this terms, the results show that Smad3 directly regulates Pten transcription and, on the other hand, that PTEN regulates the activation of the Smads, establishing a regulation loop between these two pathways that control cell proliferation and tumor suppressor functions in the endometrium. In the second part of this thesis, the effects of the presence of extracellular matrix and the establishment of correct cellular polarity in relation to TGF-β effects on endometrial cells have been studied. Unlike the effects observed in three-dimensional cultures, the lack of extracellular matrix and the consequent loss of cellular polarity causes a completely different response to TGF-β addition to these cells changing the apoptotic cell response to an induction of the epithelial-mesenchymal transition leading a brake in cell proliferation. At a molecular level, the absence of the extracellular matrix has been shown to cause an over-activation of the PI3K/Akt and the ERK/MAPK pathways, resistance to apoptosis induced by TGF-β and a promotion of the epithelial to mesenchymal transition. These results show that the correct establishment of cellular polarity due to the presence of cell matrix can act as an important tumor suppressor.
La via del TGF-β i la via PI3K/Akt/mTROC1 són dos importants reguladors de l’ homeòstasi cel·lular de les cèl·lules endometrials. Alteracions en elements d’ aquestes dues vies s’ han associat a processos relacionats amb la carcinogènesi endometrial. Per una banda, la via PI3K/Akt és una de les principals reguladores de la proliferació cel·lular en l’ endometri, aquesta via es troba alterada en un gran nombre de tumors endometrials, tant per alteracions que li confereixen guany de funció com per alteracions que provoquen la pèrdua del seu principal inhibidor, Pten. Per altra banda, la via TGF-/Smad també és capaç de regular la proliferació cel·lular en un gran nombre de teixits, el seu paper en quan a aquesta regulació és controvertit ja que pot actuar tant com a supressor tumoral com a un potent inductor de l’ EMT i la metàstasis, tot depenent del context cel·lular sobre el qual actua. És per això, que en aquesta tesi s’ analitza l’ efecte del TGF- en el mateix tipus cel·lular (cèl·lules epitelials d’ endometri) però en dos contextos diferents, un en cultiu tridimensional ben polaritzat gràcies a la presència de matriu extracel·lular i un altre en cultiu bidimensional amb una polarització cel·lular deficient. En els últims temps s’ han descrit nombrosos mecanismes d’ interacció entre aquestes dues vies que intenten explicar els efectes coordinats que es donen entre la via PI3K/Akt i la del TGF- en processos tals com proliferació i supervivència cel·lular. En aquesta tesi s’ han estudiat els efectes de la polaritat cel·lular i de les alteracions en la via PI3K/Akt i TGF-/Smad en la proliferació i la supervivència de les cèl·lules epitelials d’ endometri, així com també el seu paper en el desenvolupament i la progressió del càncer endometrial. Per assolir aquests objectius s’ han utilitzat models murins genèticament modificats i cultius tridimensionals de cèl·lules endometrials. En el cas dels cultius tridimensionals de cèl·lules d’ endometri normals el TGF- és capaç d’ induir una mort cel·lular apoptòtica. En canvi, en cultius deficients en Smad3 o Pten, hi ha una inhibició total de la mort induïda per TGF- i un marcat increment de la proliferació cel·lular. Aquests resultats recolzen un paper de supressor tumoral del TGF- quan actua sobre cèl·lules d’ endometri, i que alteracions moleculars que afecten a la seva senyalització resulten en l’ adquisició de resistència a l’ apoptosi i en un increment de la proliferació cel·lular, dos de les característiques fonamentals dels processos oncogènics. A nivell molecular, s’ han estudiat les possibles interaccions entre PTEN i les Smads. En aquest sentit, els resultats demostren que, per una banda, Smad3 regula de forma directa la transcripció de Pten i, per una altra banda, que PTEN regula l’ activació de les Smads, establint-se així un bucle de regulació entre ambdues vies que controla la proliferació cel·lular i condiciona la funció de supressor tumoral en l’ endometri. En la segona part d’ aquesta tesi, s’ han estudiat els efectes de la presència de matriu extracel·lular i l’ establiment d’ una correcta polaritat cel·lular en relació amb els efectes del TGF- en cèl·lules endometrials. A diferència del efectes observats en cultius tridimensionals, la manca de matriu extracel·lular i la conseqüent pèrdua de la polaritat cel·lular fa que l’ addició del TGF- en aquestes cèl·lules canviï completament la resposta cel·lular apoptòtica per una inducció de la transició epiteli-mesènquima i una frenada en la proliferació. A nivell molecular, s’ ha demostrat que l’ absència de matriu extracel·lular causa una sobre-activació de les vies PI3K/Akt i ERK/MAPK, resistència a l’ apoptosi induïda pel TGF- i una promoció de la transició epiteli-mesènquima. Aquests resultats demostren que el correcte establiment de la polaritat cel·lular gràcies a la presència de matriu extracel·lular pot actuar com a un important supressor tumoral.
La vía del TGF-β y la vía PI3K/Akt/mTROC1 son dos importantes reguladores de la homeostasis celular de las células endometriales. Alteraciones en elementos de estas dos vías se han asociado a procesos relacionados con la carcinogénesis endometrial. Por un lado, la vía PI3K/Akt es uno de los reguladores principales de la proliferación celular en el endometrio, esta vía se encuentra alterada en un gran número de tumores endometriales, bien por alteraciones que le confieren ganancia de función como también por alteraciones que provocan la pérdida de su principal inhibidor, Pten. Por otra parte, la vía TGF-/Smad también es capaz de regular la proliferación celular en un gran número de tejidos, su papel en esta regulación es controvertido ya que puede actuar tanto como supresor tumoral o como un potente inductor de la EMT y la metástasis, dependiendo del contexto celular en el que actúa. Es debido a esta dicotomía, que en esta tesis se analiza el efecto del TGF- sobre el mismo tipo celular (células epiteliales endometriales) pero en dos contextos diferentes, uno en cultivo tridimensional bien polarizado y otro en cultivo bidimensional con una polarización celular deficiente. En los últimos tiempos se han descrito numerosos mecanismos de interacción entre estas dos vías que intentan explicar los efectos coordinados que se dan entre la vía PI3K/Akt y la del TGF- en procesos tales como proliferación y supervivencia celular. En esta tesis se han estudiado los efectos de la polaridad celular y de las alteraciones en la vía PI3K/Akt y TGF-/Smad en la proliferación y la supervivencia de las células epiteliales de endometrio, así como también su papel en el desarrollo y progresión del cáncer endometrial. Para alcanzar estos objetivos, se han utilizado modelos murinos genéticamente modificados y cultivos tridimensionales de células endometriales. En el caso de los cultivos tridimensionales de células de endometrio normales el TGF- es capaz de inducir una muerte celular apoptótica. En cambio, en cultivos deficientes en Smad3 o Pten, hay una inhibición total de la muerte inducida por TGF- y un marcado incremento de la proliferación celular. Estos resultados apoyan un papel de supresor tumoral del TGF- cuando actúa sobre células de endometrio, así como el hecho que alteraciones moleculares que afectan a su señalización resultan en la adquisición de resistencia a la apoptosis y en un incremento de la proliferación celular, dos de las características fundamentales de los procesos oncogénicos. A nivel molecular, se han estudiado las posibles interacciones entre PTEN y las Smads. En este sentido, los resultados demuestran que Smad3 regula de forma directa la transcripción de Pten y, por otra parte, que PTEN regula la activación de las Smads, estableciéndose así un bucle de regulación entre ambas vías que controla la proliferación celular y condiciona la función de supresor tumoral en el endometrio. En la segunda parte de esta tesis, se han estudiado los efectos de la presencia de matriz extracelular y el establecimiento de una correcta polaridad celular en relación con los efectos del TGF- en células endometriales. A diferencia de los efectos observados en cultivos tridimensionales, la falta de matriz extracelular y la consecuente pérdida de la polaridad celular hace que la adición del TGF- en estas células cambie completamente la respuesta celular apoptótica por una inducción de la transición epitelio-mesénquima y provoque una parada en la proliferación. A nivel molecular, se ha demostrado que la ausencia de matriz extracelular causa una sobre-activación de las vías PI3K/Akt y ERK/MAPK, resistencia a la apoptosis inducida por TGF- y una promoción de la transición epitelio-mesénquima. Estos resultados demuestran que el correcto establecimiento de la polaridad celular gracias a la presencia de matriz extracelular puede actuar como un importante supresor tumoral.

The health benefits of long term postmenopausal HRT are well recognised, but the re-initiation of cyclical bleeding, especially when irregular or heavy, has a profound impact on continuation rates. In this thesis, this clinical problem was investigated in 256 postmenopausal women who participated in a randomised dose ranging study of sequentially administered trimegestone (4 doses) with continuous micronised oestradiol, for 6 months. A strong dose dependent modulation of the bleeding pattern was documented. Women in the higher trimegestone dose groups had better bleeding patterns compared to those on the lower doses. This was subsequently confirmed when 134 women completed a further 6 months' extraction study with a single dose of trimegestone (0.25 mg). The presence of submucous fibroids in 176 women predicted the occurrence of abnormal bleeding pattern, however, the dose of trimegestone remained the dominant factor. These clinical data led to detailed histological and immunohistochemical evaluation of the endometrial samples collected on day 24 of the last treatment cycle. The majority of the results showed remarkable similarities between the test endometria and the natural cycle. None of the parameters clearly demonstrated a dose-dependent effect of trimegestone. When the expression of metalloproteinases -1 and -3 were assessed in specimens from women who received the highest and the lowest dose of trimegestone, where the bleeding pattern was completely different, no dose effect was demonstrated. These conclusions led me to question the ability to demonstrate this apparent similarity of endometrial effect between trimegestone and progesterone. For this I have developed a primary stromal cell culture system, and examined the expression of MMP-1, and -3 mRNA in these cells. Similarity between trimegestone and progesterone was again demonstrated.

The role of C19 steroids in the human endometrium is at present unclear. In order to gain an insight into their action, radioimmunoassay procedures were developed which had sufficient specificity and accuracy to measure testosterone, 5α-DHT, oestradiol, progesterone and androstenedione in endometrial samples. Amounts of androstenedione were greater (range 1.2-20.8 ng/mg tissue) than other steroids. Samples were obtained from patients presenting with a variety of conditions: subfertility, postmenopausal bleeding, dysfunctional uterine bleeding and abdominal pain. Patients admitted for sterilisation were used as normal controls. A significant positive correlation (r = 0.80) was found between the levels of testosterone and 5α-DHT measured in the same tissue which suggests the presence of a 5α reductase enzyme. No relationship was observed in tissue steroid concentration and age of the patients. Steroid concentrations were found to be high in tissues obtained from patients with endometrial carcinomas whereas progesterone concentration being low in subfertiles. The oestrogen, progesterone and androgen receptor levels of endometrial tissues from subfertile women were also determined using the DCC technique and not the procedure based on protamine sulphate precipitation since endometrial tissue available was very small. No correlation was found between receptor binding sites and day of cycle for any of the three steroids analysed; nor was there any correlation between age and receptor binding sites. A cyclic variation followed by normal women was seen in the oestrogen and progesterone receptor concentrations in the menstrual cycle. Such a variation was also observed in subfertile women on clomiphene citrate therapy. It is concluded that normal endometrium contains measurable quantities of androgens and that a receptor for 5α-DHT is present. The difference in steroid concentrations between normal and pathological states suggest that C19 steroids may be induced in the development of abnormalities.

INTRODUCCIÓN: En la práctica clínica de la Reproducción Asistida, cada vez es más habitual la realización de ciclos de transferencia de embriones. Este incremento no solo sucede por la edad avanzada de las pacientes y la consecuente indicación de ovodonación y embriodonación, sino también por el aumento del número de ciclos de criotransferencia. La introducción de la vitrificación como técnica de criopreservación y el aumento de la supervivencia y desarrollo embrionario permiten la existencia de un excedente de embriones, que pueden ser criopreservados para su posterior transferencia. La preparación endometrial previa a la transferencia puede realizarse siguiendo el ciclo natural de la paciente o administrando estrógenos exógenos durante la fase proliferativa. La vía de administración de los estrógenos más popular es la oral, si bien la transdérmica podría ofrecer ciertas ventajas sobre ella. OBJETIVOS: Nuestro objetivo principal es confirmar que la vía transdérmica es igual o superior a la vía oral para la administración de estrógenos en los ciclos de preparación endometrial para la transferencia de embriones, teniendo como variable principal el número de días necesarios para obtener un endometrio trilaminar de más de 7 mm. Como variables secundarias, evaluaremos la comodidad de la vía y satisfacción de la paciente, los niveles plasmáticos de estradiol y las tasas de embarazo, aborto y parto. MATERIAL Y MÉTODOS: Planteamos un estudio prospectivo, randomizado y comparativo, en el que 140 pacientes que van a realizar un ciclo de transferencia de embriones son aleatorizadas en 2 grupos: al grupo I se le asignará una pauta con comprimidos de valerato de estradiol; el grupo II recibirá una pauta con parches de estradiol hemidrato. Se medirá el grosor endometrial, mediante ecografía transvaginal, el día 10+1 de ciclo y, en caso de que no se haya alcanzado un endometrio de más de 7 mm, se realizará una segunda medición el día +15. Se realizará una determinación de los niveles de estradiol en sangre el día que la línea endometrial sea superior a 7 mm. Las pacientes rellenarán un cuestionario de satisfacción, en el que evaluarán la comodidad y efectos secundarios del tratamiento. RESULTADOS: Las pacientes del grupo II alcanzaron un endometrio significativamente mayor el día del primer control que las del grupo I (7.59 mm vs 7.01 mm; p:0.026), con menores niveles de estradiol en sangre (159.16 pg//ml vs 237.14 pg/ml; p:0.000) . Las pacientes del grupo I encontraron su tratamiento más cómodo que las del grupo II. En el grupo II, se reportaron más casos de mastalgia y dermatitis. No hubo diferencias significativas en cuanto a tasa de embarazo, aborto y parto. CONCLUSIONES: El tratamiento con estrógenos transdérmicos permite alcanzar un mayor grosor endometrial en menor número de días, con menores niveles plasmáticos de estradiol, aunque es peor tolerado por las pacientes.
INTRODUCTION: In the clinical practice of Assisted Reproduction, it is more and more frequent the performance of embryos transfer’s cycles. This increment is due not only to the patient’s advanced age and the subsequent indication of egg donation and embryos donation, but also to the increasing number of frozen embryos transfer. The introduction of vitrification as cryopreservation technique and the increment of embryo survival and development allow the existence of extra embryos that can be cryopreserved for its posterior transfer. Previous endometrial preparation can be performed following patient’s natural cycle or administering exogenous estrogens during the proliferative phase. The most common way of estrogens administration is the oral one, although the transdermal way might offer some advantages over the oral one. OBJECTIVE:Our fundamental aim is to confirm that the transdermal way is equal or superior than the oral way for the estrogens administration in endometrial preparation cycles for embryos transfer, taking as principal variable the number of days necessary for obtaining a triple line endometrium thicker than 7mm. As secondary variables, we evaluate comfort and patient’s satisfaction, plasmatic levels of estradiol as well as pregnancy, miscarriage and delivery rates. MATERIALS AND METHODS: We outlined a prospective, randomized and comparative study. In this study, 140 patients who are going to undergo an embryo transfer cycle are randomized in 2 groups: group I will follow a protocol with pills of estradiol valerate; group II will receive a protocol with patches of estradiol hemihydrate. Endometrial thickness will be measured with transvaginal ultrasound on day 10 of the cycle. Should the lining not measure more than 7mm, a second measurement will be performed on day 15 of the cycle. A blood analysis of estradiol level will be performed when the lining is thicker than 7mm. Patients will complete a satisfaction survey to evaluate treatment’s comfort and side effects. RESULTS: Patients of group II reached a significantly thicker endometrium on the day of first control compared to group I’s patients (7.59 mm vs 7.01 mm; p:0.026), with lower level of estradiol in blood (159.16 pg//ml vs 237.14 pg/ml; p:0.000). Patients of group I found their treatment more comfortable than patients of group II. In group II more cases of mastalgia and dermatitis were detected. No significant differences were found in terms of pregnancy, miscarriage and birth rates. CONCLUSIONS:Treatment with transdermal estrogens allows to reach a higher endometrial thickness in less days, with lower plasmatic levels of estradiol, although it is worst tolerated by patients.

Endometrial polyps (EPs) are overgrowths of endometrial glands that are protruding into the uterine cavity. EPs are benign in nature and affect both reproductive age and postmenopausal women. The aims were to quantify the blood and lymphatic microvasculature of EPs, adjacent and distant endometria. Assess the variation of these vasculatures across the menstrual cycle. Determine if these vasculatures are related to common symptoms of EPs. It was hypothesised that Microvasculature densities of EPs would be higher compared to normal endometrium, with no variation of these vessel densities across the menstrual cycle and that it would differ in women with and without symptoms. Archived samples were collected from 20 women with confirmed endometrial polyps and 32 women without endometrial polyps during the stages of the menstrual cycle. Immunohistochemistry was performed with CD31 (blood vessels) and D2-40 (lymphatic vessels). There were no significant differences in blood vessels between endometrial polyps and adjacent, distant and control endometria (F (3,70) = 2.36, p = 0.079) and like wise in lymphatic vessel density (F (3,70) = 0.16, p = 0.920). Although there was a numerical difference in the mean blood vessel density, there were no significant differences observed in blood vessels between endometrial polyps and normal endometrium.

Abstract Apoptosis is a genetically programmed mechanism for a multicellular organism to remove cells that are unnecessary, or potentially harmful. The female reproductive system is characterised by a high rate of cellular proliferation. At the same time, apoptosis is also abundant during the normal physiological function of the ovary and endometrium. More than half of the 7 million oocytes that are produced during human ovarian development are deleted before birth and only about 400 oocytes reach the stage of ovulation during the female fertile lifespan. The fate of the non-ovulatory follicles is atresia, occurring through the mechanism of apoptosis. The endometrium goes through radical renewal processes during each menstrual cycle. Apoptosis has been suggested to participate in the regulation of endometrial cellular homeostasis. Errors in this mechanism can result in endometrial diseases such as hyperplasia and cancer. In this work, apoptosis and its regulation were studied in the human fetal and adult ovary, normal endometrium and endometrial pathologies. In fetal ovaries, apoptosis was already abundantly present in oocytes at 13 weeks of gestation. The maximum rate of apoptosis was seen between the 14th and 20th weeks, after which apoptosis decreased towards term. Ovarian Bcl-2 expression was detected in early fetal life during weeks 13 and 14. Bax expression was observed throughout the studied period, from week 13 to 40. The expression of transcription factor GATA-4, which is linked to follicular survival, was localised to the granulosa cells and was high in early fetal life and decreased somewhat towards term. In adult life apoptosis was located in the granulosa cells of the growing follicles. In ovarian biopsies from women homozygous for the inactivating C566T mutation of the FSH receptor, apoptosis or GATA-4 expression was not detected. During corpus luteum regression a peak in apoptosis was detected 10 - 12 days after the LH surge, and was preceded by an increase in 17HSD type 1 and TNF-α expression. During normal menstrual cycles, the highest rate of apoptosis was observed in the menstrual endometrium. This increase in apoptosis was preceded by a decreased Bcl-2/Bax ratio. In endometrial hyperplasia, the rate of apoptosis was similar to that seen during normal proliferation of the endometrium, but an apparent increase was observed in grade II endometrial carcinoma. In grade III carcinoma, the rate of apoptosis was lower than in grade II carcinoma but higher than in hyperplasia. These results indicate that apoptosis is the mechanism behind the substantial oocyte demise during ovarian development. During adult life, apoptosis was mainly localised to the granulosa cells of the growing follicles which do not reach the stage of a dominant follicle. In ovaries where FSH action is abolished, folliculogenesis was impaired and ovarian apoptosis was negligible. Apoptosis is also the underlying mechanism of corpus luteum regression. In the endometrium, apoptosis has a role in rejuvenating the endometrium for growth during the next endometrial cycle and in regulating cellular homeostasis.

Obesity is a global health problem and the current available evidence from the literature suggests that obese women may suffer from a wide spectrum of reproductive complications. The current understanding of obesogenic effects on the peri-implantation endometrium is limited and has become an important research topic as the emerging clinical evidence from the published studies indicate the possible role of the endometrium. The first part of this thesis addresses the clinical question of whether an early pregnancy outcome is affected by the body mass index and whether there is any difference in time taken to achieve pregnancy in obese women with recurrent miscarriage when compared to normal weight women. The results are in chapters 3 and 4, where we demonstrated that obese women were more likely to have miscarriage of empty gestational sac or anembryonic pregnancies. In recurrent miscarriage context, the obese women were more ‘super-fertile’ suggesting the possible loss of an endometrial ability to select normal from abnormal pregnancies. The second part of this thesis provides an analysis of the peri-implantation endometrial stromal compartment in normal weight and obese women. In chapter 5, using immunohistochemical methods it was shown that there was no difference in the uterine natural killer cell and macrophage density in the peri-implantation endometrium of different weight groups. This suggests that the endometrial dysfunction in obese women with reproductive failure does not appear to be immune cell mediated. In chapter 6 it was shown that the clonogenecity of endometrial mesenchymal stem cells (W5C5+) was significantly negatively correlated with the BMI. The obese women had significantly reduced cloning efficiency of W5C5+ cells when compared to normal weight women, suggesting of a possible sub-optimal regenerative capacity of the endometrium in obese women. Finally, chapter 7 showed a potential association between obesogenic environment and impaired stromal cell decidualisation. Using an in-vitro model, it was shown that there was no significant difference in the expression of decidualisation markers (PRL and IGFBP1) in the decidualising endometrial stromal cells from normal weight women when compared to high BMI women. However, when the stromal cells were decidualised in an artificial obesogenic environment, the PRL expression was significantly inhibited in the presence of supernatant from adipose tissue explants of obese women when compared to normal weight women. In summary, the findings from my work have provided an understanding of the peri-implantation endometrium in obese women and evidence to suggest that the endometrial stromal function is possibly facilitated by metabolic influences.

The present study has investigated the expression of the chemokines IL-8 and MCP-1, and COX-2 in the human non-pregnant and pregnant endometrium. Both IL-8 and MCP-1 were localised by immunohistochemistry to the perivascular cells of all blood vessel types in the endometrium. Similarly COX-2 immunostaining was present in the vasculature and glandular epithelium. Reduced immunoreactivity was detectable in the periovulatory, early and mid secretory phases. Maximal levels were identified in the premenstrual or late secretory phase, coinciding with progesterone withdrawal and in addition, with the accumulation of leukocytes in uterine stroma. In first trimester decidua, low levels of expression were observed in IL-8 and COX-2, with slightly higher levels for MCP-1. This pattern was confirmed by measurement of mRNA levels by non-competitive semi-quantitative RT-PCR. Equal loading of RNA samples was ensured by measurement of GAP-DH expression levels. RT-PCR product yield for GAP-DH, IL-8, MCP-1 and COX-2 was determined by ELISA. Maximal expression was observed in menstrual phase samples. Decidual mRNA concentrations were consistent with immunohistochemical data. The role of progesterone in regulation of local mediator expression was examined by immunohistochemical analysis of endometrium and decidua obtained from women with clinically perturbed progesterone concentrations. Additionally, RNA extracted from tissue from timed endometrial biopsies was subjected to RT-PCR for GAP-DH, IL-8, MCP-1 and COX-2 as before. 48 hours after exogenous progesterone withdrawal a significant elevation in IL-8 and COX-2 mRNA expression was detected. Further supportive evidence for an inhibitory role for progesterone was the low expression in women administered hCG to extend the lifespan of the corpus luteum.

The final, definitive version of this paper has been published in Acta radiologica by SAGE Publications Ltd, All rights reserved.Final publication is available at http://acr.sagepub.com/
Kyoto University (京都大学)
0048
新制・課程博士
博士(医学)
甲第19582号
医博第4089号
新制||医||1014(附属図書館)
32618
京都大学大学院医学研究科医学専攻
(主査)教授 山田 泰広, 教授 戸井 雅和, 教授 羽賀 博典
学位規則第4条第1項該当

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Avaliar os fatores clínicos preditivos para o desenvolvimento dos pólipos endometriais em mulheres na pós-menopausa. Métodos: Foi conduzido ensaio clínico, analítico, comparativo e transversal, envolvendo mulheres na pós-menopausa, 132 com diagnóstico anatomopatológico de pólipo endometrial e 264 sem alterações endometriais, atendidas em Hospital Público Universitário. Foram incluídas no estudo mulheres com amenorréia ≥ 12 meses e idade ≥ 45 anos, em uma proporção de 1 caso para 2 controles. Foram coletados dados clínicos, antropométricos, laboratoriais e ultrassonográficos, para avaliação dos fatores preditivos do pólipo endometrial. Para análise estatística empregou-se os testes t-Student, Qui-Quadrado e Regressão Logística (OR, odds ratio). Resultados: As pacientes com pólipo endometrial apresentaram idade mais avançada e maior tempo de menopausa quando comparadas ao controle (P<0,0001). Maior porcentagem de mulheres com pólipo era obesa (72%) quando comparadas ao controle (39%) (P<0,0001). A medida da circunferência da cintura foi maior entre as pacientes com pólipo (P=0,0001). Observou-se maior incidência de diabetes, hipertensão e dislipidemias nas pacientes com pólipo endometrial (P<0,0001). De acordo com os critérios do NCEP/ATP III, 48,5% das mulheres com pólipo e 33,3% do grupo controle foram classificadas como portadoras da síndrome metabólica (P=0,004). Em relação ao grupo controle, apresentaram maior chance de desenvolvimento de pólipo endometrial às pacientes com: IMC ≥ 25 kg/m2 (OR 4,66; IC 95% 2,16-10,05); glicose ≥ 100 mg/dl (OR 2,83; IC 95% 1,36-5,90); dislipidemia (OR 7,02; IC 95% 3,70-13,32), diabetes (OR 2,58; IC 95% 1,05-6,32); e síndrome metabólica (OR 2,76; IC 95% 1,18-6,46). Conclusão: Em mulheres na pós-menopausa, a obesidade, a dislipidemia, a hiperglicemia e a presença de síndrome metabólica foram fatores preditivos para o desenvolvimento de pólipo endometrial
Objective: To evaluate the clinical predictive factors of endometrial polyps in postmenopausal women. Methods: In this clinical, analytical, comparative and cross-sectional study 132 patients with histopathologic diagnosis of endometrial polyp were compared with 264 without endometrial alterations (control), seeking healthcare at a Public University Hospital. In the study were included women with amenorrhea ≥ 12 months and age ≥ 45 years, in a proportion of 1 case to 2 controls. Clinical, anthropometric, laboratory and ultrasonographic data were collected for evaluating the predictive factors of endometrial polyps. For statistical analysis were used the Student-t, chi-square tests and logistic regression method (odds ratio-OR). Results: Patients with endometrial polyps were older and had more time since menopause when compared to control (P <.0001). A higher percentage of women with endometrial polyps were obese (72%) compared to control (39%) (P <.0001). The measurement of waist circumference was higher among patients with polyps (P = 0.0001). We observed a higher incidence of diabetes, hypertension and dyslipidemia in patients with endometrial polyps (P <.0001). According to the NCEP/ATP III, 48.5% of women with polyps and 33.3% in the control group were classified as having metabolic syndrome (P = 0.004). Analysis of risk for endometrial polyp formation was higher in patients with: BMI ≥ 25 kg/m2 (OR 4.66; CI 95% 2.16-10.05); glucose ≥ 100 mg/dl (OR 2.83; CI 95% 1.36-5.90); dyslipidemia (OR 7.02; CI 95% 3.70-13.32); diabetes (OR 2.58; CI 95% 1.05-6.32); and metabolic syndrome (OR 2.76; CI 95% 1.18-6.46). (P = 0.019) compared to control. Conclusion: In postmenopausal women, obesity, dyslipidemia, hyperglycemia and the presence of metabolic syndrome were predictive factors for endometrial polyp

This thesis is based on the hypothesis that menstrual dysfunction, including heavy menstrual blood loss (MBL) is due to (a) up-regulated expression/synthesis of cycloxygenase enzymes and prostaglandin receptors, and (b) initiation of enhanced intracellular signalling pathways in response to prostaglandins. This thesis describes the use of an endometrial epithelial cell line to explore the molecular signalling pathways involved with the activation of a prostaglandin receptor – the prostacyclin receptor. A rapid activation of ERK1/2 signalling is demonstrated with alterations in expression of angiogenic factors via crosstalk with the epidermal growth factor receptor. By using endometrium collected from women with the complaint of heavy menstrual bleeding, the pattern of expression of the various components of the COX-prostaglandin signalling pathways present in the endometrium of women with normal and heavy MBL is elucidated. There is a significant elevation in expression of COX-1 and COX-2 mRNA in endometrium obtained from women with heavy MBL compared with endometrium obtained from women with normal MBL. Significant alterations in expression of downstream prostanoid synthase and prostanoid receptor mRNAs were also detected. Furthermore, enhanced prostaglandin stimulated production of cyclic AMP observed in endometrium of women with heavy MBL compared with normal MBL. By identifying the prostanoid receptors/signalling pathways that are responsible for disturbed endometrial function, this thesis aims to establish information that may result in the development of novel therapeutic targets for menstrual pathology.

Thesis (M.S.)--West Virginia University, 2000.
Title from document title page. Document formatted into pages; contains vii, 79 p. : ill. (some col.). Includes abstract. Includes bibliographical references (p. 68-79).

Tirtos Vilniaus universiteto Onkologijos institute gydytos pacientės, kurioms 1996 spalio mėnesį - 1998 kovo mėnesį buvo taikyta pooperacinė spindulinė terapija gama-neutronų šaltiniu gimdos kūno vėžio gydymui. Tyrimo tikslas: įvertinti gimdos kūno vėžio pooperacinio spindulinio gydymo gama-neutronų šaltiniu efektyvumą. Tyrimo uždaviniai: 1) Įvertinti vienerių, dvejų, trejų ir penkerių metų išgyvenamumą pacienčių grupėje. 2) Įvertinti vienerių, dvejų, trejų ir penkerių metų išgyvenamumą pacienčių amžiaus grupėje. 3) Įvertinti vienerių, dvejų, trejų ir penkerių metų išgyvenamumą pagal naviko charakteristikas: stadiją, morfologinį naviko tipą, naviko diferenciacijos laipsnį. 4) Įvertinti vienerių, dvejų, trejų ir penkerių metų išgyvenamumą pagal spindulinio gydymo charakteristikas: suminę židininę dozę, gydymo trukmę, laiką nuo diagnozės iki gydymo pradžios. Medžiaga ir metodai: naudotasi VU onkologijos institute esančiomis ambulatorinių kortelių ir vėžio registro duomenimis - iš viso 64 ligonių ligos istorijos: 52 pacientėms buvo taikyta pooperacinė ertmini gama-neutonų terapija + distancinė gamaterapija, 12 ertminė gama-neutronų terapija. Duomenų analizė atlikta naudojantis SPSS 11.0 versijos statistiniu duomenų analizės paketu. Skirtumui tarp grupių įvertinti taikytas Chi-kvadrato metodas. Visoms hipotezėms tikrinti taikytas reikšmingumo lygmuo 0,05. Išgyvenamumo analizė atlikta naudojant Kaplan - Meier ir išgyvenimo lentelių (life table) metodą. Rezultatai: Ertminės... [toliau žr. visą tekstą]
Women were treated with postoperative radiotherapy by using the gamma-neutrons’ source for treatment of endometrial cancer within the period from October 1, 1996 till March 1, 1998 at the Institute of Oncology of Vilnius University were surveyed in this study. Objective: to estimate the efficiency of endometrial cancer postoperative radiotherapy treatment by using the gamma-neutron source. Goals :1) to estimate one, two, three and five - year survival in a group of patients; 2) to estimate one, two, three and five - year survival according to the patients’ age; 3) to estimate one, two, three and five - year survival according to the tumor characteristics: morphologic tumor type, tumor differentiation degree; 4) to estimate one, two, three and five - year survival according to the radiotherapy characteristics: radiotherapy type, radiotherapy duration. Materials and Methods: The used materials consist of the case records collected at the Institute of Oncology of Vilnius University as well as of the data of the Cancer Register - overall 64 patients’ case records: 52 patients have had combined postoperative radiotherapy and 12 have had vaginal brachytherapy. The data were analyzed by the SPSS statistical package (Version 11.0 for Windows). The statistical assessment and analysis were performed with the qui-square (χ2), Kruscal-Wallis H test with values of p < 0,05 being defined as significant. The survival analysis was performed by using the Kaplan - Meier and life tables’... [to full text]

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
O objetivo do presente estudo foi avaliar a relação entre a intensidade de alterações degenerativas endometriais e da idade com a hemodinâmica uterina durante o período posterior à infusão de sêmen (Experimento 1) e durante a fase inicial da gestação (Experimento 2). Vascularidade, contratilidade e índices Doppler do útero foram mensurados durante as 12 primeiras horas pós-inseminação artificial (IA) no experimento1. Não foi observado (P>0,1) efeito da posição do folículo pré-ovularório sobre a hemodinâmica e contratilidade uterina. Um aumento na vascularidade foi detectado (P<0,001) em éguas velhas e com degeneração endometrial moderada (GII) e intensa (GIII) nas duas primeiras horas pós-IA. Índices de pulsatilidade (PI) e resistência (RI) uterina mantiveram-se constante durante as 12 horas pós-IA em éguas dos grupos GII, jovens e velhas (P>0,05). Um aumento na atividade contrátil do útero foi detectado (P<0,0001) nas horas 1 e 2, independente do grupo experimental (P>0,05). No experimento 2, os parâmetros citados acima, além de características luteais (área e vascularidade) e diâmetro da vesícula embrionária foram avaliados diariamente em éguas gestantes e não gestantes. Aumento e diminuição, respectivamente, na vascularidade e índices Doppler uterinos foram detectados (P<0,05) em D4 (D0=dia da ovulação) em éguas gestantes e não gestantes. Um aumento progressivo na perfusão sanguínea com pronunciada vascularidade no corno ipsi-lateral ao embrião foi observado em éguas gestantes a partir de D12 (P<0,001). Efeito da idade e da intensidade das alterações degenerativas endometriais sobre a vascularidade e índices Doppler do útero gravídico foram detectados (P<0,01). A contratilidade uterina foi máxima (P<0,001) entre D10 e D17 da gestação independente da classificação histológica do...
The overall purpose of the present study was to evaluate the relationship between endometrial degenerative changes and age on the uterine hemodynamics of mares during the post-breeding (Experiment 1) and early gestation phases (Experiment 2). Vascularity, contractility and Doppler indices from the uterus were measured hourly during the first 12 hours post-artificial insemination (AI) in experiment 1. Hemodynamics and contractility of the uterus did not change (P>0.1) by pre-ovulatory follicle position. Increased uterine vascularity was observed (P<0,001) on hours 1 and 2 in older and moderate (GII) and intense (GIII) endometrial degeneration mares. Pulsatility and resistance indices (PI and RI, respectively) did not change (P>0.05) through the experiment 1 age groups (younger and older mares) mares with moderate endometrial degeneration. Independently of the age and endometrial degenerative changes, increased uterine contractility was observed (P<0,0001) in the first two hours post-AI. In experiment 2, the same end points were measured every day. Additionally, luteal vascularity and embryonic vesicle diameter were evaluated. On D4 (D0 = day of ovulation), increased vascularity and decreased Doppler indices were detected (P <0.05) in pregnant and nonpregnant mares. A progressive increase on the uterine vascular perfusion and greater vascularity in the horn with embryo were observed in pregnant mares from D12 (P <0.001). Effect of age and intensity of endometrial degenerative changes on the vascularity and Doppler indices of the gravidic uterus were detected (P<0.01). Uterine contractility was greater (P<0.001) between days 10 and 17 of pregnancy regardless of the uterine histological classification and age of mares (P> 0.05). Mean luteal vascularity and plasmatic... (Complete abstract click electronic access below)

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
O trabalho teve por objetivo avaliar a eficiência da terapia celular no tratamento do endométrio de éguas com processo degenerativo crônico caracterizado pela presença de fibrose uterina. Foram utilizadas 10 éguas, de raça Quarto de Milha com idade entre 14 e 23 anos, com massa corpórea entre 400 a 600 Kg, e bom escore corporal com históricos reprodutivos de subfertilidade e grau severo de fibrose uterina detectada previamente pelo exame histológico realizado por meio da biópsia endometrial. O material foi coletada no dia zero (D0) antes do tratamento com células tronco mesenquimais (CTMs) e 15 dias (D15), 30 dias (D30) e 60 dias (D60) após o tratamento. As amostras foram classificadas segundo o modelo proposto por Kenney & Doig (1986) em grau de fibrose: Grau I - normal, Grau IIA – inflamação crônica leve, Grau IIB – inflamação crônica com infiltrado moderado, Grau III – inflamação crônica degenerativa grave. Estas amostras foram submetidas à confecção de lâminas para coloração de Tricrômio de Masson, Coloração de Hematoxilina e Eosina (HE) e Imunohistoquímica. A punção da medula óssea foi realizada em 10 éguas com idade entre 14 e 23 anos e o material coletado foi submetido ao Laboratório. O cultivo celular in vitro foi realizado em meio DMEM alta glicose em estufa com 37,5°C, em atmosfera úmida contendo 95% de ar e 5% de CO2. Após atingir a confluência de 80% na placa de cultivo, estas células foram removidas e injetadas intra-endometrial com uma concentração de 2x107 células\mL em um volume de 0,5 ml em 20 (vinte) diferentes pontos (sitio de administração) espaçados entre um centímetro, com o auxílio de endoscópio flexível, totalizando um volume de 10 mL aplicado. Nas avaliações realizadas, verificou-se diminuição significativa dos ninhos glandulares, mudança siginificativa de células...
The study aimed to evaluate the efficiency of cell therapy in the treatment of endometrial mares with chronic degenerative process characterized by the presence of uterine fibroids. We used 10 mares bred Quarter Horses aged between 14 and 23 years, with body mass between 400 and 600 kg, with good body and reproductive history of subfertility and severe degree of uterine fibrosis detected previously by histological examination performed by endometrial biopsy. The material was collected on day zero (D0) before treatment with mesenchymal stem cells (MSCs) and 15 days (D15), 30 days (D30) and day 60 (D60) post-treatment. The samples were classified according to the model proposed by Kenney & Doig (1986) in fibrosis: Grade I - normal, Grade IIA - mild chronic inflammation, IIB Grade - moderate chronic inflammatory infiltrate, Grade III - severe degenerative chronic inflammation. These samples were submitted for preparation of slides for staining Masson's trichrome, hematoxylin and eosin staining (HE) and immunohistochemistry. A bone marrow puncture was performed in 10 mares aged between 14 and 23 years and the collected material was submitted to the Laboratory. The in vitro cell cultivation was performed in DMEM with high glucose kiln 37.5 ° C in humidified atmosphere containing 95% air and 5% CO2. After reaching 80% confluence on plate culture, these cells were removed and injected intra-endometrial at a concentration of 2x107 cells \ ml in a volume of 0.5 ml for 20 (twenty) different point (site of administration) spaced of an inch, with the aid of flexible endoscope, for a total volume of 10 ml applied. In the assessments, there was a significant reduction in glandular nests, change polymorphonuclear cell siginificativa absent at baseline (D0) to moderate 60 days (D60); amendment of mononuclear cells in going from... (Complete abstract click electronic access below)

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O Carcinoma de Endométrio é a neoplasia epitelial maligna que acomete mulheres no pré, pós e peri -menopausa. Segundo diferenças endócrinas, metabólicas, fatores de risco e morfologia é classificado em dois grandes grupos: Tipo I (endometrióide) e Tipo II (não endometrióide). O Tipo I perfaz 90 % dos carcinomas de endométrio e sua patogenia esta ligada a exposição excessiva ao estrógeno. O Tipo II é incomum, com fatores predisponentes menos conhecidos. O sintoma mais relevante do carcinoma de endométrio é o sangramento pós-menopausa. Seu diagnóstico é feito pela comprovação histológica, associada aos exames de imagem e laboratoriais. Seu tratamento é fundamentalmente cirúrgico. O fator prognóstico mais importante é a presença ou ausência de metástase nos linfonodos regionais. Atualmente, buscam-se marcadores biológicos e teciduais que indiquem pior prognóstico. Este trabalho verificou a imunoexpressão da claudina-3 (CLDN3) e claudina-4 (CLDN4) nos carcinomas de endométrio Tipo I e Tipo II, relacionando-as com endométrio proliferativo e atrófico, aspectos clínicos, anatomopatológicos, perfil hormonal, índice de proliferação celular e expressão da p53, na tentativa de estabelecer a importância destas proteínas na progressão e agressividade tumoral e o seu valor prognóstico. Foram estudados 79 casos de carcinoma de endométrio e comparados com 74 endométrios normais. Avaliou-se a imunoexpressão das CLDNs 3 e 4, receptor estrogênico, receptor de progesterona, índice de proliferação celular (Ki67) e p53, pela técnica de imunoistoquímica. Observou-se que o padrão de coloração da membrana para CLDN3 se mostrou difuso nos carcinomas, quando comparado com os endométrios normais que exibiu padrão focal. O número de células marcadas com CLDN3 estava diminuído nos carcinomas Tipo I, porém com intensidade aumentada. Nesta análise foi possível verificar que ...
Carcinoma of the Endometrium is a malignant epithelial tumor which affects pre, peri and post-menopausal women. It is classified into two major groups, according to endocrine, metabolic risk factors and morphological differences: Type I (endometrioid) and Type II (non-endometrioid). Type I accounts for 90% of all endometrial carcinomas and its pathogenesis is linked to excessive estrogen exposure. The Type II is less common, with poorly defined predisposing factors. The most important symptom of endometrial carcinoma is postmenopausal bleeding. Diagnosis is achieved through histological evidence, in association with imaging and laboratorial exams. Treatment is primarily surgical. The most important prognostic factor is the presence or absence of metastases in regional lymph nodes. Bio and tissue markers that indicate worse prognosis are the focus of current research. This study examined the immunoexpression of claudin-3 (CLDN3) and claudin-4 (CLDN4) in endometrial carcinomas Type I and Type II, and their relation to proliferative and atrophic endometrium, clinical and pathological features, hormonal status, proliferation index and p53 expression, in an attempt to establish the importance of these proteins in tumor progression and aggressiveness and their prognostic value. Seventy-nine cases of endometrial carcinoma were studied and compared with 74 normal endometria. The immunoexpression of CLDNs 3 and 4, estrogen receptor, progesterone receptor, cell proliferation index (Ki67) and p53 were all evaluated by immunohistochemistry. Observation verified that the pattern of membrane staining for CLDN3 was diffuse in carcinomas compared with normal endometrium which presented a focal pattern. The number of cells stained with CLDN3 was lower in Type I carcinomas, while staining intensity was greater. Analysis verified that 25% of cases with high expression of CLDN4 and Ki-67 developed metastasis, while 33% of cases with greater CLDN4 staining ...

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O processo degenerativo progressivo em consequência a resposta exagerada do endométrio à exposição crônica de progesterona e estrógeno, endógeno ou exógeno, resulta em hiperplasia endometrial cística (HEC) na maioria das cadelas idosas, e consequente degeneração tecidual, distensão glandular e acúmulo de secreções, que pode resultar em contaminação bacteriana, inflamação supurativa e degenerativa do endométrio, com acúmulo de exsudato nas glândulas endometriais e lúmen uterino, bacteremia, afecção hepatorenal e toxemia. A análise oligoarrays tem sido aplicada com sucesso no estudo da expressão gênica do endométrio; o único tecido com capacidade dinâmica de sofrer remodelação em resposta a variações cíclicas de hormônios esteróides e fatores locais autócrinos e parácrinos. Objetivou-se com este estudo identificar os genes diferencialmente expressos responsáveis pela proliferação e hiperplasia no endométrio de fêmeas caninas acometidas por HEC, mucometra e piometra, comparado com o tecido endometrial normal de fêmeas caninas na fase de diestro. Foi utilizado teste estatístico SAM (Significance Analysis Of Microarray), do programa TMeV v.4.5, por meio de teste estatístico paramétrico (teste t), p<0,05 e ajuste pelo teste de Bonferroni, considerando apenas os conjuntos de genes que com False Discovery Rate iguais ou inferiores a zero, e fold-change > ou < que 3,0. O grupo HEC apresentou 33 genes com expressão aumentada (upregulated) e 45 genes com expressão diminuída (downregulated) em relação ao grupo diestro, com 15 moléculas envolvidas no desenvolvimento, crescimento e proliferação celular e morfologia tumoral. O grupo mucometra apresentou 189 genes upregulated e 150 genes downregulated em relação ao grupo diestro, sendo 26 moléculas associadas ao desevolvimento embrionário e 21 moléculas envolvidas com movimentação celular, desenvolvimento e funcionamento do sistema ...
Hyperplasic and cystic alteration of the canine endometrium compromise reproduction and fertility and may lead to a degenerative inflammation and infection of endometrium. The progressive process usually proposed as the initiating lesion, is mediated by progesterone and potentially aggravated by estrogens. A separate process caused by local uterine irritation to trophoblastic reaction and bacterial proliferation has been recently proposed as an alternate mechanism leading to pyometra. In order to identify molecular similarity and potential targets for therapeutic intervention and biomarkers for endometrium proliferative conditions in humans and dogs, gene expression profiles were performed in fifteen endometrial biopsy samples from female dogs during luteal phase (diestrus), fifteen affected by endometrial cystic hyperplasia, fifteen by mucometra and fifteen by pyometra, and processed on Affymetrix Canine 1.0 ST array. The transcriptome analysis revealed expression of 115, 23 and 284 significantly genes (p<0,05) altered in the hyperplasic compared with normal endometrium, hyperplasic and aseptic secretory compared with hyperplasic endometrium, and infected compared to hyperplasic and aseptic secretory endometrium, respectively. Gene ontology enrichment analysis revealed genes associated with the cell development, growth, proliferation, function and maintenance, and cell-to-cell signaling and interaction were altered in the hyperplasic and proliferative lesion, and also in aseptic secretory endometrium, such as elevated expression of CYR61, EGR1, FOS, GALNT14, IGKC, SLC47A2, IGFBP3, and low expression of ESR2, SOCS3 and MCOLN3. The proliferative to secretory transition revealed genes associated with immune cell trafficking, and cell-mediated immune response, such as FOSB, MAL, CCL4 and SLPI. The infected endometrium due to bacterial infection revealed elevated expression of chemokines (CCL2, CCL3, CXCS), cytokines (IL8, IL6), proteases ...

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Objetivo: Avaliar a expressão imunoistoquímica de receptores de estrogênio (RE) e progesterona (RP), de proteínas relacionadas à proliferação celular (Ki-67), à neoangiogênese (endoglina - CD105), à adesão celular (claudinas 3 e 4) e proteínas da matriz extracelular (metaloproteinases 2 e 9 - MMP 2 e MMP 9) nos pólipos endometriais e no câncer de endométrio comparativamente ao endométrio normal. Tipo de Estudo: Estudo transversal comparativo com amostra de conveniência. O levantamento foi realizado através de banco de dados do Laboratório de Patologia Clínica da Faculdade de Medicina de Botucatu. Local: Hospital das Clínicas da Faculdade de Medicina de Botucatu, Universidade Estadual Paulista Júlio de Mesquita Filho - UNESP. Pacientes: Foram realizados estudos imunoistoquímicos de 30 amostras de pólipos endometriais sem atipias e de 30 amostras de adenocarcinoma endometrial do tipo endometrioide e confrontados com os resultados da análise de 30 amostras de endométrio normal (grupo controle). Intervenções: Dados epidemiológicos, clínicos e antropométricos foram levantados através de análise dos prontuários. Para análise dos casos de adenocarcinoma de endométrio e dos controles foi empregada a técnica de tissue microarray (TMA). Os blocos de parafina, com os cortes do maior fragmento de lesão polipoide e os blocos receptores de TMA foram utilizados para avaliação imunoistoquímica de RE, RP, CD105, Ki-67, claudinas 3 e 4, MMP-2 e MMP-9. Resultados Principais: Identificou-se diferença significativa entre os grupos na expressão de RE (P<0,001) e RP (P<0,05), do Ki-67 (P<0,001), do CD105 (P<0,001) e da claudina 3 (P<0,001). Não foram identificadas diferenças nos marcadores pesquisados entre pólipos e câncer de endométrio (P≥0,05). A expressão de MMP-2 e MMP-9 foi praticamente ausente nos três grupos. Conclusões: Nas amostras pesquisadas, não foram demonstradas diferenciações entre os...
Objective: To evaluate the immunohistochemical expression of estrogen receptors (RE), progesterone receptors (RP), cell proliferation (Ki-67), neoangiogenesis (endoglin - CD105), cell adhesion (claudin 3 and 4) and extracellular matrix proteins (metalloproteinases 2 and 9 - MMP 2 and MMP 9) in endometrial polyps and endometrial cancer compared to normal endometrium. Design: Comparative cross-sectional study with occasional sample. The survey was conducted from the database of the Clinical Pathology Laboratory of Botucatu Medical School - Unesp. Setting: Clinics Hospital from Botucatu Medical School, São Paulo State University - UNESP. Patients: Were performed immunohistochemical studies of 30 samples of endometrial polyps without atypia and 30 samples of endometrioid adenocarcinoma of endometrium confronted with the results of 30 normal endometrial samples (control group). Interventions: Epidemiological, clinical and anthropometric data were collected through analysis of medical records. To analyze the cases of endometrial adenocarcinoma and controls we used the tissue microarray technique (TMA). Paraffin blocks, with the larger fragment of polypoid lesions and recipient blocks of TMA were used to evaluate immunohistochemical expression of ER, PR, CD105, Ki-67, claudin 3 and 4, MMP-2 and MMP-9. Main Results: We identified a significant difference between groups in ER (P <0.001) and PR expression (P<0.05), Ki-67 (P<0.001), CD105 (P<0.001) and claudin 3 (P<0.001). No difference was found between polyps and endometrial cancer (P≥0,05). The expression of MMP-2 and MMP-9 was virtually absent in all three groups. Conclusions In the studied samples according to the immunohistochemical parameters evaluated no differentiation between polyps and endometrial cancer was observed. There was no expression of MMP-2 and -9 in endometrial tissues analyzed. Further studies are necessary to better understand ...
FAPESP: 2012/17297-3

Orientador: Cezinande de Meira
Banca: Sony Dimas Bicudo
Banca: João Carlos Pinheiro Ferreira
Banca: Lucianao Andrade de Silva
Banca: Rubens Paes de Almeida
Resumo: O objetivo do presente estudo foi avaliar a relação entre a intensidade de alterações degenerativas endometriais e da idade com a hemodinâmica uterina durante o período posterior à infusão de sêmen (Experimento 1) e durante a fase inicial da gestação (Experimento 2). Vascularidade, contratilidade e índices Doppler do útero foram mensurados durante as 12 primeiras horas pós-inseminação artificial (IA) no experimento1. Não foi observado (P>0,1) efeito da posição do folículo pré-ovularório sobre a hemodinâmica e contratilidade uterina. Um aumento na vascularidade foi detectado (P<0,001) em éguas velhas e com degeneração endometrial moderada (GII) e intensa (GIII) nas duas primeiras horas pós-IA. Índices de pulsatilidade (PI) e resistência (RI) uterina mantiveram-se constante durante as 12 horas pós-IA em éguas dos grupos GII, jovens e velhas (P>0,05). Um aumento na atividade contrátil do útero foi detectado (P<0,0001) nas horas 1 e 2, independente do grupo experimental (P>0,05). No experimento 2, os parâmetros citados acima, além de características luteais (área e vascularidade) e diâmetro da vesícula embrionária foram avaliados diariamente em éguas gestantes e não gestantes. Aumento e diminuição, respectivamente, na vascularidade e índices Doppler uterinos foram detectados (P<0,05) em D4 (D0=dia da ovulação) em éguas gestantes e não gestantes. Um aumento progressivo na perfusão sanguínea com pronunciada vascularidade no corno ipsi-lateral ao embrião foi observado em éguas gestantes a partir de D12 (P<0,001). Efeito da idade e da intensidade das alterações degenerativas endometriais sobre a vascularidade e índices Doppler do útero gravídico foram detectados (P<0,01). A contratilidade uterina foi máxima (P<0,001) entre D10 e D17 da gestação independente da classificação histológica do... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The overall purpose of the present study was to evaluate the relationship between endometrial degenerative changes and age on the uterine hemodynamics of mares during the post-breeding (Experiment 1) and early gestation phases (Experiment 2). Vascularity, contractility and Doppler indices from the uterus were measured hourly during the first 12 hours post-artificial insemination (AI) in experiment 1. Hemodynamics and contractility of the uterus did not change (P>0.1) by pre-ovulatory follicle position. Increased uterine vascularity was observed (P<0,001) on hours 1 and 2 in older and moderate (GII) and intense (GIII) endometrial degeneration mares. Pulsatility and resistance indices (PI and RI, respectively) did not change (P>0.05) through the experiment 1 age groups (younger and older mares) mares with moderate endometrial degeneration. Independently of the age and endometrial degenerative changes, increased uterine contractility was observed (P<0,0001) in the first two hours post-AI. In experiment 2, the same end points were measured every day. Additionally, luteal vascularity and embryonic vesicle diameter were evaluated. On D4 (D0 = day of ovulation), increased vascularity and decreased Doppler indices were detected (P <0.05) in pregnant and nonpregnant mares. A progressive increase on the uterine vascular perfusion and greater vascularity in the horn with embryo were observed in pregnant mares from D12 (P <0.001). Effect of age and intensity of endometrial degenerative changes on the vascularity and Doppler indices of the gravidic uterus were detected (P<0.01). Uterine contractility was greater (P<0.001) between days 10 and 17 of pregnancy regardless of the uterine histological classification and age of mares (P> 0.05). Mean luteal vascularity and plasmatic... (Complete abstract click electronic access below)
Doutor

This work has shown that 11βHSD-1 mRNA is present at highest levels in the menstrual phase of the cycle and in first trimester deciduas, the times when an inflammatory response is evident. 11βHSD-2 mRNA and protein are present at all stages of the cycle, and also in first trimester deciduas. GR mRNA and protein are highly expressed throughout the cycle. MR mRNA expression varies across the cycle in a pattern similar to progesterone expression. 11βHSD-1 mRNA expression is increased in response to IL-1α and cortisol, and GR mRNA shows a similar trend. 11βHSD-1 and MR expression are not altered by IL-1α or cortisol.3βHSD-1 mRNA has been shown to be present only in first trimester deciduas; 3βHSD-1 is not detectable by these methods. Immunohistochemistry using an antibody which detects both 3βHSD-1 and -2 has shown low levels of protein in the tissues studied. AKR1C1-3 mRNAs are expressed throughout the menstrual cycle; all three enzymes are predominantly expressed in the secretory phase. AKR1C3 is localised to the glandular and surface epithelial cells, and vascular endothelium. AKR1C4 mRNA is not detectable in the endometrium at any stage of the menstrual cycle. Expression of steroid-metabolising enzymes is perturbed in the endometrium of users of a Levonorgestrel intra-uterine system, and also the following GnRH antagonist treatment for sub-fertility. These studies have shown that the endometrium has the ability to precisely regulate its balance of steroid hormone availability at a local level, and that this balance may be altered following administration of exogenous steroids. Further functional studies such as knockout or knockdown of these enzymes would expand this knowledge and fully elucidate steroid hormone metabolism and pre-receptor signalling.

Both the endometrium and the trophoblast secrete a number of proteins. Some of these, e.g. human chorionic gonadotrophin (hCG) from the trophoblast and pregnancy-associated endometrial α2-globulin (α2-PEG) from the endometrium, are secreted into the bloodstream and may act as endocrines. Moreover, they may also function as paracrines, signalling to adjacent tissues. Several models have been developed, by which the paracrine activity of the endometrium and the trophoblast might be examined in vitro. These include, cultures of endometrium and trophoblast, dual chamber superfusion of membranes and dual perfusion of individual placental cotyledons. The purpose of the present study was to examine some aspects of experimental design on the results from these models and consider the factors that might govern the rate of release. A good many trophoblast proteins have been characterised and methods devised for their quantitative measurement, however, the study of endometrial proteins is less advanced. As a first step, a method was developed for the measurement of α2-PEG, the major secretory protein of the secretory endometrium during the first trimester of pregnancy. This thesis describes the development of a competitive ELISA for measuring α2-PEG. This assay was employed for measuring α2-PEG in the medium from decidual cultures. In this thesis, release of protein in vitro has been examined as a mode of basal unstimulated secretion in vivo. The release is compounded of at least two simultaneous processes, biosynthesis and the transport of formed material to the outside of the cell. The concept is put forward that protein release is determined by a dynamic balance between biosynthesis of new protein and diffusion of protein down the concentration gradient between cell cytoplasm and the surrounding medium.

Decidualization of the endometrium is essential for successful pregnancy and in human females of reproductive age, it occurs every month following the post-ovulatory rise in progesterone levels. The decidualization of primary human endometrial stromal cells (hESCs) can be recapitulated by treatment with cAMP and progestins, which results in changes in gene expression that give rise to phenotypes that favour implantation and survival of the conceptus. MicroRNAs (miRNAs) are a diverse class of small, non-coding RNA molecules that regulate gene expression post-transcriptionally and have important roles in many biological processes. Expression profiling revealed several miRNAs to be regulated during decidualization, and alterations in miRNA pathway components were also found. Although induction of Dicer suggested increased capacity to produce mature miRNAs, endogenous miRNA silencing became restricted upon decidualization of hESCs due to the down-regulation of the Argonaute proteins, catalytic components of the RNAinduced silencing complex. This was reflected in the regulation of miRNA target genes, which only appeared to be subject to miRNA-dependent regulation in undifferentiated hESCs. Moreover, the regulation of the androgen receptor (AR), a nuclear hormone receptor known to modulate the expression of a subset of decidual genes, was not dependent on miRNAs during decidualization. Rather, an RNA binding protein, poly(C)- binding protein 1 (PCBP1), regulated AR expression in hESCs during decidualization and in LNCaP cells. Additionally, decidualizing hESCs export miRNAs in exosome-sized vesicles that can be taken up by a variety of cells, including trophoblast and vascular cells, representing a novel mode of communication that may coordinate responses across different cell types at the feto-maternal interface.

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A expressão das interleucinas no útero e das catelicidinas no plasma seminal podem ser responsáveis pela magnitude e persistência da resposta inflamatória uterina. Os objetivos desse estudo foram verificar a presença das catelicidinas no plasma seminal e caracterizar a resposta imune uterina pela expressão de interleucinas inflamatórias. O plasma seminal foi obtido por coleta de sêmen de 15 garanhões e submetido ao SDSPAGE e DOT-blotting. As éguas foram submetidas a um ciclo estral sem desafio uterino (T1), com infusão de plasma seminal (T2) e inseminação artificial (T3). Exame ultrassonográfico, citologia exfoliativa uterina e colheita de células para a expressão do RNAm das interleucinas por qPCR foram realizados durante 3 ciclos consecutivos aleatoriamente, 24 (M1), 48 (M2) e 72 horas (M3) após a indução da ovulação. A expressão do RNAm no endométrio das éguas resistentes e susceptíveis foi verificada para as interleucinas IL-1β, IL-6, IL-8, IL-10 e TNF-α. A concentração total de proteínas no plasma seminal dos garanhões variou de 6,3 a 25,6 mg/mL. O SDS-PAGE revelou a existência de 3 bandas protéicas dentro do intervalo de peso molecular previamente citado na literatura para as catelicidinas equinas. Os garanhões avaliados mostraram quantidades de eCATH1 < 339,6ng, eCATH2 < 141,3ng e eCATH3 < 283,4ng. No momento M2 não foi possível diferenciar as éguas resistentes das susceptíveis pela citologia ou ultrassonografia independente da infusão com plasma seminal, inseminação artificial ou da ovulação, mas quando há presença de espermatozoides no útero a resposta inflamatória é mais intensa. Durante o momento M3, as éguas susceptíveis apresentaram maior volume de fluido uterino acumulado em todos os tratamentos com menor capacidade de limpeza uterina. Foi verificada maior expressão do RNAm das interleucinas pró-inflamatórias nas éguas susceptíveis e a expressão do RNAm da interleucina-10 é...
In the equine species, uterine interleukin expression and seminal plasma cathelicidins can modulate the magnitude and persistence of the post-breeding inflammatory reaction. The objectives were to verify the presence of cathelicidins in the stallion seminal plasma and characterize the uterine immune response by pro and anti-inflammatory interleukins. Seminal plasma was obtained by semen collection of 15 stallions and submitted to SDS-PAGE and DOT-blotting tests. Three experiments were designed to study interleukin expression during baseline (T1), after seminal plasma infusion (T2) and artificial insemination (T3). Endometrial cytology and cell collection as well as ultrassonographic evaluations were performed to access interleukin mRNA through qPCR during three randomly assigned consecutive estrous cycles, 24 (M1), 48 (M2) and 72 hours (M3) after an ovulation was induced. Endometrial mRNA expression in mares resistant and susceptible to persistent post-breeding endometritis was identified for the following pro-inflammatory cytokines: IL-1β, IL-6, IL-8, IL-10 e TNF-α. Total seminal plasma protein concentrations ranged from 6.3 to 25.6 mg/mL. SDS-PAGE analysis identified the existence of three proteic bands with molecular weight similar to what was previously reported for equine cathelicidins. Evaluated stallions showed values < 339.6 ng for eCATH1, < 141.3 ng for eCATH2 and < 283.4 for eCATH3. At M2, resistant and susceptible mares could not be separated through cytology or ultrassonography, even though the presence of sperm strongly stimulates uterine inflammatory reaction. At M3, susceptible mares showed higher uterine fluid accumulation and lower uterine clearance. Pró-inflammatory mRNA expression was higher in susceptible mares and interleukin-10 mRNA expression was similar between resistant and susceptible mares

Orientador: Marco Antonio Alvarenga
Coorientador: João Pessoa Araújo Junior
Banca: Frederico Ozanam Papa
Banca: Fabiana Ferreira de Souza
Banca: Rubens Paes de Arruda
Banca: Eduardo Malschitzky
Resumo: A expressão das interleucinas no útero e das catelicidinas no plasma seminal podem ser responsáveis pela magnitude e persistência da resposta inflamatória uterina. Os objetivos desse estudo foram verificar a presença das catelicidinas no plasma seminal e caracterizar a resposta imune uterina pela expressão de interleucinas inflamatórias. O plasma seminal foi obtido por coleta de sêmen de 15 garanhões e submetido ao SDSPAGE e DOT-blotting. As éguas foram submetidas a um ciclo estral sem desafio uterino (T1), com infusão de plasma seminal (T2) e inseminação artificial (T3). Exame ultrassonográfico, citologia exfoliativa uterina e colheita de células para a expressão do RNAm das interleucinas por qPCR foram realizados durante 3 ciclos consecutivos aleatoriamente, 24 (M1), 48 (M2) e 72 horas (M3) após a indução da ovulação. A expressão do RNAm no endométrio das éguas resistentes e susceptíveis foi verificada para as interleucinas IL-1β, IL-6, IL-8, IL-10 e TNF-α. A concentração total de proteínas no plasma seminal dos garanhões variou de 6,3 a 25,6 mg/mL. O SDS-PAGE revelou a existência de 3 bandas protéicas dentro do intervalo de peso molecular previamente citado na literatura para as catelicidinas equinas. Os garanhões avaliados mostraram quantidades de eCATH1 < 339,6ng, eCATH2 < 141,3ng e eCATH3 < 283,4ng. No momento M2 não foi possível diferenciar as éguas resistentes das susceptíveis pela citologia ou ultrassonografia independente da infusão com plasma seminal, inseminação artificial ou da ovulação, mas quando há presença de espermatozoides no útero a resposta inflamatória é mais intensa. Durante o momento M3, as éguas susceptíveis apresentaram maior volume de fluido uterino acumulado em todos os tratamentos com menor capacidade de limpeza uterina. Foi verificada maior expressão do RNAm das interleucinas pró-inflamatórias nas éguas susceptíveis e a expressão do RNAm da interleucina-10 é...
Abstract: In the equine species, uterine interleukin expression and seminal plasma cathelicidins can modulate the magnitude and persistence of the post-breeding inflammatory reaction. The objectives were to verify the presence of cathelicidins in the stallion seminal plasma and characterize the uterine immune response by pro and anti-inflammatory interleukins. Seminal plasma was obtained by semen collection of 15 stallions and submitted to SDS-PAGE and DOT-blotting tests. Three experiments were designed to study interleukin expression during baseline (T1), after seminal plasma infusion (T2) and artificial insemination (T3). Endometrial cytology and cell collection as well as ultrassonographic evaluations were performed to access interleukin mRNA through qPCR during three randomly assigned consecutive estrous cycles, 24 (M1), 48 (M2) and 72 hours (M3) after an ovulation was induced. Endometrial mRNA expression in mares resistant and susceptible to persistent post-breeding endometritis was identified for the following pro-inflammatory cytokines: IL-1β, IL-6, IL-8, IL-10 e TNF-α. Total seminal plasma protein concentrations ranged from 6.3 to 25.6 mg/mL. SDS-PAGE analysis identified the existence of three proteic bands with molecular weight similar to what was previously reported for equine cathelicidins. Evaluated stallions showed values < 339.6 ng for eCATH1, < 141.3 ng for eCATH2 and < 283.4 for eCATH3. At M2, resistant and susceptible mares could not be separated through cytology or ultrassonography, even though the presence of sperm strongly stimulates uterine inflammatory reaction. At M3, susceptible mares showed higher uterine fluid accumulation and lower uterine clearance. Pró-inflammatory mRNA expression was higher in susceptible mares and interleukin-10 mRNA expression was similar between resistant and susceptible mares
Doutor

Kyoto University (京都大学)
0048
新制・課程博士
博士(医学)
甲第9461号
医博第2474号
新制||医||795(附属図書館)
UT51-2002-G219
京都大学大学院医学研究科外科系器官外科学(婦人科学産科学)専攻
(主査)教授 中畑 龍俊, 教授 丹羽 太貫, 教授 藤井 信吾
学位規則第4条第1項該当

NFkB pathway intermediates were identified in endometrium and decidua. The mRNA expression profiles of the inhibitory protein, IkBa, and an upstream kinase, TANK binding kinase 1, suggest that the pathway is activated during menstruation. Additionally, intermediates involved in the NFkB activating pathway are differentially regulated in the first trimester decidua. The pathway that mediates proinflammatory signaling (MEKK1-IKKb) to NFkB is downregulated in decidua consistent with the local immunosuppression which occurs during pregnancy. In contrast, intermediates involved in morphogenic signaling (NIK-IKKa) to NFkB are increased. This suggests a role in the expression of molecules crucial to successful pregnancy. The T47D cell line expresses high levels of progesterone receptor and was used as a cell model to study the effects of progesterone on the NFkB pathway. IkBa mRNA expression was found to be increased by progesterone while other pathway intermediates were unaffected by progesterone over the timecourse investigated. CD40 is a proinflammatory signaling molecule that activates NFkB. CD40 was detected in the perivascular region of endometrium. Previously, it has been reported that chemokine expression is upregulated in this region premenstrually and the detection of CD40 in this area suggests a role in the control of inflammatory mediator expression during menstruation. Messenger RNA expression for CD40 and its ligand, CD40L, was increased in decidua suggesting a role similar to that of NIK-IKKa. Secretory leukocyte protease inhibitor (SLPI) is an anti-inflammatory and antimicrobial molecule that also has inhibitory effects on the NFkB pathway. SLPI was localized to the glandular epithelium of endometrium from the mid-late secretory phase and was also detected in first trimester decidua. SLPI may provide antimicrobial protection at the time of implantation and during pregnancy. The detection of NFkB pathway intermediates, the CD40-CD40L system and SLPI in human endometrium supports a role for interfacing control mechanisms in the regulation of inflammatory mediators in the uterus.

Thesis (Ph. D.)--University of North Carolina at Chapel Hill, 2007.
Title from electronic title page (viewed Mar. 26, 2008). "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Pathology and Laboratory Medicine, School of Medicine." Discipline: Pathology and Laboratory Medicine; Department/School: Medicine.

The endometrium undergoes recurrent cycles of dynamic remodelling, involving breakdown and scarless repair, proliferation and differentiation, including decidualisation of the stroma, during the menstrual cycle. Extensive studies have characterised how the steroid hormones oestrogen and progesterone acting via their nuclear receptors coordinate these remarkable changes. Although a few previous studies have postulated a role for androgens the impact of androgens on endometrial function remains understudied. The studies described in this thesis aimed to 1) identify cellular processes, pathways and networks regulated by androgens in human androgen receptor-positive endometrial stromal cells (hESCs), 2) investigate the potential for regulation and determine the regulation of putative dihydrotestosterone (DHT)-regulated gene expression by androgen in hESCs, 3) investigate the expression and regulation of putative androgen-regulated genes in the human endometrium across the menstrual cycle and in early pregnancy and 4) explore the role of androgens in modulating metformin-induced gene expression associated with decidualisation of hESCs. Analysis of data from a whole genome array conducted previously in the laboratory using primary hESCs treated with DHT for 2 or 8 hours identified time dependant putative androgen-regulated mRNAs (34 and 268 genes, respectively). Thereafter, all work was completed by the author. Gene ontology and functional based bioinformatic analyses of the putative androgen-regulated gene sets revealed potential androgen regulation of a variety of cell processes, pathways and networks including those associated with gene transcription, signal transduction pathways (such as phosphatidylinositol, oestrogen receptor alpha (ERα) and Wnt signalling), cancer pathways, metabolism, cell cycle, development, apoptosis/survival. In addition, various transcription factors (e.g. AR, c-Myc, SP1, ERα, p53, E2F1, RUNX2, CREB1 and STAT3) were associated with androgen regulation in hESCs. Consensus androgen receptor binding sites were identified in the promoter sequences of 18 genes by transcription factor binding site sequence analysis. Direct DHT regulation of ten of 15 of these genes was validated in endometrial stromal cells using qRTPCR. Of these genes, RGS2, SIK1, and SNCAIP mRNAs were confirmed as DHT-regulated in hESCs by use of an AR inhibitor (flutamide) and in addition, were not found to be regulated by oestradiol. Discovery bioinformatics predicted these genes may interact in a gene network involving AR and the cAMP transduction pathway. Expression of the 15 putative androgen-regulated genes was confirmed by qRTPCR in intact human endometrial tissue (13 novel) and 9 of these genes were regulated in association with decidualisation i.e. either in the secretory phase, the time at which decidualisation begins and/or in first trimester decidua. Protein expression of RGS2, SIK1 and Synphilin-1 (encoded by SNCAIP) was confirmed by immunohistochemistry in endometrial tissues and protein expression also appeared greater in decidua. Regulation of putative androgen-regulated gene expression by decidualisation was confirmed in 4 out of 8 genes by employing a model of reduced in vivo decidualisation i.e. decidua from ectopic pregnancies. Regulation of 5 out of 7 genes was confirmed in decidualised hESCs (RGS2, SIK1, SLC6A6, SNCAIP and AXIN2) but expression of these genes was not altered by DHT inclusion during decidualisation. Finally, only a high metformin concentration enhanced hESC decidualisation and putative androgen-regulated gene expression (4 genes) in decidualised hESCs. In comparison, in the presence of DHT, a lower clinically relevant metformin concentration (100μM) did enhance decidualisation marker expression but did not alter expression of putative androgen-regulated genes. In summary, these studies have revealed new insights into androgen action in the human endometrium. Studies in hESCs 1) predicted the pathways and interacting transcription factor regulatory networks that may be androgen-dependent in this cell type, these were associated with cell differentiation, apoptosis and proliferation, 2) identified novel putative androgen-regulated genes expressed in hESCs and in endometrial tissues, 3) showed putative androgen-regulated genes are regulated by DHT (possibly via AR) in endometrial stromal cells, some of which are also regulated in association with decidualisation and 4) showed that androgens may enhance decidualisation during exposure to the commonly used drug metformin. Collectively, these new findings support a physiological role for androgens in endometrial function and provide a series of new avenues for further studies of the regulation of differentiation and proliferation.