Made available in DSpace on 2014-06-11T19:29:14Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-08-07Bitstream added on 2014-06-13T19:59:04Z : No. of bitstreams: 1 montechiesi_df_me_botfmvz.pdf: 745136 bytes, checksum: eafa1c97155a42da695e74f237e9a6fb (MD5) / Universidade Estadual Paulista (UNESP) / O presente estudo teve como objetivo verificar o efeito da aspiração de folículos 25mm sobre a concentração de progesterona plasmática em éguas. O crescimento folicular foi acompanhado diariamente e um único folículo ovariano foi aspirado, exceto quando uma co-dominância foi observada. Neste caso, ambos os folículos foram aspirados no mesmo momento quando atingiram o diâmetro esperado nos respectivos grupos: F ³25mm (n = 6), F ³30mm (n = 6), F ³35mm (n = 6), F pré-ovulatório (n = 6). O Grupo controle (n = 5) não foi submetido à aspiração, acompanhando-se as ovulações espontâneas. A avaliação ultrassonográfica dos ovários foi realizada a cada 24 horas iniciando-se 48 horas antes da aspiração folicular (D0 = dia da aspiração), continuando até a ovulação subsequente. Para a análise estatística, foi utilizada a análise de variância de perfil seguida do método de Tukey com nível de significância de 5% para todas as variáveis, exceto: (1) animais que responderam à aspiração folicular atingindo concentrações de 2ng/mL de progesterona; (2) presença ou ausência de estrutura lútea visualizada pela ultrassonografia, onde foi utilizado o teste do qui-quadrado. A concentração de progesterona foi >2ng/mL, entre os dias 4,0±0,4 e 7,3±0,5 após a aspiração. O intervalo entre a aspiração e a luteólise foi de 16,0±0,5 a 19,0±1,4 dias e entre a aspiração e a ovulação foi de 17,2±2,8 a 23±0,5 dias. A concentração máxima de progesterona alcançada variou entre 6,4±2,6 e 10,9±1,8ng/mL e ocorreu entre os dias 8,7±3,4 e 11,5±1,2. A aspiração folicular em todos os grupos permitiu que as células foliculares se transformassem em estrutura hormonalmente ativa, produtora de progesterona em níveis compatíveis com o diestro. / The present study aims to verify the follicles 25mm ablation effect on plasmatic progesterone concentration in mares. Follicular growth was daily accompanied and only one follicle was ablated, excepted when codominance was observed. At this case, both follicles were ablated at the same moment when achived the expected diameter for the following groups: F ³25mm (n = 6), F ³30mm (n = 6), F ³35mm (n = 6), F pre-ovulatory (n = 6). Control group (n = 5) was not submitted to ablation and the spontaneous ovulations were followed. Ultrasound evaluations of the ovaries were done every 24h beginning 48h after follicular ablations (D0 = day of ablation), continuing until subsequent ovulation. For statistical analysis, profile analysis followed by Tuckey method was used with a significance level of 5% for all variables excepted for: (1) animals that responded to follicular ablation achieving progesterone concentrations of 2ng/mL; (2) presence or absence of a luteum structure observed by ultrasound were analysed by Qui-square test. Interval between aspiration and luteolysis varied from 16,0±0,5 to 19,0±1,4 days and between ablation and next ovulation varied from 17,2±2,8 to 23±0,5 days. Maximum concentration of progesterone varied from 6,4±2,6 and 10,9±1,8ng/mL and occurred between days 8,7±3,4 and 11,5±1,2. In all groups, follicular ablation allowed follicular cells to become an active hormonal structure, which produced progesterone in concentrations similar to diestrous.

Orientador: José Carlos Peraçoli / Coorientador: Maria Teresinha de Oliveira Cardoso / Banca: Adriano Bueno Tavares / Banca: Francisco Diogo Rios Mendes / Resumo: A taxa de sucesso de ovulação com uso do citrato de clomifene é excelente, alcançando valores entre 80% e 85%, porém a taxa de gravidez é de apenas 20% a 40%. Quando o citrato de clomifene é usado como indutor de ovulação, em mulheres portadoras de síndrome dos ovários policísticos, a literatura não define qual o momento ideal de ruptura folicular que se correlaciona com sucesso de gravidez. Identificar o diâmetro de ruptura folicular de pacientes com síndrome de ovários policísticos (SOP) estimuladas com citrato de clomifeno que melhor se correlacione com ocorrência de gravidez. Estudo corte transversal realizado entre janeiro a dezembro de 2010, que incluiu 104 pacientes com ciclos ovulatórios após uso de citrato de clomifeno, acompanhadas com ultrassonografia até determinar-se o diâmetro de ruptura folicular, que foi posteriormente correlacionado com a ocorrência ou não de gravidez. Para o estudo da associação entre as variáveis aplicou-se o teste do qui-quadrado e para as variáveis quantitativas a análise de variância seguida do método de Tukey ou teste t de Student, com valor de p <0,05. No grupo de pacientes com ruptura folicular com menos de 25 mm (n: 54) a taxa de gravidez foi de 35,2% e no grupo com ruptura folicular com mais de 25 mm (50) essa taxa foi de 34%, sem diferencia estatística significativa. Quando relacionamos aleatoriamente os diferentes diâmetros de ruptura folicular com a ocorrência de gravidez também não houve diferença significativa. O presente estudo permitiu concluir que, a taxa de gravidez de mulheres portadoras de síndrome dos ovários policísticos estimuladas com citrato de clomifene, não tem correlação com o diâmetro de ruptura folicular / Abstract: The success rate of ovulation using clomiphene citrate is excellent, reaching values between 80% and 85%, but the pregnancy rate is only 20% to 40%. When clomiphene citrate is used to induce ovulation in women with polycystic ovary syndrome (PCOS), the literature does not define what the optimal timing of follicular rupture that correlates with successful pregnancy is. To identify the diameter of follicular rupture in patients with PCOS stimulated with clomiphene citrate that best correlates with the occurrence of pregnancy. Cross-sectional study conducted between January and December 2010, which included 104 patients with ovulatory cycles after the use of clomiphene citrate, followed by ultrasound to determine the diameter of follicular rupture, which was later correlated with the occurrence of pregnancy or not. To the study of the association between variables the chi-square test for quantitative variables and the analysis of variance followed by Tukey's method or Student's t test, with p <0.05 was applied. In patients with follicular rupture less than 25mm (n = 54) the pregnancy rate was 35.2%, in the group with follicular rupture with more than 25mm (50) this rate was 34%, no statistically significant difference was found. When relating the different diameters of randomly follicular rupture with the occurrence of pregnancy there was no significant difference. This study concluded that the pregnancy rate of women with PCOS stimulated with clomiphene citrate does not correlate with the diameter of follicular rupture / Mestre

The purpose of this study was to examine the relationships between age, gender and fatigue resistance, and further, to determine the relative influence that estrogen status, membrane excitability, absolute force and muscle length have over the development of fatigue. A total of 48 subjects, classified by age and gender participated in this study; 12 young males (25.3 ± 2.1 yrs.), 12 young females (23.5 ± 2.1 yrs.), 12 elderly males (71.7 ± 5.6 yrs.) and 12 elderly females (69.5 ± 4.6 yrs.). The young females were all eumenorrheic, not taking oral contraceptives and tested in the mid-follicular phase of the menstral cycle. None of the elderly females were on hormone replacement therapy. A 3 minute paradigm of intermittent maximal voluntary contractions (MVC) was used to fatigue the adductor pollicis muscle, in which 5s MVC's were alternated with 2s rest periods. In addition, maximal twitches were evoked in each 2s rest period. No gender difference in fatigability was evident between young males and young females when considering the fatigue index of the evoked twitch (FI-PT) (young males: 39.8 ± 26.7%, young females: 36.6 ± 19.0%). There was also no gender difference in fatigability found between elderly males and elderly females when considering the FI-PT (elderly males: 24.9 ± 26.6%, elderly females: 16.4 ± 48.9%). However, potentiation of the evoked twitches during fatigue may have confounded these measures. When considering the changes in voluntary force during fatigue, there was a strong trend for a gender by time interaction between young males and young females.(p=0.06), which suggests that the former were more fatigable. The trend for this gender difference was also apparent in the voluntary fatigue index (FI-MVC) (young males: 44.7 ± 10.5%, young females: 37.8 ± 14.1 %; p=0.12). Similarly, young males had a significant decrease in M-wave amplitude during the fatigue protocol and a trend for a decrease in M-wave area (p=0.08), while young females showed no significant decreases in either M-wave measure during the fatigue protocol. There was no gender difference found between the elderly males and elderly females when considering the FI-MVC (elderly males: 24.2 ± 10.7%, elderly females: 26.3 ± 14.5%). Both groups also showed small but significant reductions in theM-wave amplitude during the fatigue protocol, although M-wave area was well maintained. The fact that a strong trend for a gender difference in fatigability was found in the young subjects but not the elderly subjects, suggests that estrogen may possess fatigue resisting properties, even during short duration exercise in which glycogen depletion is not a concern. With respect to the age-related differences in fatigue, elderly males were found to be significantly more fatigue resistant than young males as indicated by the FI-MVC (p<0.01), and the significant age by time interaction during the fatigue protocol (p<0.01). In contrast, only a trend was found for an age by time interaction between the young and elderly females during the fatigue protocol (p=0.06). This trend for an age-related difference in fatigue amongst women was also reflected in the FI- MVC (p=0.13). / Thesis / Master of Science (MSc)

T follicular helper cells (T-FH) are a specialized subset of CD4 T cells that reside in B cell follicles and promote B cell maturation into plasma cells and long-lived memory B cells. During chronic infection prior to the development of AIDS, HIV-1 (HIV) replication is largely concentrated in T-FH. Paradoxically, T-FH numbers are increased in early and midstages of disease, thereby promoting HIV replication and disease progression. Despite increased T-FH numbers, numerous defects in humoral immunity are detected in HIV-infected individuals, including dysregulation of B cell maturation, impaired somatic hypermutation, and low quality of antibody production despite hypergammaglobulinemia. Clinically, these defects are manifested by increased vulnerability to bacterial infections and impaired vaccine responses, neither of which is fully reversed by antiretroviral therapy (ART). Deficits in T-FH function, including reduced HIV-specific IL-21 production and low levels of co-stimulatory receptor expression, have been linked to these immune impairments. Impairments in T-FH likely contribute as well to the ability of HIV to persist and evade humoral immunity, particularly the inability to develop broadly neutralizing antibodies. In addition to direct infection of T-FH, other mechanisms that have been linked to T-FH deficits in HIV infection include upregulation of PD-L1 on germinal center B cells and augmented follicular regulatory T cell responses. Challenges to development of strategies to enhance T-FH function in HIV infection include lack of an established phenotype for memory T-FH as well as limited understanding of the relationship between peripheral T-FH and lymphoid tissue T-FH. Interventions to augment T-FH function in HIV-infected individuals could enhance immune reconstitution during ART and potentially augment cure strategies.

Orientador: Fabiana Ferreira de Souza / Resumo: No ciclo estral das vacas ocorre uma série de eventos que se repetem até o impedimento da luteólise, pela presença do embrião no útero, que provoca alterações no organismo materno de ordens hormonais, anatômicas e comportamentais. Apesar disso, a colheita de oócitos em vacas gestantes é uma alternativa para a produção in vitro de embriões. No entanto, em vista do perfil hormonal durante a gestação, os fatores envolvidos no desenvolvimento folicular podem ser comprometidos. Em vista disso, este estudo objetivou descrever o perfil proteico do líquido folicular de vacas gestantes. Foram colhidos ovários de 36 vacas gestantes em terço inicial de gestação. O líquido folicular foi puncionado e o diâmetro de cada folículo foi mensurado pela ultrassonografia, sendo classificados em três categorias ≤6,4mm, 6,5mm a 8,9mm e ≥9. Após duas centrifugações para remoção dos componentes celulares, as proteínas totais foram mensuradas e a eletroforese unidimensional realizada sob condições desnaturantes e redutoras. As bandas foram recortadas e a digestão in gel realizada objetivando a espectrometria de massas. A concentração de estrógeno e progesterona foi mensurada no líquido folicular para determinar a viabilidade dos folículos. Foram encontradas 46 bandas de proteínas na eletroforese em 67 amostras de líquido folicular. A média da concentração ± desvio padrão da progesterona foi de 71,32 ± 80,06 ng/mL e estrógeno de 27,57 ± 30,62 ng/mL, considerando todos os folículos. Hou... (Resumo completo, clicar acesso eletrônico abaixo) / Mestre

Diss. (sammanfattning) Stockholm : Karol. inst., 2006. / Härtill 4 uppsatser.

Submitted by DIEGO SOUZA MOURA MOURA (diosmoura@yahoo.com.br) on 2016-12-09T13:03:07Z No. of bitstreams: 1 Diego Souza Moura Dissertação depois da defesa para arquivar on line.pdf: 16424035 bytes, checksum: fdfeaf59a20fcfe3cbd802420a342a7b (MD5) / Approved for entry into archive by Felipe Augusto Arakaki (arakaki@reitoria.unesp.br) on 2016-12-13T10:54:04Z (GMT) No. of bitstreams: 1 moura_ds_me_bot.pdf: 16424035 bytes, checksum: fdfeaf59a20fcfe3cbd802420a342a7b (MD5) / Made available in DSpace on 2016-12-13T10:54:05Z (GMT). No. of bitstreams: 1 moura_ds_me_bot.pdf: 16424035 bytes, checksum: fdfeaf59a20fcfe3cbd802420a342a7b (MD5) Previous issue date: 2016-03-29 / No ciclo estral das vacas ocorre uma série de eventos que se repetem até o impedimento da luteólise, pela presença do embrião no útero, que provoca alterações no organismo materno de ordens hormonais, anatômicas e comportamentais. Apesar disso, a colheita de oócitos em vacas gestantes é uma alternativa para a produção in vitro de embriões. No entanto, em vista do perfil hormonal durante a gestação, os fatores envolvidos no desenvolvimento folicular podem ser comprometidos. Em vista disso, este estudo objetivou descrever o perfil proteico do líquido folicular de vacas gestantes. Foram colhidos ovários de 36 vacas gestantes em terço inicial de gestação. O líquido folicular foi puncionado e o diâmetro de cada folículo foi mensurado pela ultrassonografia, sendo classificados em três categorias ≤6,4mm, 6,5mm a 8,9mm e ≥9. Após duas centrifugações para remoção dos componentes celulares, as proteínas totais foram mensuradas e a eletroforese unidimensional realizada sob condições desnaturantes e redutoras. As bandas foram recortadas e a digestão in gel realizada objetivando a espectrometria de massas. A concentração de estrógeno e progesterona foi mensurada no líquido folicular para determinar a viabilidade dos folículos. Foram encontradas 46 bandas de proteínas na eletroforese em 67 amostras de líquido folicular. A média da concentração ± desvio padrão da progesterona foi de 71,32 ± 80,06 ng/mL e estrógeno de 27,57 ± 30,62 ng/mL, considerando todos os folículos. Houve expressão diferenciada de proteínas nas diferentes categorias de folículos. Na espectrometria de massas foram encontradas diversas proteínas, sendo as principais apolipoproteina, angiotensinogênio, heat shock protein, glutationa e suas isoformas. / Estorus cycle of cows has a series of events that are repeated until the luteolysis impediment, because the presence of embryo in the uterus, which causes hormonal, anatomical and behavioral orders, changes in the maternal organism. Nevertheless, the ovum pick-up in pregnant cows is an alternative to the in vitro production of embryos. However, in view of the hormonal profile during pregnancy, the factors involved in follicular development may be impaired. Thus, this study aimed to describe the protein profile of follicular fluid from pregnant cows. Ovaries were collected from 36 pregnant cows with gestational age 1-4 months. Follicular fluid was punctured and the diameter of each follicle was measured by ultrasonography, classified into three categories ≤6.4mm, 6.5 to 8.9 mm and ≥9. After two centrifugation to remove the cellular components, total protein were measurement and unidimensional electrophoresis was performed under denaturing and reducing conditions. The bands were cut and digestion in gel was carried out aiming to mass spectrometry. The concentration of estrogen and progesterone in follicular fluid was measured to determine the viability of follicles. We found 46 protein bands in electrophoresis in 67 follicular fluid samples. The mean ± standard deviation of progesterone concentration was 71.32 ± 80.06 considering all follicles. There was differential expression of proteins in different categories of follicles. In mass spectrometry were found many proteins, being main apolipoprotein, angiotensinogen, heat shock protein, glutathione and their isoforms.

Luteinising hormone (LH) is a crucial regulator of ovarian follicle maturation, ovulation and luteinisation. Development of healthy follicles and fertile ovulation can only occur within a specific range of circulating LH concentrations, with differing upper and lower limits depending on the stage of the oestrous cycle. The objective of the three studies in this thesis was to investigate the effects of both physiological and non-physiological circulating LH levels on equine follicular maturity by examining ovulatory and steroidogenic capacity, gene expression profiles and miRNA expression in ovulatory-size follicles at various stages of the oestrous cycle and/or in response to supplementation with LH. The aim of the first study was to investigate the hypothesis that deficient circulating LH is a primary cause for the inability of equine follicles to ovulate during the physiological anovulatory season. A LH-rich equine pituitary fraction (eLH) given twice daily to early transitional mares did not restore steroidogenic capacity of the ovulatory-size follicle or advance the onset of the natural breeding season; however, it significantly stimulated follicular growth to a level similar to that occurring during the normal oestrous cycle. The results demonstrated that a deficiency in LH is critically involved in reduced follicle growth during the anovulatory season. The second study examined the effects of elevated circulating LH levels early during follicle development on follicle maturation and ovulatory ability in cycling mares, with the hypothesis that excessive LH would disrupt ovulation and produce haemorrhagic anovulatory follicles (HAFs). Treatment with eLH or a luteolytic dose of prostaglandin F2α (to stimulate an increase in endogenous levels of LH) did not have any effects on follicle growth or ovulation, but did impair follicular production of androstenedione and insulin-like growth factor 1 (IGF1), suggesting a deleterious effect of high LH on follicle and oocyte maturation. The third study examined the expression of different follicular factors associated with follicle maturation as well as microRNAs (miRNAs) in ovulatory-size follicles naturally developing under different LH milieus (oestrus, dioestrus and spring transitional period). Progesterone and IGF1 were significantly reduced in follicles developing in a low LH environment (dioestrus and transition). All four miRNAs measured, miR-378, miR-542, miR-202 and miR-21 were found at higher levels in subordinate follicles than in preovulatory follicles during oestrus. In addition miR- 202 and miR-21 were significantly increased in transitional follicles relative to oestrous follicles. The results of this study indicate that follicles developing during both the spring transitional and dioestrous periods are developmentally immature and suggested potential important roles of miRNAs in follicle maturation in the horse. In summary, although LH is a key factor promoting follicular growth, it is by itself not sufficient to restore steroidogenic activity in transitional follicles. Elevated LH levels during follicle development do not disrupt ovulation, but induce changes in follicular fluid factors related to follicle maturation and oocyte quality. Follicles developing under different LH milieus show altered miRNA expression, suggesting an important role of miRNAs in follicle maturation.

The focus of the present thesis was on ultrasonographic, endocrine and molecular characterization of ovarian antral follicular waves in sheep. Transrectal ultrasonography and computer assisted image analysis were used to determine the feasibility of detecting ovulation and the forming corpus luteum (CL) and to non-invasively monitor CL differentiation and growth. High resolution transrectal ultrasonography and hormone measurements were used to assess changes in numbers of small ovarian antral follicles and their relationships to the emergence of follicular waves in cyclic ewes and to correlate pulsatile secretion of gonadotropins with follicular growth in a wave, during the mid to late-luteal phase of the ovine estrous cycle. A series of experiments were conducted, using treatment with injections of ovine follicle stimulating hormone (oFSH) and measurement of serum concentrations of FSH, in cyclic and anestrous sheep, to investigate the existence of follicular dominance. We also evaluated the characteristics of secretory patterns of FSH that are critical for follicular wave emergence, in anestrous ewes. The possible existence of an endogenous rhythm of FSH secretion, independent of ovarian antral follicular dynamics, was studied in ovariectomized ewes. Finally, ovarian antral follicles at defined stages of growth in a follicular wave (based on transrectal ultrasonographic observations) were collected from separate groups of sheep by ovariectomy, to profile the expression patterns of steroidogenic enzymes (3¦Â-HSD, 17¦Á-OH and aromatase) using immunohistochemistry and gray-scale densitometric analysis. <p> The results of the present studies showed that it is possible to detect ovulation and visualize developing CL as early as 12-24 h after ovulation in the ewe. Changes in echotexture of the CL were closely associated with its morphological and functional characteristics, and we concluded that computer assisted image analysis holds promise for the noninvasive monitoring of CL differentiation and growth. Follicles reaching ovulatory diameter (¡Ý 5 mm) emerged and grew in a wave-like pattern in sheep, but without variation in the number of small follicles (1-3 mm in diameter), as seen in cattle. We concluded that all follicles that are recruited to grow beyond 2-3-mm in diameter, to 4-mm diameter in a wave, succeed in reaching an ovulatory diameter of ¡Ý 5 mm in the ewe. The emergence and growth of ovarian antral follicles in follicular waves, in sheep, do not require changes in LH secretion and may perhaps involve changes in the follicular sensitivity to LH. The largest follicle of a wave, in sheep, appears to have limited effects on other small follicles and on the time of emergence of the next follicular wave. Thus, functional dominance, as is present in cattle, may be absent in sheep. An endogenous rhythm for periodic peaks in serum FSH concentrations that is independent of ovarian follicular dynamics may exist in sheep. The expression patterns of steroidogenic enzymes, in the theca and granulosa compartments of antral follicles growing in each follicular wave in the ewe, paralleled serum estradiol concentrations, with the exception of the concentrations of 3¦Â-HSD in granulosa cells, which increased continuously from follicles 3 mm in diameter to the preovulatory follicle after the LH surge. The largest follicle of any follicular wave, irrespective of the stage of the cycle, would appear to be mature enough to ovulate if a gonadotropin surge is provided.

For this thesis our objectives were to establish an efficient method of ovarian synchronization and superstimulation in bison, and determine the effects of gonadotropin treatments on oocyte collection efficiency and quality in bison. In the first study we conducted two experiments to develop an efficient protocol for synchronization of follicular wave emergence during the anovulatory season. In Experiment 1, we compared the synchronizing effect of follicular ablation (n = 9) and treatment with 2 mg estradiol (E-) 17β in oil (n = 10), while in Experiment 2, we compared follicular ablation (n = 9) and treatment with 2 mg E-17β + 100 mg progesterone (P4; n = 10). Results showed that the degree of synchrony did not differ between ablation and hormone treatment groups in either Experiment, but follicular wave emergence was more synchronous in both treatment groups compared to the untreated control phase. The second study was conducted to develop an efficient method for ovarian superstimulation and oocyte collection during the anovulatory and ovulatory seasons. During the anovulatory season, one experiment was conducted in two replicates to compare the superstimulatory effect of 2500 IU of eCG (n = 10) given intramuscularly vs two doses of 200 mg of pFSH each (n = 10) given subcutaneously. Additionally, the effect of 25 mg of pLH given 24 hours prior oocyte collection on oocyte quality and collection rate was evaluated for each superstimulatory treatment. Results showed that treatment with pFSH induced a higher superstimulatory response and more cumulus oocyte complexes (COC) collected than did eCG during the anovulatory season. Furthermore, treatment with pLH increased the proportion of expanded COC that were collected with ultrasound-guided follicular aspiration. Two experiments were conducted during the ovulatory season, to develop an efficient protocol for superstimulation and oocyte collection. In Experiment 1, we compared the effect of two intramuscular doses of 200 mg of pFSH in saline (n = 11) vs two intramuscular doses of 200 mg of pFSH in a proprietary slow release formulation (SRF; n = 11). In Experiment 2, we compared the effect of a single dose of 2500 IU eCG intramuscularly vs two doses of 200 mg of pFSH administered subcutaneously. Results showed that a 2-dose regime of pFSH, diluted in either saline or a slow-release formulation induced a similar superstimulatory ovarian response in wood bison, while bison given a single-dose of 2500 IU eCG had a significantly lower ovarian response. In summary, synchronization of follicle wave emergence can be effectively accomplished in wood bison during the anovulatory season and follicular ablation, E-17β and E-17β + P4 treatments all shortened, and decreased the variability in the interval to follicular wave emergence. In addition, oocyte collection by transvaginal ultrasound-guided follicle aspiration from superstimulated bison was feasible and practical. Finally, treatment with pFSH was more effective than eCG to induce ovarian superstimulation for ultrasound-guided follicle aspiration in wood bison during both the anovulatory and ovulatory seasons.