Le pronostic de certains cancers s’est considérablement amélioré avec l’arrivée sur le marché des anticorps thérapeutiques. Devant l’essor de ces nouveaux médicaments associé à l’identification de nouveaux biomarqueurs, de nouvelles perspectives émergent pour l’imagerie moléculaire in vivo. En effet, disposer de nouveaux traceurs moléculaires spécifiques de ces biomarqueurs permettra de caractériser l’hétérogénéité des cellules cancéreuses, de suivre l’expression de ces marqueurs au cours de l’évolution de la tumeur, mais également de suivre l’efficacité d’un traitement sur la régression tumorale du patient. Pour répondre à cette évolution de diagnostic moléculaire in vivo, il convient de développer de nouvelles sondes moléculaires. L'objectif de ma thèse répond à ce nouveau besoin avec l'ingénierie et le marquage d'un format d'anticorps recombinant adapté à l'imagerie in vivo : le diabody 12G4 dirigé contre le récepteur de l’hormone antimüllérienne (AMH), marqueur de certains cancers de la sphère génitale. / Prognosis of cancers dramatically improved with the development on the market of therapeutic antibodies. With the increase of these new biodrugs, associated with the identification of new biomarkers, new opportunities emerge for the in vivo molecular imaging.. Indeed, to use new molecular tracers specific of tumoral biomarkers will allow to study and characterize the cancer heterogeneity, to monitor the expression of these markers during the tumor evolution, but also to check the treatment effectiveness on patients’ tumoral regression. To answer this evolution of in vivo molecular diagnosis, it is favorable to develop new molecular probes. The aim of this thesis answers this new need with the engineering and labeling of a new recombinant antibody format suitable to in vivo imaging : the 12G4 diabody directed against the type II human receptor for the anti-Müllerian hormone, a biomarker of some cancers of the genital area.

Introdução. A Síndrome Pré-menstrual (SPM) é caracterizada por sintomas de estresse somáticos e comportamentais que se desenrolam logo após a ovulação, atingindo um pico máximo próximo à data da menstruação e desaparecendo em média em até quatro dias após o início do sangramento. A presença da formação do corpo lúteo seria necessária para a presença dos sintomas, contudo essa dinâmica dos hormônios da fase lútea associados à SPM é pouco conhecida. O hormônio antimülleriano (HAM) é um dos melhores marcadores de declínio da capacidade reprodutiva das mulheres. Até o momento o mesmo não foi correlacionado a sintomas da SPM. A percepção de piora dos sintomas da SPM na perimenopausa é sugerida, existindo a possibilidade de que com a menor função ovariana (redução do HAM) exista uma exacerbação da SPM durante essa fase da vida reprodutiva da mulher. Objetivo: avaliar o padrão de SPM em mulheres com idade superior a 35 anos e correlacionar seus sintomas com a redução de HAM. Métodos: Estudo observacional, transversal, com uma amostra de conveniência composta por mulheres recrutadas através da mídia local. O questionário Registro Diário da Intensidade de Problemas (DRSP) de 2 meses foi utilizado para o diagnóstico de SPM. As participantes (N = 101) foram avaliadas em seus perfis antropométricos (peso, altura, índice de massa corporal) e social (paridade, nível educacional, uso de contracepção oral). Depois de completar o questionário DRSP por 2 meses, as mulheres retornaram no período prémenstrual para a coleta do sangue. Entre as participantes que utilizaram contracepção hormonal, foi solicitado um período de wash out de um mês para a coleta de sangue e dosagem de HAM no soro. O HAM foi analisado usando um kit comercial ELISA, de acordo com as instruções do fabricante. A análise estatística foi realizada na versão SPSS 18.0. O nível de significância adotado para todas as análises foi estabelecido em 5%. Resultados: A análise de HAM mostrou uma redução significativa após a idade de 35 anos, e não foi relacionada à severidade dos sintomas de SPM. Os sintomas físicos na fase lútea, mas não os emocionais, foram maiores em mulheres com mais de 35 anos de idade. A análise de Spearman mostrou uma correlação positiva entre um grupo de ≥35 anos e uma correlação negativa entre o uso de contracepção oral com piora dos sintomas físicos da SPM. Conclusão: este estudo mostrou que não houve piora dos sintomas emocionais entre as mulheres com SPM após os 35 anos de idade, apenas piora dos sintomas físicos, que necessitam uma investigação mais aprofundada. Não há relação entre os sintomas físicos e emocionais com níveis de HAM nas mulheres com SPM. / Background. Premenstrual Syndrome (PMS) is characterized by somatic and behavioral stress symptoms that take place soon after ovulation, reaching a peak near the time of menstruation and disappears on average within 4 days after the start of the bleeding. The presence of the corpus luteum formation would be needed for the presence of symptoms, however, these dynamics luteal phase of hormones associated with PMS is not well known. The anti-müllerian hormone (AMH) is one of the best markers of decline in women's reproductive capacity, and AMH has not been correlated with PMS symptoms. The perception of worsening of PMS symptoms in perimenopause is suggested, with the possibility that with less ovarian reserve (reduction of AMH) there is an exacerbation of PMS during this phase of the woman's reproductive life. Objective: To evaluate the pattern of PMS in women aged over 35 years, and correlate its symptoms to the reduction of AMH. Methods: It was an observational, cross-sectional study with a convenience sample composed of women recruited through local media. The 2-months DRSP questionnaire was used for PMS diagnosis. The participants (N=101) were evaluated in their anthropometric (weight, height, body mass index) and social (parity, educational level, use of oral contraception) profiles. After completing the DRSP questionnaire for 2 months, women returned in the pre-menstrual period for serum collection. Among participants using hormonal contraception, a one-month washout period was requested for serum collection and AMH dosage. AMH was analyzed using an ELISA commercial kit, according the manufacturer’s instructions. Statistical analysis was carried out in SPSS version 18.0. Significance level adopted for all analyzes was set at 5%. Results: AMH analysis showed a significant reduction after the age of 35 years, and it was not related to PMS symptoms severity. Physical symptoms in luteal stage were greater in women aged over 35 years, emotional symptoms were equal in both ages. Spearman’s analysis showed a positive correlation between ≥ 35years group and a negative correlation between the use of oral contraception with PMS physical symptoms worsening. Conclusion: this study showed that there is no worsening of emotional symptoms among women with PMS after 35 years of age, with a greater physical complaint, that needs further investigation. There is no relation between the physical and emotional symptoms with AMH levels in PMS women.

Les téléostes, le clade possédant le plus d’espèces parmi les vertébrés, emploient une saisissante diversité de mécanismes de détermination du sexe, incluant des mécanismes génétiques et environnementaux. Des études récentes ont identifié de nombreux nouveaux régulateurs génétiquesdu développement sexual des poissons et ont introduit la notion de “terrain de jeux évolutif”. Le but de ce projet de thèse est de donner une vue complète des dynamiques évolutives des déterminants du sexe dans un ordre de téléostes, les Esociformes. Dans notre espèce focale, Esox lucius, nous avons identifié une duplication d’un membre de la famille des TgfB – une famille qui a émergé en tant que fondamentale dans la régulation du sexe chez les téléostes – comme MSD (gène controlant la détermination du sexe). Nous avons obtenu des preuves fonctionnelles du rôle de ce gène en tant que MSD, de son interaction avec des facteurs environnementaux, ainsi que des nouvelles informations sur les processus évolutifsCe gène est perdu dans une population de la même espèce en Amérique du Nord, mais il est conservé parmi les autres Esociformes. En parallèle, d’autres systèmes de détermination du sexe ont été identifiés dans des espèces proches. De plus, dans des clades plus distants comme Umbra et Dallia, le MSD d’Esox n’est pas présent, et d’autres mécanismes implicants de nouveaux MSD ont été identifiés, complefixiant l’histoire évolutive des MSD dans ce groupe, qui reflète la plasticité génétique observée dans les téléostes en général. / Teleost fishes, the most species-rich clade among vertebrates, employs an astonishing diversity of sex-determining (SD) mechanisms, including environmental and genetic systems. Recent studies identified many new genetic regulators in fish sexual development that lead to the notion of an 'evolutionary playground'.This thesis project aims to provide a complete picture of the evolutionary dynamic of SD systems within a small order of teleost, Esociformes. In our focal species Esox lucius, we idenWhile the gene is lost in another population of the same species rapidly possibly during post-glacial recolonization process, it is well conserved among different species. Meanwhile, additional transition of SD system have also been identified in a sister Esox species. Moreover, in the most distant genera Dallia and Umbra, the Esox master SD gene is not present and we found different SD mechanisms with novel SD genes that adds additional layers of complexity to this group, which mirrors the observed high genetic plasticity in teleost SD

Introdução. A Síndrome Pré-menstrual (SPM) é caracterizada por sintomas de estresse somáticos e comportamentais que se desenrolam logo após a ovulação, atingindo um pico máximo próximo à data da menstruação e desaparecendo em média em até quatro dias após o início do sangramento. A presença da formação do corpo lúteo seria necessária para a presença dos sintomas, contudo essa dinâmica dos hormônios da fase lútea associados à SPM é pouco conhecida. O hormônio antimülleriano (HAM) é um dos melhores marcadores de declínio da capacidade reprodutiva das mulheres. Até o momento o mesmo não foi correlacionado a sintomas da SPM. A percepção de piora dos sintomas da SPM na perimenopausa é sugerida, existindo a possibilidade de que com a menor função ovariana (redução do HAM) exista uma exacerbação da SPM durante essa fase da vida reprodutiva da mulher. Objetivo: avaliar o padrão de SPM em mulheres com idade superior a 35 anos e correlacionar seus sintomas com a redução de HAM. Métodos: Estudo observacional, transversal, com uma amostra de conveniência composta por mulheres recrutadas através da mídia local. O questionário Registro Diário da Intensidade de Problemas (DRSP) de 2 meses foi utilizado para o diagnóstico de SPM. As participantes (N = 101) foram avaliadas em seus perfis antropométricos (peso, altura, índice de massa corporal) e social (paridade, nível educacional, uso de contracepção oral). Depois de completar o questionário DRSP por 2 meses, as mulheres retornaram no período prémenstrual para a coleta do sangue. Entre as participantes que utilizaram contracepção hormonal, foi solicitado um período de wash out de um mês para a coleta de sangue e dosagem de HAM no soro. O HAM foi analisado usando um kit comercial ELISA, de acordo com as instruções do fabricante. A análise estatística foi realizada na versão SPSS 18.0. O nível de significância adotado para todas as análises foi estabelecido em 5%. Resultados: A análise de HAM mostrou uma redução significativa após a idade de 35 anos, e não foi relacionada à severidade dos sintomas de SPM. Os sintomas físicos na fase lútea, mas não os emocionais, foram maiores em mulheres com mais de 35 anos de idade. A análise de Spearman mostrou uma correlação positiva entre um grupo de ≥35 anos e uma correlação negativa entre o uso de contracepção oral com piora dos sintomas físicos da SPM. Conclusão: este estudo mostrou que não houve piora dos sintomas emocionais entre as mulheres com SPM após os 35 anos de idade, apenas piora dos sintomas físicos, que necessitam uma investigação mais aprofundada. Não há relação entre os sintomas físicos e emocionais com níveis de HAM nas mulheres com SPM. / Background. Premenstrual Syndrome (PMS) is characterized by somatic and behavioral stress symptoms that take place soon after ovulation, reaching a peak near the time of menstruation and disappears on average within 4 days after the start of the bleeding. The presence of the corpus luteum formation would be needed for the presence of symptoms, however, these dynamics luteal phase of hormones associated with PMS is not well known. The anti-müllerian hormone (AMH) is one of the best markers of decline in women's reproductive capacity, and AMH has not been correlated with PMS symptoms. The perception of worsening of PMS symptoms in perimenopause is suggested, with the possibility that with less ovarian reserve (reduction of AMH) there is an exacerbation of PMS during this phase of the woman's reproductive life. Objective: To evaluate the pattern of PMS in women aged over 35 years, and correlate its symptoms to the reduction of AMH. Methods: It was an observational, cross-sectional study with a convenience sample composed of women recruited through local media. The 2-months DRSP questionnaire was used for PMS diagnosis. The participants (N=101) were evaluated in their anthropometric (weight, height, body mass index) and social (parity, educational level, use of oral contraception) profiles. After completing the DRSP questionnaire for 2 months, women returned in the pre-menstrual period for serum collection. Among participants using hormonal contraception, a one-month washout period was requested for serum collection and AMH dosage. AMH was analyzed using an ELISA commercial kit, according the manufacturer’s instructions. Statistical analysis was carried out in SPSS version 18.0. Significance level adopted for all analyzes was set at 5%. Results: AMH analysis showed a significant reduction after the age of 35 years, and it was not related to PMS symptoms severity. Physical symptoms in luteal stage were greater in women aged over 35 years, emotional symptoms were equal in both ages. Spearman’s analysis showed a positive correlation between ≥ 35years group and a negative correlation between the use of oral contraception with PMS physical symptoms worsening. Conclusion: this study showed that there is no worsening of emotional symptoms among women with PMS after 35 years of age, with a greater physical complaint, that needs further investigation. There is no relation between the physical and emotional symptoms with AMH levels in PMS women.

Introdução. A Síndrome Pré-menstrual (SPM) é caracterizada por sintomas de estresse somáticos e comportamentais que se desenrolam logo após a ovulação, atingindo um pico máximo próximo à data da menstruação e desaparecendo em média em até quatro dias após o início do sangramento. A presença da formação do corpo lúteo seria necessária para a presença dos sintomas, contudo essa dinâmica dos hormônios da fase lútea associados à SPM é pouco conhecida. O hormônio antimülleriano (HAM) é um dos melhores marcadores de declínio da capacidade reprodutiva das mulheres. Até o momento o mesmo não foi correlacionado a sintomas da SPM. A percepção de piora dos sintomas da SPM na perimenopausa é sugerida, existindo a possibilidade de que com a menor função ovariana (redução do HAM) exista uma exacerbação da SPM durante essa fase da vida reprodutiva da mulher. Objetivo: avaliar o padrão de SPM em mulheres com idade superior a 35 anos e correlacionar seus sintomas com a redução de HAM. Métodos: Estudo observacional, transversal, com uma amostra de conveniência composta por mulheres recrutadas através da mídia local. O questionário Registro Diário da Intensidade de Problemas (DRSP) de 2 meses foi utilizado para o diagnóstico de SPM. As participantes (N = 101) foram avaliadas em seus perfis antropométricos (peso, altura, índice de massa corporal) e social (paridade, nível educacional, uso de contracepção oral). Depois de completar o questionário DRSP por 2 meses, as mulheres retornaram no período prémenstrual para a coleta do sangue. Entre as participantes que utilizaram contracepção hormonal, foi solicitado um período de wash out de um mês para a coleta de sangue e dosagem de HAM no soro. O HAM foi analisado usando um kit comercial ELISA, de acordo com as instruções do fabricante. A análise estatística foi realizada na versão SPSS 18.0. O nível de significância adotado para todas as análises foi estabelecido em 5%. Resultados: A análise de HAM mostrou uma redução significativa após a idade de 35 anos, e não foi relacionada à severidade dos sintomas de SPM. Os sintomas físicos na fase lútea, mas não os emocionais, foram maiores em mulheres com mais de 35 anos de idade. A análise de Spearman mostrou uma correlação positiva entre um grupo de ≥35 anos e uma correlação negativa entre o uso de contracepção oral com piora dos sintomas físicos da SPM. Conclusão: este estudo mostrou que não houve piora dos sintomas emocionais entre as mulheres com SPM após os 35 anos de idade, apenas piora dos sintomas físicos, que necessitam uma investigação mais aprofundada. Não há relação entre os sintomas físicos e emocionais com níveis de HAM nas mulheres com SPM. / Background. Premenstrual Syndrome (PMS) is characterized by somatic and behavioral stress symptoms that take place soon after ovulation, reaching a peak near the time of menstruation and disappears on average within 4 days after the start of the bleeding. The presence of the corpus luteum formation would be needed for the presence of symptoms, however, these dynamics luteal phase of hormones associated with PMS is not well known. The anti-müllerian hormone (AMH) is one of the best markers of decline in women's reproductive capacity, and AMH has not been correlated with PMS symptoms. The perception of worsening of PMS symptoms in perimenopause is suggested, with the possibility that with less ovarian reserve (reduction of AMH) there is an exacerbation of PMS during this phase of the woman's reproductive life. Objective: To evaluate the pattern of PMS in women aged over 35 years, and correlate its symptoms to the reduction of AMH. Methods: It was an observational, cross-sectional study with a convenience sample composed of women recruited through local media. The 2-months DRSP questionnaire was used for PMS diagnosis. The participants (N=101) were evaluated in their anthropometric (weight, height, body mass index) and social (parity, educational level, use of oral contraception) profiles. After completing the DRSP questionnaire for 2 months, women returned in the pre-menstrual period for serum collection. Among participants using hormonal contraception, a one-month washout period was requested for serum collection and AMH dosage. AMH was analyzed using an ELISA commercial kit, according the manufacturer’s instructions. Statistical analysis was carried out in SPSS version 18.0. Significance level adopted for all analyzes was set at 5%. Results: AMH analysis showed a significant reduction after the age of 35 years, and it was not related to PMS symptoms severity. Physical symptoms in luteal stage were greater in women aged over 35 years, emotional symptoms were equal in both ages. Spearman’s analysis showed a positive correlation between ≥ 35years group and a negative correlation between the use of oral contraception with PMS physical symptoms worsening. Conclusion: this study showed that there is no worsening of emotional symptoms among women with PMS after 35 years of age, with a greater physical complaint, that needs further investigation. There is no relation between the physical and emotional symptoms with AMH levels in PMS women.

O hormônio anti-Mülleriano (AMH) é um marcador de reserva ovariana e função testicular. A aplicação clínica desse hormônio requer padronização adequada de valores de referência, de acordo com o imunoensaio utilizado. Os objetivos deste estudo foram: validar o imunoensaio AMH Gen II (Beckman Coulter Company, TX, USA), estabelecer valores normais de AMH em homens e mulheres saudáveis, avaliar a influência do uso de contraceptivos hormonais, tabagismo e índice de massa corpórea (IMC) sobre os valores de AMH e verificar as concentrações séricas desse hormônio em pacientes com síndrome de Turner (ST), síndrome dos ovários policísticos (SOP) e em meninos com criptorquidismo submetidos ao teste de estímulo com rhCG (gonadotrofina coriônica humana recombinante). A validação do ensaio AMH Gen II foi realizada utilizando protocolo simplificado, conforme as recomendações do Clinical and Laboratory Standards Institute. Para estabelecer valores normais de AMH, 133 mulheres e 135 homens saudáveis foram selecionados prospectivamente. Além disso, 66 pacientes com ST, 29 com SOP e 31 com criptorquidismo foram estudados. A sensibilidade analítica e funcional do ensaio AMH Gen II foram 0,02 e 0,2 ng/mL, respectivamente. Os coeficientes de variação intra e interensaio variaram entre 5,2% - 9,0% e 4,6% - 7,8% em diferentes concentrações, respectivamente. A linearidade, paralelismo e estabilidade das amostras apresentaram percentual de recuperação entre 80% - 120%. O ensaio AMH Gen II correlacionou-se fortemente com o ensaio Immunotech anteriormente utilizado (r = 0,9; p < 0,001). No sexo feminino, os valores séricos de AMH demonstraram declínio progressivo com o aumento da idade (r = -0,4; p < 0,001). Por outro lado, os valores de AMH não diferiram entre usuárias e não usuárias de contraceptivos hormonais, entre tabagistas e não tabagistas, bem como entre mulheres obesas e não obesas. No sexo masculino, a idade não influenciou as concentrações séricas de AMH; no entanto, os valores de AMH foram significativamente reduzidos com o aumento do IMC (r = -0,3, p = 0,008), contudo, esses valores permaneceram dentro do intervalo observado no grupo de indivíduos com peso normal. Todas as pacientes com ST apresentaram valores indetectáveis de AMH. Em contraste, na SOP as concentrações de AMH foram significativamente maiores do que nos controles femininos, mas com 76% de sobreposição entre os dois grupos. Em meninos com criptorquidismo, a concordância entre os valores basais de AMH e a testosterona pósestímulo com rhCG foi de 74%, houve correlação positiva significativa entre os valores basais de AMH e a testosterona pós-estímulo (r = 0,5; p < 0,001). Em conclusão, o ensaio AMH Gen II é confiável para a determinação sérica do AMH. Valores normativos de AMH apresentaram ampla faixa de normalidade em ambos os sexos. Valores reduzidos de AMH em homens obesos encontraram-se dentro da faixa de indivíduos normais. Na ST, o potencial uso do AMH para avaliar a reserva ovariana exige estudos longitudinais. A utilização do AMH como um critério diagnóstico da SOP deve ser associada aos critérios já estabelecidos. Em meninos com criptorquidismo, um AMH normal provê informação útil sobre a função testicular, mas não exclui a necessidade do teste de estímulo com rhCG e pacientes com resultados discordantes necessitam de acompanhamento clínico / The anti-Müllerian hormone (AMH) is a marker of ovarian reserve and testicular function. The clinical application of this hormone requires proper standardization of reference values according to the immunoassay used. The aims of this study were: to validate the AMH Gen II immunoassay (Beckman Coulter Company, TX, USA), to establish reference AMH values in healthy men and women, the influence of hormonal contraceptive use, smoking and body mass index (BMI) on the values of AMH and to check serum concentrations of this hormone in patients with Turner syndrome (TS), polycystic ovary syndrome (PCOS) and in boys with cryptorchidism underwent stimulation test rhCG (recombinant human chorionic gonadotropin).The validation of the AMH Gen II assay was performed using simplified protocol following the recommendations of the Clinical and Laboratory Standards Institute. To establish reference AMH values, 133 healthy women and 135 men were prospectively selected. In addition, 66 patients with TS, 29 with PCOS and 31 with cryptorchidism were studied. The analytical and functional sensitivity of the AMH Gen II assay was 0.02 and 0.2 ng / mL, respectively. Intra and inter-assay coefficients of variation in different concentrations ranged between 5.2-9.0% and 4.6-7.8%, respectively. The linearity, parallelism and stability studies showed recovery % between 80 and 120%. The AMH Gen II assay strongly correlated with the previously used Immunotech assay (r = 0.9, p < 0.001). In females, serum AMH showed a progressive decline with increasing age (r = -0.4, p < 0.001). On the other hand, AMH values did not differ between users and nonusers of hormonal contraceptives, smokers and nonsmokers, obese and non-obese. In males, age did not influence the levels of serum AMH, however, AMH values were significantly reduced with increasing BMI (r = -0.3, p = 0.008), but, these values were within the range observed in the group of subjects with body weight. All TS patients showed undetectable AMH levels. In contrast, in PCOS AMH values were significantly higher than in women controls, but with overlap of 76% between the two groups. In boys with cryptorchidism, the concordance between baseline AMH and testosterone after stimulation with rhCG was 74%, there was a significant positive correlation between baseline AMH values and testosterone values after stimulation (r = 0.5, p < 0.001). In conclusion, AMH Gen II assay is reliable for determining the serum AMH. Reference AMH values showed a wide range in both sexes. Reduced values of AMH in obese men were within the range of normal individuals. In ST, the potential use of AMH to assess ovarian reserve requires longitudinal studies. The use of AMH as a diagnostic criterion of SOP must be based on the association with the criteria already established. In boys with cryptorchidism, a normal AMH provides useful information on testicular function, but does not exclude the need for stimulation test rhCG, patients with discordant results require follow-up

Polycystic ovary syndrome (PCOS) is a complex clinical condition characterized by hyperandrogenism and chronic oligo/anovulation. Infrequent ovulation and metabolic alterations in women with PCOS are associated with subfertility and probably increased miscarriage rates compared with normal fertile women. The overall risk of developing type 2 diabetes and impaired glucose tolerance (IGT) is three- to sevenfold higher in PCOS women, and the onset of glucose intolerance seems to occur at an earlier age than in healthy controls. Women with PCOS also have several risk factors for cardiovascular disease, although it is unclear whether they actually experience more cardiovascular events than other women. Very few studies assessing the long-term reproductive and metabolic consequences in older women with previously confirmed PCOS have been conducted. In this long-term follow-up of women with PCOS, 84 women with a diagnosis of PCOS between 1987 and 1995 and age at the follow-up &gt; 35 years and an age-matched population-based group of control women participated. Data on reproductive outcome, ovarian reserve, endothelial function, insulin sensitivity and beta-cell function were collected. According to our results most women with PCOS had given birth and the rate of spontaneous pregnancies was relatively high. The rate of miscarriages was not increased in PCOS patients and the ultrasound findings together with increased levels of anti-müllerian hormone suggested that their ovarian reserve is superior to women of similar age. PCOS women displayed signs of endothelial dysfunction, but this was largely due to the increased prevalence of independent risk factors for cardiovascular disease such as increased BMI, triglycerides and blood pressures. IGT and type 2 diabetes occurred more often in PCOS women. Free androgen levels and beta-cell function decreased over time whereas insulin sensitivity remained unchanged. Obesity at young age and progressive weight-gain rendered them more prone to be insulin resistant at the follow-up. Beta-cell function was increased in PCOS women in comparison with control subjects but declined over time. Independent of PCOS phenotype at the index assessment and persistence of PCOS symptoms at the follow-up investigation, premenopausal women with PCOS had lower insulin sensitivity and increased beta cell function in comparison with control subjects. Conclusion: The long-term reproductive outcomes of PCOS are similar compared to women with normal ovaries. Although symptoms and androgen levels are normalized over time, women with PCOS continue to display reduced insulin sensitivity and increased beta-cell function and they also have an increased risk of IGT and type 2 diabetes.

Treatment success in IVF-ICSI is mainly limited by female age, but differences in ovarian reserve (OR; the remaining pool of oocytes and their quality) between individuals modify treatment prerequisites among women of similar age. OR may be assessed by OR tests (ORTs). The main aims of this work were to study menstrual cycle length (MCL), basal levels of circulating gonadotrophins, antral follicle count (AFC) and serum Anti-Müllerian hormone (AMH) levels and their associations with and prognostic capacities regarding IVF-ICSI outcome in large cohorts of unselected women. Age-adjusted MCL was positively and linearly associated with pregnancy rates (PRs), live-birth rates (LBRs) and ovarian response to controlled ovarian hyperstimulation. An MCL of &gt;34 days almost doubled the LBR compared with an MCL of &lt;26 days. The grouped variable ‘combined FSH and LH levels’ was superior to both individual gonadotrophin levels and the LH:FSH ratio. The highest mean PR was seen in connection with a combination of FSH &lt;6.7 U/l with LH &gt;4.9 U/l; PRs were lowest when FSH-LH levels were opposite to this (high-low) and intermediate when FSH-LH levels were low-low or high-high. Associations with LBR and ovarian response were similar as those for PR. AFCs and serum AMH levels were positively and log-linearly associated with PR, LBR and ovarian response. Success rates levelled out above AFC 30 or AMH 5 ng/ml. Treatment outcome was superior among women with polycystic ovaries. Among the studied ORTs, logAFC and logAMH concentration correlated most strongly. After multivariate testing, entering all studied ORTs, AMH and female age remained independently associated with LBR. AMH + AFC + age predicted both poor and excessive ovarian responses with high accuracy. Adjusting for age and oocyte yield, all ORTs remained significant for LBR, implying that ORTs also capture information on oocyte quality. In conclusion, measures of OR are strongly associated with PR, LBR and ovarian response in a log-linear fashion, and partly reflect oocyte quality. The OR spectrum is continuous, from small ‘oligofollicular’ ovaries (the low extreme) to polycystic ovaries (the high extreme). Among the studied ORTs, AMH together with age provide the most powerful basal estimate for IVF/ICSI outcome.

Abstract Anti-Müllerian hormone (AMH) was identified originally in connection with its role in male sexual differentiation. In females, AMH is secreted by ovarian granulosa cells of growing follicles and its serum levels correlate well with the remaining number of follicles, thus reflecting ovarian reserve. The aim of this study was to explore the expression and secretion of AMH in human ovarian development and in various disorders resulting in decreased reproductive function. In fetal ovaries, AMH expression was found to be initiated at midgestation and it increased gradually towards term. In serum samples from infant girls, transient postnatal activation of the pituitary-ovarian axis was found and it occurred later in premature infants, reflected in lower serum AMH levels and lower numbers of growing follicles. This immaturity resulted in insufficient feedback from ovary to pituitary and may explain the higher levels of follicle-stimulating hormone (FSH) in these girls. Ovarian follicle reserve is typically diminished in women with primary ovarian insufficiency (POI). Assay of serum AMH may be used to identify women with POI and existing follicles, as women with FSH receptor mutation who were known to have follicles had serum AMH levels in low normal range and girls/women with Turner syndrome mosaicism had higher serum AMH levels compared with those with other karyotypes. The diagnostics and follow-up of ovarian granulosa cell tumors (GCTs) are challenging as a result of the rarity of the disease and late possible recurrences. Assay of serum AMH combined with assay of the currently used marker inhibin B was shown to improve follow-up of GCTs in individual patients, compared with serum inhibin B measurement alone. The analyses revealed that AMH and inhibin B assays perform similarly in detecting macroscopic tumors. Continuous use of combined hormonal contraceptives inhibited ovarian activity independently of the administration route, as serum AMH levels decreased significantly and similarly during oral, transdermal or vaginal administration. The decrease of serum AMH levels indicates that AMH is secreted partially from FSH-dependent follicles. The study provides novel information on AMH secretion in ovarian development, and the use of AMH assay in assessing ovarian reserve and detecting ovarian disorders such as ovarian insufficiency and GCTs. / Tiivistelmä Naisen munasarjassa munarakkuloiden granuloosasolut erittävät anti-Müllerian hormonia (AMH), ja sen pitoisuudet seerumissa heijastavat jäljellä olevien munarakkuloiden määrää. Tässä tutkimuksessa tarkasteltiin AMH:n eritystä ja ilmentymistä munasarjan kehityksen aikana ja erilaisissa munasarjan toiminnan häiriötiloissa. Tuloksina havaittiin, että AMH:a ilmentyy sikiökaudella munasarjassa jo keskiraskaudesta eteenpäin. Syntymän jälkeen otetuissa seeruminäytteissä todettiin ohimenevä aivolisäkkeen ja munasarjan aktivaatio, joka ilmeni myöhemmin ennenaikaisesti syntyneillä tytöillä. Heillä havaittiin vähemmän kasvavia munarakkuloita ja matalammat seerumin AMH-pitoisuudet syntymän jälkeen kuin täysiaikaisilla tytöillä. Tämä kypsymättömyys johtui todennäköisesti munasarjan riittämättömästä palautejärjestelmästä aivolisäkkeeseen, mikä voi selittää korkeammat follikkelia stimuloivan hormonin (FSH) pitoisuudet ennenaikaisesti syntyneillä tytöillä. Ennenaikainen munasarjojen toiminnan hiipuminen voi johtua esim. kromosomi- tai geenivirheestä. Naisilla, joilla on virheellinen FSH-reseptori, tiedetään olevan munarakkuloita jäljellä, ja tulokset osoittivat seerumin AMH-pitoisuuksien olevan lähes normaali näillä naisilla. Myös tytöillä ja naisilla, joilla on todettu Turnerin oireyhtymän mosaikismi, AMH-pitoisuudet olivat merkittävästi korkeammat verrattuna muihin karyotyyppeihin. Täten AMH-määrityksen avulla voidaan mahdollisesti löytää ennenaikaisesta munasarjojen toiminnan hiipumisesta kärsivien joukosta ne naiset, joilla on vielä jäljellä munarakkuloita. Munasarjan granuloosasolukasvaimen diagnostiikka ja seuranta ovat haastavia kasvaimen myöhäisen uusiutumistaipumuksen ja harvinaisuuden vuoksi. Tulokset osoittivat, että seerumin AMH-määrityksen yhdistäminen tällä hetkellä käytössä olevaan inhibiini B -määritykseen voisi parantaa yksittäisten potilaiden seurantaa makroskooppisen kasvaimen toteamisen osalta. Tutkimus osoitti hormonaalisten yhdistelmäehkäisyvalmisteiden jatkuvan käytön vähentävän munasarjan aktiivisuutta merkittävästi annostelureitistä riippumatta, koska seerumin AMH-pitoisuudet laskivat samalla tavalla ehkäisypilleriä, -rengasta ja -laastaria käyttävillä naisilla. Tutkimus toi uutta tietoa AMH:n erityksestä munasarjan kehityksen aikana sekä AMH-määrityksen käytön mahdollisuuksista munarakkuloiden määrän arvioinnissa ja eri häiriötilojen tunnistamisessa.

Introdução: A síndrome antifosfolípide é uma doença autoimune caracterizada por eventos trombóticos e/ou obstétricos adversos associados à presença de anticorpos antifosfolípides. Considera-se síndrome antifosfolípide primária (SAFP) quando não há outra doença autoimune ou inflamatória associada. Objetivos: Avaliar marcadores de reserva ovariana em mulheres com SAFP e a associação entre estes marcadores e dados clínicos, laboratoriais e anticorpo anti-corpo lúteo (anti-CoL). Métodos: Realizou-se estudo transversal em 18 pacientes com SAFP e 24 controles. A reserva ovariana foi avaliada na fase folicular precoce através das dosagens de hormônio folículo estimulante (FSH), hormônio luteinizante (LH), estradiol e hormônio anti-Mülleriano (HAM), pela técnica de ELISA, e contagem ultrassonográfica de folículos antrais (CFA). O anti-CoL foi avaliado através de immunoblot. Todas as análises foram realizadas após suspensão de contraceptivo hormonal por, no mínimo, 6 meses e retorno das menstruações. O teste t foi utilizado para comparar médias ± desvio padrão e o teste de Mann-Whitney, para comparar medianas (variação). O teste exato de Fisher foi utilizado para comparar diferenças entre variáveis categóricas. Adotou-se nível de significância de 5% (p<0,05). Resultados: A média da idade foi similar em pacientes com SAFP e controles (33,0 ± 5,0 vs. 30,4 ± 7,0 anos, p=0,19). Houve maior frequência de CFA baixa (<=10) (56% vs. 22%, p=0,04) e muito baixa (<=5) (37% vs. 9%, p=0,04) em pacientes com SAFP quando comparadas aos controles. Em relação ao HAM, observou-se tendência de maior frequência de concentração sérica reduzida (<1,0 ng/mL), baixa (<0,5 ng/mL) e muito baixa (<0,2 ng/mL) em pacientes com SAFP (p=0,08; p=0,07 and p=0,07; respectivamente). As concetrações séricas de FSH, LH e estradiol foram semelhantes em pacientes e controles (p>0,05). Não houve associação entre baixa reserva ovariana e tipos específicos de anticorpos antifosfolípides. A presença do anti-CoL foi observada apenas em pacientes com SAFP (11% vs. 0%, p=0,177) e não foi relacionada a parâmetros de reserva ovariana. Conclusões: Pacientes com SAFP apresentaram reserva ovariana diminuída, com prevalência maior do que 50%, o que reforça o aconselhamento reprodutivo e planejamento familiar / Introduction: Antiphospholipid syndrome is an autoimmune disease characterized by thrombosis and/or pregnancy morbidity associated with antiphospholipid antibodies. Primary APS (PAPS) is diagnosed when no other autoimmune or inflammatory diseases are present. Objective: to determine ovarian reserve in PAPS women and to evaluate the association between ovarian reserve tests and clinical and laboratorial parameters, and anti-corpus luteum antibody (anti-CoL). Methods: In this cross sectional study, 18 PAPS patients and 24 healthy women were evaluated at early follicular phase with measurement of follicle stimulating hormone (FSH), estradiol, and anti-Müllerian hormone (AMH), carried out by ELISA test, and sonographic antral follicle count (AFC). Serum measurement of anti-CoL was determined by immunoblot analysis. All analyses were performed after at least 6 months from the last intake of hormonal contraceptive and resumption of menstruation. Data were compared by t-test and Mann-Whitney test in continuous variables and by Fisher\'s exact test in categorical variables. The level of significance was set at 5% (p<0.05). Results: The mean age was comparable in PAPS and controls (33.0 ± 5.0 vs. 30.4 ± 7.0 years; p=0.19). Regarding ovarian reserve tests, the frequencies of low AFC (<= 10) (56% vs. 22%, p=0.04) and very low AFC (<= 5) (37% vs. 9%, p=0.04) were significantly higher in PAPS patients than controls. Trends of higher frequencies of reduced (<1.0 ng/mL), low (<0.5 ng/mL) and negligible (<0.2 ng/mL) AMH levels were found in PAPS patients (p=0.08; p=0.07 and p=0.07; respectively). FSH, LH and estradiol were similar in patients and controls. There was no association between low ovarian reserve and specific types of antiphospholipid antibodies. Anti-CoL was solely observed in PAPS patients (11% vs. 0%; p=0.177) and was not related to ovarian reserve tests. Conclusion: Women suffering from PAPS possessed reduced ovarian reserve, with prevalence greater than 50%, emphasizing fertility counseling and family planning