Submitted by Edineia Teixeira (edineia.teixeira@unioeste.br) on 2017-12-18T17:51:48Z No. of bitstreams: 2 TATIANE_MATTIA2017.pdf: 1152114 bytes, checksum: 712fc2801c953f31d127b53677f3a645 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-12-18T17:51:48Z (GMT). No. of bitstreams: 2 TATIANE_MATTIA2017.pdf: 1152114 bytes, checksum: 712fc2801c953f31d127b53677f3a645 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-02-10 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Periodontal disease is one of the most important diseases around the world. Its progress occurs due to a combination of factors which affect the sustainable tissues of teeth, and which leads to the development of periodontal pockets and to the reabsorption of the alveolar bone. This can also result in tooth loss. By considering obesity as one of the systemic diseases which pre-establishes a series of complications and as a risk factor to other chronic pathologies, several studies have suggested that this condition is also an important risk factor for the development of periodontal diseases. It is known that the periodontal disease is immune-inflammatory when there is an imbalance between the immune host responses and the causative agents, and that it may be intensified when associated to systemic factors and conditions. This study aims at evaluating the effect of obesity induced by monosodium glutamate (MSG) over the periodontal tissues of rats having induced periodontitis. In this study, 33 rats were divided, initially, into two groups which were submitted to intradermal injections in the cervical region using 1,25g/kg/day of saline solution (group CON), and 1,25g/kg/day of MSG solution (group B), in their first 5 days of life. At 70 days, the periodontal disease was induced with the placement of ligatures in first inferior premolars of both sides of animals. This served as a gingival irritation for 30 days, and provided the cumulus of bacterial plaque, and consequently, the development of periodontal disease. After this procedure, the animals were sub-divided into 4 groups: control group without ligatures (CON); control group with ligatures (CONLIG); obese without ligature (OB), and obese with ligature (OBLIG). At 100 days, the rats were weighed and measured related to the naso-anal length in order to determine the Lee index. Through the vaginal cytology, the female rats which were in proestrus in their estrous cycles were euthanized. After that, the perigonadal and retro perigonadal fats were collected and weighted. It was also collected blood from the right brain stem. The conditions of the estradiol and progesterone were determined by using specific kits. The left and right hemi mandibles were withdrawn and submitted to a radiographic and histological analysis, respectively. A sample of the gingival tissue was collected in order to dosage the cytokines. The data were analyzed and evaluated through the ANOVA and Tukey. Obesity was confirmed by Lee index and by retroperitoneal and perigonadal fat (p<0.05). At the radiographic analysis, it was verified the efficacy of the induction of periodontal diseases, showing the insertion loss in the groups submitted to the experimental periodontitis, with an alveolar bone loss smaller in the group OBLIG when compared to the group CONLIG (p<0.05). This was also proved by the histological analysis. By analyzing the gingival tissue, no differences between CONLIG and OBLIG groups were found. In the hormonal analysis, the concentrations of LH and FSH were reduced to the OB and OBLIG groups. According to the results, it is possible to suggest that the obesity model may have some protector effect over the alveolar bone loss in the cases of induction of periodontitis. Such condition also may interfere negatively in the plasmatic concentrations of some female sexual hormones / Doença periodontal trata-se de uma das doenças orais mais importantes no mundo. Sua progressão ocorre devido a uma combinação de fatores que atingem os tecidos de sustentação dos dentes, levando à formação de bolsas periodontais e indução de reabsorção do osso alveolar, podendo resultar em perdas dentárias. Considerando a obesidade como uma doença sistêmica, que predispõe o indivíduo a uma série de complicações e sendo citada como um fator de risco para várias patologias crônicas, diversos estudos têm sugerido que essa condição vem se revelando como um importante fator de risco também para o desenvolvimento da doença periodontal. Sabendo-se que a doença periodontal é imunoinflamatória, podendo ser expressa quando ocorrer um desequilíbrio entre a resposta imune do hospedeiro com o agente causador e que ela pode ser exacerbada quando associada a fatores ou condições sistêmicas, o objetivo deste estudo foi avaliar o efeito da obesidade induzida por glutamato monossódico (MSG) sobre os tecidos periodontais de ratas com periodontite induzida. Para a realização deste estudo, 33 ratas foram divididas inicialmente em dois grupos que foram submetidas a injeções intradérmicas na região cervical de 1,25g/kg/dia de solução salina (grupo CON) e 4g/kg/dia de solução de MSG (grupo OB), nos primeiros 5 dias de vida. Aos 70 dias, foi induzida a doença periodontal com a colocação de ligadura nos 1os molares inferiores de ambos os lados dos animais, que atuou como irritante gengival por 30 dias, favorecendo o acúmulo de placa bacteriana e consequente desenvolvimento da doença periodontal. Após esse procedimento, os animais foram subdivididos em 4 grupos: grupo controle sem ligadura (CON); grupo controle com ligadura (CONLIG); obeso sem ligadura (OB), obeso com ligadura (OBLIG). Aos 100 dias de vida, as ratas foram pesadas e mensurado o comprimento nasoanal para determinação do índice de Lee. Através da citologia vaginal as ratas que se encontravam em proestro do ciclo estral foram sacrificadas. Após a eutanásia, as gorduras perigonadal e retroperitonial foram coletadas e pesadas, também foi coletado sangue do tronco cerebral direito e as concentrações de estradiol e progesterona foram determinadas utilizando-se kits específicos. As hemimandíbulas direita e esquerda foram coletadas e submetidas à análise radiográfica e histológica, respectivamente, bem como a remoção de uma amostra do tecido gengival para a dosagem de citocinas. Os dados obtidos foram analisados e avaliados através dos testes Anova e Tukey. A obesidade foi confirmada pelo índice de Lee e pelas gorduras retroperitonial e perigonadal (p< 0,05). Na análise radiográfica, verificou-se a efetividade da indução da doença periodontal, mostrando a perda de inserção nos grupos submetidos a periodontite experimental, com perda óssea alveolar menor no grupo OBLIG quando comparado ao grupo CONLIG (p<0,05), fato esse também comprovado pela análise histológica. Ao analisar o tecido gengival, não foram encontradas diferenças entre os grupos CONLIG e OBLIG. Na análise hormonal, as concentrações de LH e FSH estavam reduzidas nos grupos OB e OBLIG. De acordo com os resultados encontrados, é possível sugerir que o modelo de obesidade pode exercer efeito protetor sobre a perda óssea alveolar nos casos de periodontites induzidas. Tal condição também pode interferir negativamente nas concentrações plasmáticas de alguns hormônios sexuais femininos.

Orientadores: Sophie Françoise Mauricette Derchain, Cássia Raquel Teatin Juliato / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-19T13:57:52Z (GMT). No. of bitstreams: 1 Hartman_CaioAugusto_M.pdf: 1648028 bytes, checksum: c76142fd3621c277879783c1e0cfdcb1 (MD5) Previous issue date: 2012 / Resumo: Introdução: O câncer de ovário é a mais letal das neoplasias ginecológicas e representa a quinta causa mais comum de morte por câncer em mulheres. A sobrevida é baixa e não ultrapassa 40% após 5 anos e está diretamente relacionada ao estádio da doença ao diagnóstico. Não há até hoje, nenhum método de rastreamento que se mostrou efetivo na redução da mortalidade por esta neoplasia. Entretanto, a ultrassonografia (US) e o CA-125 são muito utilizados na diferenciação das tumorações anexiais. Apesar de todos os estudos realizados com US, não existe um consenso sobre os critérios a serem utilizados na diferenciação pré-operatória destes tumores. Frente ao achado de um tumor anexial, o ginecologista deve avaliar o risco de malignidade do mesmo, para instituir um preparo pré-operatório mais adequado, prever a extensão do procedimento e suas possíveis complicações. Objetivo: avaliar os critérios ultrassonográficos de Timmerman et al. e os valores do CA-125 como indicadores de risco para malignidade em mulheres com tumores anexiais. Sujeitos e métodos: este é um estudo de corte transversal no qual foram incluídas 105 mulheres, com 112 tumores anexais. Estas foram submetidas a coleta de sangue periférico para dosagem de CA-125 e exame de ultrassonografia (US). O exame de US foi realizado utilizando descrição padronizada. Informações sobre mais de 40 variáveis morfológicas e de Doppler foram coletadas. A seguir, os tumores anexiais foram classificados segundo 5 critérios de benignidade: (B1) cisto unilocular, (B2) presença de componentes sólidos menores que 7mm, (B3) presença de sombra acústica, (B4) tumor multilocular com paredes lisas medindo menos que 100mm e (B5) ausência de fluxo ao Doppler (índice de cor 1). Os 5 critérios de malignidade foram: (M1) tumor sólido irregular, (M2) presença de ascite, (M3) presença de pelo menos 4 projeções papilíferas, (M4) tumor multilocular sólido irregular com maior medida ? 100mm e (M5) alto fluxo ao Doppler (índice de cor 4). O padrão-ouro foi considerado o resultado do exame anátomo patológico das peças cirúrgicas. Resultados: dos 112 tumores, 81 (72,3%) eram benignos e 31 (27,7%) malignos. Os critérios ultrassonográficos foram aplicáveis a 91 (81,2%) dos tumores e resultaram em sensibilidade de 90% e especificidade de 87%. Nos tumores não classificáveis segundo os critérios, utilizamos a avaliação subjetiva para classificá-los como benignos ou malignos e obtivemos sensibilidade de 66,7% e especificidade de 75%. O CA-125 apresentou sensibilidade de 69% e especifidade de 87,8%. Nos tumores ultrassograficamente classificados como benignos, tanto a idade como os valores do CA-125 não contribuíram adicionalmente para a detecção dos tumores histologicamente malignos. Já no grupo de tumores ultrassonograficamente classificados como malignos, a idade e os valores de CA-125 contribuíram significativamente para a detecção de tumores histologicamente malignos (p = 0.025). Conclusões: A maioria dos tumores pode ser corretamente classificada segundo os critérios ultrassonográficos, com sensibilidade e especificidade semelhantes a do estudo original de Timmerman. O CA-125 isoladamente apresentou menor desempenho que o US na discrimação de tumores anexiais. Quando associamos o CA-125 à idade e aos critérios ultrassonográficos em um modelo de regressão logística, obtivemos melhora no desempenho na discriminação dos tumores ultrassograficamente malignos / Abstract: Introduction: Ovarian cancer is the deadliest gynecologic neoplasm and it is the fifth leading cause of cancer-related deaths in women. The survival rate is low, not exceeding 40% after 5 years and is directly related to tumor stage at the time of diagnosis. To date there is still no screening method that is effective at reducing mortality from this neoplasm. However, ultrasonography and CA-125 are widely used in the differentiation of adnexal tumors. Despite all studies conducted with US, there is no consensus on the criteria to be used in the preoperative differentiation of these tumors. When an adnexal tumor is found, the gynecologist should assess the risk of tumor malignancy to institute a more appropriate preoperative preparation, predict the extension of the surgical procedure along with its potential complications. Objective: to evaluate ultrasound criteria of Timmerman et al. and CA-125 values as indicators of malignancy risk in women with adnexal tumors. Subjects and methods: a prospective study was conducted, including 105 women with 112 adnexal tumors. These women underwent peripheral blood collection for CA-125 measurement and ultrasound (US). US evaluation was performed by using standardized classification. Information on more than 40 morphologic and Doppler variables was obtained. Adnexal tumors were then classified according to 5 features of benign disorders: (B1) unilocular ovarian cyst, (B2) presence of solid components in which the largest solid component is < 7mm, (B3) presence of acoustic shadows, (B4) multilocular tumor with smooth walls measuring less than 100mm and (B5) no detectable Doppler flow (color score 1). The 5 features of malignancy were: (M1) irregular solid tumor, (M2) presence of ascites, (M3) presence of at least 4 papillary projections, (M4) irregular solid multilocular tumor with largest measurement ? 100mm and (M5) high color content on Doppler exam (color score 4). Histopathological analysis of surgical specimens was considered the gold standard. Results: among the 112 tumors, 81 (72.3%) were benign and 31 (27.7%) were malignant. Ultrasound criteria were applicable to 91 (81.2%) of the tumors and resulted in a sensitivity of 90% and specificity of 87%. In tumors not classifiable according to criteria, we used subjective assessment to classify these tumors as benign or malignant, obtaining a sensitivity of 66.7% and specificity of 75%. CA-125 had a sensitivity of 69% and specificity of 87.8%. In tumors sonographically classified as benign, both age and CA-125 values did not contribute any further to the detection of histologically malignant tumors. In contrast, in the group of tumors classified as malignant on ultrasound, the age and the CA-125 values contributed significantly to the detection of histologically malignant tumors (p = 0.025). Conclusions: The majority of tumors may be correctly classified according to ultrasound criteria, with sensitivity and specificity similar to those of the original study by Timmerman. Measurement of CA-125 levels alone showed a worse performance than US evaluation in discriminating adnexal tumors. CA-125 measurement associated with age and ultrasound criteria in a logistic regression model, resulted in a better performance in discriminating the sonographically malignant tumors / Mestrado / Oncologia Ginecológica e Mamária / Mestre em Ciências da Saúde

Les microARNs (miARNs), petits ARNs non codants, sont des fins régulateurs de l’expression des gènes. Les miARNs jouent des rôles essentiels dans les processus biologiques physiologiques mais aussi pathologiques. Cependant, leurs rôles durant l’ovogenèse chez les vertébrés demeurent peu documentés. D’une manière similaire aux gènes ovocytes-spécifiques, nous avons proposé l’hypothèse que les miARNs ovaire-prédominants joueraient des rôles essentiels durant l’ovogenèse et/ou le développement embryonnaire précoce. L’objectif de ma thèse était, dans un premier temps, d’identifier les miARNs ovaire-prédominants chez le médaka (Oryzias latipes). Ensuite, dans un deuxième temps, de caractériser le rôle de MiR202, un miARN gonades-prédominant chez les vertébrés. Par une analyse transcriptionnelle à grande échelle, nous avons identifié 66 miARNs ovaire-prédominants chez le médaka, qui, pour la plupart, n’ont jamais été identifiés ni chez le poisson ni dans l’ovaire. Neuf d’entre eux ont été validés par PCRq. Parmi ces derniers, 3 miARNs (MiR4785, MiR6352 et MiR729) présentent une expression ovarienne stricte. De plus, nous avons mis en évidence une isoforme de MiR202 qui est ovaire-prédominante. L’analyse de l’expression de MiR202 durant l’ovogenèse et le développement embryonnaire a montré une expression de ce dernier durant tous les stades de l’ovogenèse. Cependant, il n’est détecté que durant les premiers stades de développement embryonnaire, précédent l’activation du génome zygotique, en cohérence avec un potentiel effet maternel. Afin de caractériser la fonction de MiR202, nous avons réalisé une inactivation fonctionnelle de ce dernier chez le médaka par le système CRISPR/Cas9. La déplétion de mir202 a causé une baisse de fécondité et un arrêt du développement avant le stade une cellule. Une analyse transcriptionnelle globale des ovaires mir202-/-, nous a permis d’identifier plusieurs gènes modulés par MiR202. Parmi eux, six3, cible potentielle de MiR202, semble réguler plusieurs gènes essentiels durant l’ovogenèse ou le développement embryonnaire. Ces travaux nous ont permis d’identifier plusieurs miARNs ovaire-prédominants. Parmi eux, nous avons montré que MiR202 joue un rôle essentiel durant l’ovogenèse et sa contribution en tant que gène à effet maternel est indispensable au développement embryonnaire précoce chez le médaka. / MicroRNAs (miRNAs) are small non-codant RNAs that emerged as key regulators of gene expression. MiRNAs play important roles in both normal physiological and pathological pathways in many organisms. The involvement of miRNAs in vertebrate oogenesis remains however poorly documented. Based on the assumption that ovarian-specific or ovarian-predominant genes usually play important roles in oogenesis or early development in vertebrates, we searched for ovarian-predominant miRNAs in the medaka (Oryzias latipes) ovary, in one hand. In another hand, we studied the function of MiR202, a gonadal predominant miRNA in vertebrates. Using genome-wide expression analysis, we identify 66 miRNAs predominantly expressed in the ovary, most of them have never been described neither in fish nor in ovaries. Nine were validated by QPCR. Among them, 3 miRNAs exhibit a strict ovarian expression (MiR4785, MiR6352 and MiR729). Further, we identify a novel miR202 isomiR that exhibits an ovarian predominant expression. MiR202 expression analyses during oogenesis and early embryonic development revealed an expression in all oogenesis stages. However, it was only detected in early developmental stages before onset of zygotic genome activation (ZGA), suggesting that this MiR202 is maternally inherited in medaka. To decipher MiR202 function, CRISPR/Cas9 system was used to functionally inactivate this miRNA in medaka. Mir202 depletion causes a reduced fecundity and an early embryonic developmental arrest. Global gene expression profiling of mir202-/- ovaries revealed that many genes are regulated by MiR202. Among them, six3, that could be a putative target of MiR202, seems to be involved in the regulation pathway of many genes that are essential in oogenesis and embryonic development. During my PhD, we identify many ovarian-predominant miRNAs. Among them, we showed that MiR202 plays an essential role during oogenesis and plays a key role during early embryonic development as a maternal effect gene.

With meiosis spanning from embryonic development to the end of reproductive life in females, scientists have faced considerable limitations in studying female meiosis and the effects of toxicants on the developing oocyte. Over the last half century, various culture methods have been developed with the aim of studying the mechanisms of early ovary development, as well as for use in reproductive toxicology. However, very few of the established embryonic ovary culture systems have been used to investigate potential reproductive toxicants on the embryonic ovary, in particular when compared with the vast number of in vitro reproductive toxicity studies on the post-natal ovary. Here, a novel test compound, a topoisomerase II inhibitor: AstraZeneca Test Compound (AZTC), was used to investigate the efficacy and validity of ovarian culture methods when compared with in vivo reprotoxicity studies. AZTC was selected due to preliminary in vivo studies demonstrating its detrimental effects on spermatogenesis in male rats. AZTC targets bacterial type II topoisomerases that might have mammalian homologues involved in meiosis. Topoisomerase-II α was expressed within the female germ cells pre-natally, but became localised to the granulosa and stroma cells post-natally. This occurred both in vivo and in vitro. Ovaries from female rats exposed pre-natally to AZTC in vivo were analysed histologically and a significant increase in the number of primordial follicles was observed within the ovaries, as well as an increase in the number of unhealthy follicles. A novel mouse embryonic ovary culture system was developed by adapting, improving and bridging existing available culture techniques. The culture system supported growth of pre-meiotic mouse germ cells through prophase I of meiosis, the formation of primordial follicles and initiation of follicle growth. Cultured ovaries contained follicles at stages in comparable ratios to those in vivo and appeared morphologically normal and healthy. The culture also supported meiotic progression of oocytes to the pachytene stage, albeit with a slight delay. AZTC was used to validate the novel embryonic ovary culture by comparing the results with those from the in vivo study, where AZTC exposure had also occurred during embryonic development. Similar results were consistently observed between the in vivo and in vitro studies. In vitro effects of AZTC on the post-natal mouse ovary were also investigated, where neonatal mouse ovaries cultured with AZTC had fewer primordial follicles and more unhealthy follicles than did control ovaries. AZTC therefore demonstrated different effects when exposure occurred pre-natally vs. post-natally. The embryonic ovary culture was then used to examine the effects of another topoisomerase II inhibitor, etoposide, on the pre-natal ovary. Etoposide is a chemotherapy agent and has previously been prescribed to pregnant women. A significant reduction in the size of the follicle pool was observed in exposed cultured embryonic ovaries, where primordial and transitional follicles were targeted. Overall, establishment of post-natal culture systems have become a useful addition to in vivo reproductive toxicology studies. The embryonic ovary culture system developed here could become a valuable and powerful tool to screen potential reproductive toxicants, as well as to study the dynamics and regulation of early ovary development.

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.

In the present study, the effects of clomiphene citrate (CC) on ovulation, ovarian growth and ovarian steroidogenesis were examined. Ovulation in rats in response to PMSG was completely blocked by administration of three daily treatments of 1.0 mg CC/rat, but was restored by administration of hCG as a preovulatory LH surge substitute. When the number of treatment days was reduced to two days, 1.0 mg of CC enhanced ovulation in response to PMSG, whereas treatment for one day with the same dose of CC did not affect ovulation. The effects of CC on ovulation appear to be dose-dependent. The effects of CC on ovarian growth were similar to the effects of CC on ovulation. The ovarian growth induced by PMSG was inhibited by high doses of CC, while a lower dose had no effect. The inhibition of ovarian growth in terms of ovarian weight by a high dose of CC was restored by hCG given as a preovulatory LH surge. Treatment duration with CC appears to have an important influence on ovarian growth. Three daily treatments with high doses of CC significantly inhibited ovarian growth. However, when the number of treatment days was reduced from three to two, the opposite results were obtained in that CC significantly stimulated ovarian growth. The effects of CC on ovarian steroidogenesis in response to PMSG were dose-dependent. A higher dose of CC significantly stimulated estradiol-17β biosynthesis. Clomiphene citrate did not show any inhibitory effects on progesterone production. Progesterone production was stimulated by hCG in CC treated rats. Lower doses of CC stimulated progesterone and androgen production. Further studies on this are necessary. Histological examination of the ovary revealed that CC selectively inhibited the development of nondominant follicles. The dominant follicles were unaffected as for they were able to develop to the mature stage. These results suggest that the effects of CC on ovulation, ovarian growth and ovarian steroidogenesis are dose-dependent and affected by treatment duration. Clomiphene citrate is assumed to exert its action via a gonadotropic mechanism. / Medicine, Faculty of / Obstetrics and Gynaecology, Department of / Graduate

O objetivo do estudo foi confirmar se a redução da fecundidade dos mosquitos Aedes aegypti infectados por Plasmodium gallinaceum ocorre por morte das células foliculares dos ovários. Mosquitos infectados produzem menos ovos quando comparado aos mosquitos sadios. Uma explicação é a redução da viabilidade celular que ocorre nos ovários de fêmeas 18, 22 e 24 horas após o repasto sanguíneo infectado (RSI) como foi observado pela técnica MTT. Utilizando o acridine orange, não foi possível observar a morte das células foliculares no intervalo de 18 horas, mas 22 e 24 horas após o RSI essas células estão em morte em relação ao mesmo intervalo do repasto sanguíneo controle (RSC). A análise do DNA fragmentado foi realizada através do TUNEL. Ovários de 22 e 24 horas após RSC e RSI foram negativos nas regiões dos cortes histológicos examinados. Sendo assim, podemos concluir que, utilizando esses ensaios foi possível identificar a morte das células foliculares como uma resposta a redução da fecundidade, porém não foi possível determinar que o tipo de morte é apoptose. / The objective of this study was to confirm the hypothesis that the fertility reduction in Plasmodium gallinaceum-infected Aedes aegypti occurs by follicular cells death. A significant reduction in the number of eggs laid by infected mosquitoes was confirmed. It was observed a reduction of viable cells in 18, 22 and 24 hours PBM infected by MTT assay. It was not possible to observe cell death in ovary tissue 18 hours PBM infected, but the follicular cells showed orange color 22 and 24 hours indicating they are in death in relation to the same interval of PBM control. To determine if these cells exhibit apoptosis, we use the TUNEL which mark the fragmented DNA, a characteristic of the apoptosis process. Ovaries 22 and 24 hours PBM infected and control were negative for TUNEL marker from ovary histological preparations. Thus, we conclude that fecundity reduction occurs as a response to follicular cells death caused by P. gallinaceum infection but it was not possible to affirm if the type of follicular cells death is apoptosis.

Common features of polycystic ovary syndrome (PCOS), including hyperandrogenism, ovarian dysfunction and obesity, can be highly distressing. We compared 40 women with PCOS to women with infertility but not PCOS, and to women with neither PCOS nor infertility, on measures of depression and body image. Women with PCOS reported higher depression scores and greater body dissatisfaction (p < .001) than comparison group women. Body image was strongly associated with depression overall, even after controlling body mass. Among women with PCOS, body dissatisfaction measures and education explained 66 percent of the variance in depression, suggesting explanations of the PCOS-depression link should consider the role of potentially mediating psychosocial variables.

Cell clearance is critical for the resolution of inflammation. Defects in cell clearance can result in pathologies associated with chronic inflammation. Cells are cleared by phagocytes. These phagocytes can be either professional phagocytes whose main function is to engulf, or non-professional phagocytes, which have other tissue-resident functions but can engulf when needed. While the molecular biology of cell clearance has been heavily studied, the differences between mechanisms of engulfment by non-professional and professional phagocytes are not yet known. The study of cell clearance by non-professional phagocytes is important due to the presence of these cells in all tissues of the human body. The Drosophila ovary is a genetically tractable, in vivo model system to study engulfment by non-professional phagocytes. In response to protein deprivation, apoptosis is induced in the germline and the surrounding follicle cells engulf the dying corpses. The back and forth signaling that occurs between the dying germline and engulfing epithelial follicle cells is critical for efficient death and clearance. The germline must display eat-me signals for phagocyte recognition. The interaction of the eat-me signal and the engulfment receptor allows follicle cells to activate downstream signaling for internalization and corpse degradation. Interestingly, engulfment receptors have an additional role in driving germline death progression. The research in this dissertation focuses on the signals that drive the reciprocal signaling between non-professional phagocytes and dying cells. The results of this study demonstrate that Draper, an engulfment receptor, induces germline cell death by activating a Shark/JNK/NADPH oxidase signaling axis that is dependent on a YxxL motif. The results from this work indicate that the apoptotic machinery is activated and nurse cell nuclei are degraded in response to Draper overexpression, but Draper-induced cell death occurs independent of effector caspases, indicating that Draper induces cell death via other death pathways. An unbiased, high throughput approach to identify novel death-inducing receptors and ligands is also described. This study describes the reciprocal signaling mechanisms between engulfing and dying cells and opens up new avenues for targeting engulfment-mediated cell death. / 2021-11-08T00:00:00Z

Engulfment of dead cells by epithelial cells is crucial for the health of an organism. Defective engulfment can lead to serious conditions such as retinitis pigmentosa and asthma. In the Drosophila melanogaster ovary, protein starvation induces apoptotic cell death of germline cells, which are then engulfed by adjacent epithelial follicle cells. The follicle cells synchronously enlarge approximately four-to-five fold as they engulf the dying germline, suggesting that significant changes are required. However, the molecular changes needed to drive enlargement and engulfment by epithelial cells are still poorly understood. In this dissertation, I determined the role of integrins in engulfment, the interactions between the core engulfment machinery and the corpse processing pathway, and the roles of GTPases in engulfment by epithelial cells. First, I found that the integrin heterodimer, αPS3/βPS, becomes apically enriched and is required in the epithelial follicle cells for engulfment. αPS3/βPS is trafficked in a polarized fashion using much of the same machinery as migrating cells, suggesting similarities between engulfing and migrating cells. The canonical corpse processing pathway has been well-characterized, however, little is known about how the core engulfment machinery interacts with the corpse processing pathway. I found that the phagocytic receptor Draper is present on the phagocytic cup and early phagosomes, whereas integrins are maintained on the cell surface. Engulfment mutants fell into three distinct categories based on their specific effects on internalization and phagosome maturation, suggesting that components of the core engulfment machinery are required for distinct steps of corpse processing. Last, I investigated the roles of the Rho family GTPases during engulfment. Strikingly, Rac2 becomes induced during engulfment whereas Rac1 does not change its expression pattern. Both Rac1 and Rac2 are required for engulfment, however, Rac1 has defects in both enlargement and vesicle uptake whereas Rac2 only has defects in enlargement. Furthermore, I found that Rac1 and Rac2 may have differential effects on the Jun kinase signaling pathway, which suggests complexity in the regulatory network controlling engulfment in epithelial cells. Together, this work has provided a greater understanding of the molecular changes required within epithelial cells for proper engulfment. / 2016-12-01T00:00:00Z