<p>Picropodophyllin (PPP) is an analogue of the anti tumour lignan podophyllotoxin with the unique ability to selectively inhibit the receptor of Insulin like growth factor 1(IGF-1). IGF-1 is believed to play an important part in development of the endometrium facing implantation. With PPP treated mice, studies can be made to measure gene expression from tissue of both treated and untreated mice to compare the role of IGF-1 regarding ovulation, implantation and receptivity. The aim of this study was to analyze gene expression of some steroid hormone receptors and cytokines in ovaries from mice treated with PPP. In this study, seven mice were treated with PPP at different times and tissue was collected. PCR-primers for cDNA sequences of estrogene receptor α, estrogene receptor β, progesterone receptor A, progesterone receptor B, growth hormone receptor, interleukin 1 α, interleukin 1 β, tumour necrosis factor α and androgen receptor were used. Real Time PCR was run with the samples and gene expression was measured. The results of this study showed that the inhibition of IGF-1 receptor interacted with IGF-1 which lead to altered levels of estrogene receptor alpha, progesterone receptor, growth hormone receptor and androgen receptor that can decrease ovulation. The results also showed the differences in gene products between treated and untreated samples, suggesting that IGF-1 plays an important role regarding ovulation.</p> / <p>Studier med hjälp av den selektiva insulinlika tillväxtfaktor 1 receptorn (IGF-1R) antagonisten; picropodof?phyllin (PPP), hur samspelet mellan livmoderslemhinnan och implantationsprocessen, samt hur ovulationen påverkas av insulinlika tillväxtfaktorn 1 (IGF-1) kan nu utföras. IGF-1 tros ha en viktig roll för den reproduktiva processen, där den påverkar ovulation, implantation och embryoutveckling. IGF-familjen består av tre ligander; insulin, IGF-1 och IGF-2. IGF transporteras bundet till bindarprotein (IGFBP). Medlemmarna i IGF receptorfamiljen kan binda IGF-1, IGF-2 och insulin fast med olika affinitet. PPP som är en cykloligan, är en analog från podofyllotoxin och fungerar som en syntetisk IGF-1 receptorantagonist, som selektivt inhiberar receptorns aktivitet. PPP tros även kunna nedreglera genexpression av receptorn. Tre tidigare projektarbeten har utförts på vävnader från möss injicerade med PPP. Tyngdpunkterna i dessa arbeten har legat på immunhistokemiska studier av IGF-1 i reproduktionsorgan från möss, uttryck av IGF-1, dess receptor och bindarprotein 1 i ovarier och uterus efter behandling med PPP. I denna studie användes vävnad samt cDNA från sju möss behandlade med PPP, i olika stadier av reproduktionen samt även icke behandlade möss. Studiens syfte var att med sanntids-PCR jämföra genuttryck från östrogenreceptor α och β, progesteronreceptor A och B, tillväxthormonreceptor, Interleukin 1 α och β, ’tumor necrosis’ faktor α samt androgenreceptor i vävnad från PPP-behandlade och obehandlade möss och genom de erhållna resultaten från ovarievävnaden utläsa effekten på ovulationen och från uterusvävnaden effekten på implantation och receptivitet. Studieresultaten visade att IGF-1s frånvaro gav förändrade nivåer av genprodukter, som medförde minskad ovulationen. Studien visade att IGF-1s roll vid ovulationen var väsentlig.</p>

The signalling cascade induced by the binding of neurotrophins (NGF, BDNF, NT3 and NT4) to their high-affinity tyrosine kinase receptors (TrkA, B and C) is well documented to be important for neuronal cell survival, proliferation and differentiation. Evidence has accumulated demonstrating the importance of these signalling pathways in nonneuronal tissues, including the ovary where all neurotrophins and their receptors are expressed. In the mouse, effects on ovulation have been demonstrated but the role of Trk signalling in neonatal ovary development is less clear. Previous work had found that TrkB expression is upregulated at the time of follicle formation in the mouse and transgenic mice null for the TrkB receptor demonstrate significant loss of oocytes neonatally (TrkB knockouts, KO, die shortly after birth). This thesis examines the phenotype of the TrkB KO using morphological, histological and surgical techniques with the aim being to further investigate the role of TrkB signalling in oocyte survival, and to contribute to our understanding of neonatal ovary development. The main questions addressed are: 1) what developmental defects are occurring on a morphological level that result in the phenotype of the TrkB KO; 2) can these defects be quantified; and 3) what are the longterm survival prospects for TrkB KO oocytes. Morphological assessment revealed that TrkB KO ovaries exhibit poorer follicle health than their Controls and this was confirmed by assessment of basement membrane (BM) composition. TrkB KO brain and kidney were also assessed and found to have similarly affected BM. It is well known that cells require contact with the BM to maintain survival, thus it is postulated that TrkB signalling contributes to oocyte survival through regulation of the BM. Due to the postnatal lethality of the mutation, TrkB KO ovaries were transplanted to ascertain long-term oocyte survival. Unexpectedly it was found that TrkB KO oocytes are able to survive and follicles grow as well as they do in the Control transplants. Consequently, the in vivo effect has to be indirect. It is known that oocytes in the neonatal ovary undergo an increased rate of cell death but it is not known how the cell debris is removed. A novel observation of a neonatal ovarian immune response has been made in this thesis and is postulated to be a physiological mechanism for cell debris clearance. In conclusion, this thesis has demonstrated that signalling through TrkB has an effect on regulating BM in the ovary and other organs, but that surprisingly it has an indirect effect on oocyte survival.

The layer of cells lining the outer surface of the mammalian ovary, the ovarian surface epithelium (OSE), is a constant feature throughout the dynamic tissue remodeling that occurs throughout the reproductive cycle (follicle growth, ovulation, corpora lutea formation and pregnancy). Abnormal development of these cells is responsible for 90% of all epithelial ovarian cancers in women and epidemiological studies have shown that susceptibility to ovarian cancer is negatively correlated with increasing pregnancy. Little is known about how OSE cells are affected at each stage of the cycle, so the main aim of this study was to determine how the reproductive cycle affected proliferation and degeneration of OSE cells. This study utilised three animal models each with a different type of reproductive cycle: a mono-ovular seasonal breeder (Sheep), a mono-ovular polyoestrous breeder (Cow) and a poly-ovular non human primate (marmoset) to allow comparisons to be made. Comparison of OSE proliferative activity was made in sheep and marmoset at each stage of the cycle including pregnancy and anoestrous. The bovine model was used to investigate apoptotic cell death. Proliferative activity of somatic cells within the sheep ovary was monitored throughout the reproductive cycle by detection of cell cycle markers PCNA and Ki67 using immunohistochemistry. The pattern of OSE proliferation was correlated with the pattern of follicle development at each stage (sheep and marmoset). During pregnancy cell proliferation was significantly lower in OSE and in granulosa cells, reflecting a suppression of mature follicle development during these stages whereas in cycling animals proliferation was increased. Differences in OSE proliferation were observed in relation to the local underlying tissue environment in both sheep and marmoset. Epithelial cell rupture and regeneration enhanced the hormonal mitogenic action on epithelial cells, which showed highest proliferation over corpora lutea in each animal model. To test the hypothesis that these changes are mediated by hormones or growth factors ovine OSE cells were cultured and proliferative activity monitored after treatment with several factors: fetal calf serum (FCS), follicular fluid from follicles of varying sizes, corpora lutea extracts, recombinant human IGF-1, oestradiol and progesterone. IGF alone was demonstrated to have an affect on increasing proliferation of cultured OSE cells. Levels of FSHr and LHr were monitored by quantitative real- time PCR and it was demonstrated that the concentration of gonadotrophin receptors in OSE, increased prior to and after ovulation, at which time the in vivo OSE proliferation also peaked. The in situ apoptosis index was determined in bovine tissue using TUNEL throughout the regular cycle, and at mid and late-pregnancy stages. The results showed that pregnancy induced apoptotic activity in OSE cells and up regulated the tumour suppressor gene p53. Cultured bovine OSE cells also exhibited an increased level of apoptosis following progesterone treatment. Since p53/p53 gene expression in OSE over the corpora lutea producing progesterone also increased, this progesterone-mediated apoptosis may be mediated through an up-regulation of p53 synthesis. The effect of pregnancy and low production of gonadotrophins in the regulation of OSE cell morphology and activity was further investigated in the marmoset monkey (a non-human primate) treated with GnRH antagonist and infused with BrdU to monitor proliferative activity. OSE proliferation was correlated to ovarian events (follicular growth, ovulation and luteinization) and this was suppressed during pregnancy. Inhibition of gonadotrophin secretion by treatment with a GnRH antagonist also markedly inhibited OSE proliferation. Taken together these studies support the hypothesis that pregnancy and periods of anovulation reduce proliferation of OSE cells and alter the pattern of apoptotic cell death and that this effect is independent of species and reproductive pattern. Suppression of gonadotrophins and other growth factors during pregnancy could enhance p53-mediated apoptosis of damaged and mitogenic cells arising from repeated ovulations. This effect may partly explain why increasing number of pregnancies in woman reduces the chance of epithelial ovarian cancers.

The optical design for a dual modality endoscope based on piezo scanning fiber technology is presented including a novel technique to combine forward-viewing navigation and side viewing OCT. Potential applications include navigating body lumens such as the fallopian tube, biliary ducts and cardiovascular system. A custom cover plate provides a rotationally symmetric double reflection of the OCT beam to deviate and focus the OCT beam out the side of the endoscope for cross-sectional imaging of the tubal lumen. Considerations in the choice of the scanning fiber are explored and a new technique to increase the divergence angle of the scanning fiber to improve system performance is presented. Resolution and the necessary scanning density requirements to achieve Nyquist sampling of the full image are considered. The novel optical design lays the groundwork for a new approach integrating side-viewing OCT into multimodality endoscopes for small lumen imaging.

BACKGROUND: Polycystic Ovary Syndrome (PCOS) is an endocrine disrupting disorder affecting at least 10 percent of reproductive-aged women. In many cases, women develop comorbidities such as diabetes, cardiovascular disease, and other metabolic disorders. In North America and Europe, it takes several years and multiple doctors for women to receive a diagnosis of PCOS. This results in lost time for risk-reducing interventions. Menstrual tracking applications are one potential tool to alert women of their risk for PCOS and prompt them to seek further evaluation from a medical professional. OBJECTIVE: The objective was to develop the Irregular Cycles Feature (ICF) on the mobile phone application Clue®, which generates a probability of a user’s risk for PCOS. The secondary aim was to assess the accuracy of the tool by testing the feature on virtual test subjects. METHODS: A literature review was conducted to generate a list of signs and symptoms of PCOS. Probabilities were assigned to each variable and built into a Bayesian Network. The Irregular Cycles Feature, an adaptive questionnaire, was then developed in order to detect high-risk PCOS patients. The ICF detected at risk Clue® users through self-reported menstrual cycles and answers to medical history questions. Upon completion of the questionnaire, a Result Screen is displayed to the user. The Screen is a summary of the individual’s probability of having PCOS. For each eligible user, a Doctor’s Report is also generated. This is a screen containing information regarding menstrual irregularities and a brief medical history to be used by a medical professional in order to make a final diagnosis. Both the Result Screen and Doctor’s Report disclose information about PCOS and detailed explanations for consulting a medical provider. A brief statistical validation was then performed to compare the output of the network to predictions made by a physician-scientist using a correlation coefficient, a p-value, and a Pearson’s coefficient. RESULTS: The Irregular Cycles Feature successfully predicts probability of PCOS based on eight test cases. The correlation between the network’s calculation and the assessment made by a board-certified reproductive endocrinology/infertility physician-scientist was 0.82, with a p-value of less than 0.05. The Pearson’s coefficient was calculated to be 0.69. These values indicate that the ICF made statistically significant predictions when compared to the physician-scientist. CONCLUSIONS: The ICF provides consumer-friendly ways to improve interactions between medical providers and patients. The tool can be adapted to capture other causes of menstrual irregularities and can serve as an important mechanism for drawing attention to potentially hazardous health problems. Further validation studies will be conducted to confirm the utility of the ICF with Clue® users, particularly amongst those who receive an official diagnosis from a medical professional. / 2020-06-17T00:00:00Z

Polycystic ovary syndrome is an endocrine disorder with symptoms of hyperandrogenism, insulin-resistance and anovulation in women which creates a disposition for other disorders, like Type 2 Diabetes. Drosophila Melanogaster flies have short generation, low cost-management and share several signaling pathways with humans and are therefore an excellent tools to study disease aetiology – yet no such model exists for PCOS. This thesis aimed to create such PCOS-like disease model through treatment of Drosophila Melanogaster adults and larvae with 5-α Dihydrotestosterone and/or high sugar diet. The samples were studied by measuring weight, fecundity, total glycerol concentration and Real-Time quantitative Polymerase Chain Reaction (RT qPCR) using PCOS-related orthologue genes. While previous studies have frequently studied the effects of high sugar diets on Drosophila Melanogaster – there are no previous studies that have studied the effects of 5-α Dihydrotestosterone, with or without high sugar diet, on the weight, fecundity and gene expression of Drosophila Melanogaster. Results showed that larva raised on high sugar diet had traits resembling an insulin-resistant phenotype while the fecundity of flies treated with high sugar diet was significantly lower than those treated with standard sugar diet and DHT. RT qPCR showed overexpression of the standard sugar diet and DHT groups in the investigated genes EcR, Med, TOLL, THADA and FOXO. The conclusion, based on the results, is that insulin-resistance was produced but that more studies are required before a complete PCOS-like disease model is possible in Drosophila Melanogaster.

Rainhas de Apis mellifera acasalam com 10-20 zangões. O parentesco médio entre as operárias filhas da rainha, portanto, atinge valores bastante divergentes entre operárias da mesma patrilínea (r=0,75) e operárias de diferentes patrilíneas (r=0,29), o que deve resultar em conflitos de interesse no que diz respeito à reprodução. Neste estudo abordamos esta questão sociogenética ao nível molecular, por análise de microssatélites em dois ninhos de abelhas africanizadas, envolvendo mais especificamente a influência paterna na ativação dos ovários em operárias que ficaram órfãs após perda da sua rainha. Detectamos existência de uma preferência dentre as linhagens paternas, quanto à ativação dos ovários destas operárias e a correlação do número de ovaríolos em operárias com o seu genótipo. Utilizando o primer A88 e A24 conseguimos distinguir um mínimo de 14 patrilíneas diferentes na população de operárias dos ninhos. A partir da análise de zangões filhos das rainhas dos ninhos 65 e 67, conseguimos identificar os alelos maternos. Além disso, conseguimos distinguir 20 patrilíneas diferentes em zangões filhos de operárias órfãs do ninho 65, e 14 patrilíneas diferentes em zangões filhos de operárias órfãs do ninho 67. Analisando o grau de ativação de ovários em operárias órfãs com idade de 15 e 21 dias, detectamos diferenças entre as colmeias com respeito ao status ovariano dessas. Correlacionando-se o grau de ativação do ovário e número de ovaríolos, com as patrilíneas detectadas, foi possível distinguir patrilíneas cuja ativação ovariana e número de ovaríolos foi baixo (patrilíneas com os alelos 4/1, 4/2, 4/3, 4/4 - alelos referentes aos microssatélites A88/A24), como também, se encontrou patrilínea onde ambos aspectos foram elevados (principalmente patrilínea com os alelos 2/4). E essas mesmas patrilíneas com alto status ovariano foram mais representadas após a orfandade, através de seus filhos. E por fim, através da análise estatística, pôde-se comprovar pelo teste exato de Fisher, que essas diferenças de representatividade das patrilíneas antes, e após a orfandade, não ocorreram aleatoriamente. Esses resultados mostram a alta eficiência da metodologia empregada e, possibilitaram comprovação de que há diferenças genéticas na probabilidade de operárias se tornarem poedeiras. / A honeybee queen normally mates with 10-20 drones. Mean kinship relations, therefore, attain quite distinct values when comparing kinship between workers within the same patriline (r=0,75) to the corresponding values for workers between different patrilines (r=0,29). This discrepancy results in conflicts with respect to reproductive interests. In the present study, we employed microsatellites as molecular markers to approach this sociogenetic question in two honey bee nuclei. In detail, we asked whether differences in ovary activation, previously observed in queenless workers, can be attributed to patriline differences. Additionally, we asked whether differences in ovariole numbers in worker bees also correlate with their patriline genotype. We obtained the following results: Utilizing microsatellite primer A88 and A24 we could distinguish a minimum of 14 different worker patrilines in the two nuclei. The two maternal alleles for nuclei 65 and 67 were identified from the genotypes of the queen’s sons. We could distinguish at least 20 different patrilines in the sons of queenless workers in nucleus 65, and 14 patrilines in the sons of queenless workers in nucleus 67. Analyzing the degree of ovary activation in 15 and 21-day-old queenless workers, we detected differences in ovary status between the two nuclei. When correlating the degree of ovary activation and number of ovarioles in marked queenless workers, we were able to detect patrilines whose ovary status and ovarioles number was low (patrilines with the alleles 4/1, 4/2, 4/3, 4/4 - microsatellites A88/A24), in contrast, there were also patrilines where both parameters were high (especially in patriline with the alleles 2/4). It was exactly these patrilines which produced most offspring (drones) after queen removal. And finally, by Fisher’s exact test, we could show that these representation differences before and after orphanhood did not occur randomly. These results show the high efficiency of the methodology used, and made it possible to demonstrate that there are genetic differences in the probability of workers to become egg layers.

O câncer de ovário é a quinta maior causa de óbitos relacionados ao câncer em mulheres em todo o mundo, com pacientes tendo taxas de sobrevivência extremamente baixas. MicroRNAs (miRNAs) são RNAs não codificadores em torno de 22 ribonucleotídeos que desempenham um papel crítico na regulação da expressão gênica em praticamente todos os processos biológicos. Os miRNAs induzem o silenciamento de seus alvos através da repressão de tradução ou clivagem de mRNA. Neste trabalho verificamos que o miR-450a e o miR-450b-5p, dois miRNAs pouco explorados em tumores ginecológicos, foram regulados de forma negativa no câncer de ovário e nas linhagens de câncer de colo do útero. O objetivo deste trabalho foi verificar a hipótese de que tanto o miR-450a como o miR- 450b-5p atuam como supressores tumorais em cânceres de tecido reprodutivo. Para testar esta hipótese, utilizamos ensaios in vitro e um modelo xenográfico in vivo. Usando um vetor de expressão lentiviral, miR-450a e miR-450b-5p foram superexpressos de forma estável nas linhagens A2780, OVCAR-3 e SKOV-3 (linhagens tumorais de ovário) e C33 e SiHa (linhagens tumorais de câncer cervical). A taxa de anoikis foi analisada nas células submetidas a placas de baixa aderência durante 24 horas: a superexpressão de miR-450a resultou em menor viabilidade nas linhagens SiHa, OVCAR-3, A2780 e SKOV-3. Em relação à proliferação celular, as células A2780 que superexpressam o miR-450a proliferaram a uma taxa maior em comparação com as células controle. No entanto, no ensaio clonogênico essas células apresentaram menor número de colônias que eram visivelmente maiores. Por outro lado, o miR-450b diminuiu a taxa de proliferação na linhagem SKOV-3, o que foi corroborado no ensaio clonogênico. Também foram medidas as taxas de migração e invasão celular usando ensaio transwell. A superexpressão de miR-450a ou miR-450b-5p resultou em maior potencial de migração da linha celular C33, mas menor taxa de migração e invasão nas linhagens A2780 e SKOV3. Na linhagem OVCAR-3, a superexpressão do miR-450a aumentou o número de células capazes de invadir, já o miR-450b-5p induziu uma maior taxa de migração. A superexpressão do miR-450b-5p na linhagem SiHa resultou em menor potencial de invasão em comparação com o grupo controle correspondente. Após estes ensaios in vitro, decidimos continuar nossa análise apenas com células A2780 e SKOV-3, que apresentaram um padrão mais parecido nos ensaios. No ensaio in vivo, as células A2780 transduzidas foram injetadas intraperitonealmente em camundongos imunodeficientes de 9-11 semanas e, após 4 semanas, o peso de tumores recuperados foi comparado. Neste modelo xenográfico de câncer de ovário, verificamos que ambos os miRNAs diminuíram o crescimento tumoral, o que corrobora nossos ensaios in vitro. Também verificamos que a maioria dos alvos diretos encontrados por meio do método do PAR-CLIP e aqui validados do miR-450a estão relacionados ao metabolismo energético nas mitocôndrias (TIMMDC1, ACO2, ATP5B), além dos genes PAPSS1, CITED2, MAML1 e VIM, o marcador canônico do processo de EMT. O alvo direto para miR-450b, ME1, também está relacionado ao metabolismo energético. Até o momento, nossa análise verificou que o miRNAs miR- 450a e miR-450b-5p são comprovadamente relevantes para o processo tumorigênico de tumores epiteliais de ovário e agem como supressores tumorais principalmente pela regulação de genes que atuam no metabolismo energético. / Ovary cancer is the fifth largest cause of cancer-related deaths for women worldwide, with patients presenting extremely low 5-year survival rates. MicroRNAs (miRNAs), non-coding RNAs around 22 ribonucleotides long, play a critical role in gene expression regulation in virtually all biological processes. miRNAs induce gene silencing of their targets through translation repression or mRNA cleavage. We showed that miR-450a and miR-450b-5p, two functionally underexplored miRNAs in gynecologic tumors, were downregulated in ovarian cancer and cervical cancer cell lines. The aim of this work was to verify the hypothesis that both miR-450a and miR- 450b-5p display tumor suppressor functions in reproductive tissue cancers. For this purpose, we utilized in vitro assays and an in vivo xenographic model. Using a lentiviral expression vector, miR-450a and miR-450b-5p were stably overexpressed in A2780, OVCAR-3 and SKOV-3 (ovarian cancer cell lines) and C33 and SiHA (cervical cancer cell lines). The anoikis rate was analyzed in those cells submitted to low adherence plates for 24 hours: miR-450a overexpression resulted in lower viability in SiHa, OVCAR-3, A2780 and SKOV-3 cells. Regarding to cell proliferation, A2780 cells overexpressing miR-450a proliferated at higher rate compared to the control counterpart. However, these cells presented fewer but noticeable larger colonies at the clonogenic assay. On the other hand, miR-450b decreased the proliferation rate in SKOV-3, what was corroborated in the clonogenic assay, wherein the number of colonies were reduced at the same cell line. We also measured cell migration and invasion rates using transwell assays. Overexpression of miR-450a or miR-450b-5p resulted in higher migration potential of C33 cell line but lower migration and invasion rates in A2780 and SKOV3 cell lines. In OVCAR-3 cell line, miR-450a overexpression caused an increase in number of invaded cells and miR-450b-5p overexpressed cells migrated in higher number than control group. The overexpression of miR-450b-5p in SiHa cell line resulted in lower invasion potential compared to correspondent control group. After these in vitro experiments, we decided to continue our analysis only with A2780 and SKOV-3 cells, which had the most similar pattern. In the in vivo assay, transduced A2780 cells were injected intraperitoneally in 9-11 weeks immunodeficient mice and the weight of dissected tumors were compared after 4 weeks. In this in vivo xenographic model of ovarian cancer, we verified that both miRNAs impaired tumor growth, which corroborates our in vitro assays. We also verified that most of the direct targets found through PARCLIP technique validated in this work for miR-450a are related to energetic metabolism in mitochondria (TIMMDC1, ACO2, ATP5B) besides PAPSS1, CITED2, MAML1 and VIM, the canonical marker of EMT. The direct target for miR-450b, ME1, is also related to energetic metabolism. To date, our analysis points to the scenario of miR-450a and miR-450b-5p playing functions like tumor suppressors proven to be relevant to the tumorigenic process of ovarian epithelial tumors. Additionally, they do that mostly by regulating genes responsible by the energetic metabolism.

A hiperinsulinemia e a resistência à insulina (RI) são descritas em pacientes portadoras da PCOS, independentemente do índice de massa corporal. O exercício físico parece não ser capaz de reverter completamente a RI nessas pacientes, sugerindo uma resposta sub-ótima ao estímulo do treinamento físico nessa população. Assim, o objetivo desse trabalho foi investigar os efeitos agudos do exercício aeróbio sobre a expressão de proteínas envolvidas na sinalização intracelular para captação de glicose em pacientes com PCOS comparadas a mulheres sem comorbidades e sem a PCOS (grupo CTRL). No período basal, as voluntárias realizaram avaliações do perfil lipídico, glicêmico e hormonal, capacidade física e composição corporal. Após a intervenção, as voluntárias realizaram dosagens sanguíneas para a avaliação do perfil inflamatório e glicêmico em resposta ao exercício físico. Além disso, as voluntárias foram submetidas à biopsias musculares, para analises da expressão de proteínas envolvidas na sinalização intracelular para a captação de glicose. A expressão proteica da PI3-k não demonstrou diferenças significantes em resposta à sessão aguda de exercício aeróbio no grupo PCOS. Todavia, o grupo CTRL demonstrou um aumento significante da ativação dessa proteína em resposta à sessão aguda de exercício aeróbio (p = 0.018), bem como uma tendência de diferença significante da atividade dessa proteína na condição PÓS (p = 0.073) no grupo CTRL em comparação ao grupo PCOS. A fosforilação da AS160 Thr 642 foi significante maior no grupo CTRL em resposta a sessão aguda de exercício aeróbio (p = 0.043), enquanto, uma resposta inalterada da fosforilação da AS160 Thr 642 foi observada no grupo PCOS em resposta ao exercício aeróbio agudo. O grupo CTRL apresentou um aumento da fosforilação dessa proteína significante maior que o grupo PCOS na condição PÓS (p = 0.05). Os presentes achados demonstram um prejuízo na sinalização intracelular para captação de glicose no músculo esquelético ao nível da proteína PI3-k p85 e AS160 Thr 642 em resposta ao exercício aeróbio agudo nas mulheres com PCOS comparada as mulheres CTRL. Entretanto, a translocação do GLUT-4 não está prejudicada em resposta ao exercício aeróbio agudo nas mulheres com PCOS comparada as mulheres CTRL. Esses dados sugerem que defeitos de sinalização em proteínas específicas da sinalização insulínica não impedem a efetiva translocação de GLUT-4 no músculo esquelético de pacientes com PCOS / Hyperinsulinemia and insulin resistance (IR) are described in PCOS patients regardless of body mass index. Exercise does not seem to be able to completely reverse the IR in these patients, suggesting a sub-optimal response to the stimulus of exercise training in this population . The objective of this study was to investigate the acute effect of aerobic exercise on the protein expression involved in intracellular signaling for glucose uptake in patients with PCOS compared with women without PCOS (CTRL group). At baseline, subjects underwent assessments of lipid, glucose and hormone profile, physical fitness and body composition. After the intervention, the volunteers performed blood measurements for the assessment of inflammatory markers and glycemic profile in response to acute aerobic exercise. In addition, the volunteers underwent skeletal muscle biopsies for analysis of the protein expression involved in intracellular signaling for glucose uptake. PI3-k expression showed no significant differences in response to acute exercise bout in the PCOS group. However, the CTRL group showed a significant increase in activation of this protein in response to acute exercise bout (p = 0.018) and a trend toward significant difference in activity of this protein after exercise (p = 0.073) in the CTRL group compared to the PCOS group. AS160 Thr 642 phosphorylation was significantly higher in the CTRL group in response to acute exercise bout (p = 0.043), while an unchanged response of AS160 Thr 642 phosphorylation was observed in the PCOS group in response to acute aerobic exercise. CTRL group showed an increase in phosphorylation of this larger than PCOS group significant protein in the period after the acute exercise bout (p = 0.05). The present findings demonstrate a loss in intracellular signaling for glucose uptake in skeletal muscle at the level of the protein PI3 -k p85 and AS160 Thr 642 in response to acute aerobic exercise in women with PCOS compared CTRL women. However, GLUT - 4 translocation is not impaired in response to acute aerobic exercise in women with PCOS compared CTRL women. These data suggest that specific defects of insulin signaling do not impairment the effective GLUT - 4 translocation in skeletal muscle of patients with PCOS, probably by activation of compensatory molecular mechanisms. Keywords: polycystic ovary syndrome, exercise, obesity, insulin

O estudo do díptero Rhynchosciara americana forneceu informações importantes sobre a biologia cromossômica, por apresentar cromossomos politênicos e amplificação gênica em diferentes órgãos. Dentre os órgãos que apresentam especializações do ciclo celular, o ovário é um dos mais interessantes, possuindo características únicas, como o desenvolvimento sincrônico dos folículos ovarianos e o fato de apresentar uma única e gigante célula nutridora conectada ao ovócito através de um canal citoplasmático. Une em si: meiose no ovócito, célula nutridora com poliploidia seguido de politenia e células foliculares com mitose. Células poliplóides e politênicas possuem cópias extras de DNA genômico através de repetidos ciclos de fase S sem que ocorra a divisão celular, um processo denominado endoreplicação. Os ciclos celulares, são dirigidos pela oscilação da ativação de complexos ciclina/Cdk. Estudos mostraram que a ciclina A atua em células endoreplicativas em Drosophila e a ciclina B inibe ciclos endoreplicativos induzindo a divisão celular. Torna-se relevante investigar a associação dessas moléculas reguladoras do ciclo celular com os ciclos endoreplicativos que ocorrem na ovogênese de R. americana. Perfis de expressão das ciclinas A e B foram detectados nos ovários por RT-PCR ao longo do desenvolvimento. Ensaios de incorporação do nucleosídeo timidina mostraram elevada atividade proliferativa das células foliculares para formar o folículo e o fim da atividade endoreplicativa nas células nutridoras em pupas de 4 dias. Preparações de imunolocalização proteica em ovários na fase de pupa revelaram acúmulo de ciclina A no citoplasma das células nutridoras e dos ovócitos, e acúmulo de ciclina B no citoplasma e na vesícula germinal do ovócito, atuando nos mecanismos meióticos. O estudo de proteínas relacionadas ao ciclo celular nesse modelo é importante para um melhor entendimento dos ciclos celulares incomuns presentes em diferentes órgãos de insetos. / Study of the diptera Rhynchosciara americana has provided important information about chromosome biology, for it displays polytene chromosomes and gene amplification in different organs. Among the organs that possess cell cycle specializations, the ovary is one of the most interesting ones, showing unique characteristics, such as the synchronous development of follicles and the presence of a single giant nurse cell connected to the oocyte through a cytoplasmic channel. This organ gathers: meiosis in the oocyte, nurse cell with polyploidy followed by polyteny, and follicle cells in mitosis. Polyploid and polytene cells have extra copies of genomic DNA obtained via sequential cycles of S phase not followed by cell division, a process called endoreplication. Cell cycles are driven by the oscilation in the activation of different cyclin/CDK complexes. Studies have shown that Cyclin A acts in endoreplicating cells in Drosophila and Cyclin B inhibits endoreplicative cycles, inducing cell division. Thus, it is relevant to investigate the association between these cell cycle-regulating proteins and the endoreplication cycles that occur during the oogenesis of R. americana. Expression profiles of cyclins A and B were evaluated in the ovary via RT-PCR throughout the development. Thymidine nucleoside incorporation assays showed high proliferative activity in follicle cells to build the follicle and the end of endoreplicative activity in nurse cells of 4-day-old pupae. Protein immunolocalization in ovary at the stage of pupa has shown accumulation of Cyclin A in the cytoplasm of nurse cells and oocytes, and accumulation of Cyclin B in the cytoplasm and germinal vesicle of the oocyte, acting on meiosis mechanisms. The study of proteins related to cell cycle in this model is important for a better understanding of uncommon cell cycle in different insect organ.