Navegando por Palavras-chave "Ducto Mesonéfrico"

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    Expressão Espaço-Temporal Do Receptor De Glicocorticoide No Desenvolvimento Epididimário
    (Universidade Federal de São Paulo (UNIFESP), 2017-07-31) Thimoteo, Daniel Soares [UNIFESP]; Avellar, Maria Christina Werneck De [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
    O epidídimo é um órgão do trato reprodutor masculino responsável pela maturação, transporte, estocagem e proteção dos espermatozoides e, assim fundamental para fertilidade masculina. O desenvolvimento embrionário e pós-natal do epidídimo é um evento complexo, que envolve vários fatores modulatórios, incluindo os androgênios que são primários para a estabilização e diferenciação morfológica do ducto mesonefrico e manutenção da sua morfologia e funções ao longo da vida pos-natal e adulta. Além dos androgênios, outros hormônios esteroidais, como os glicocorticoides, também têm sido mostrados com relevância e potencial papel na regulação fisiológica do desenvolvimento e função epididimária e, consequentemente, fertilidade masculina. Nosso grupo de pesquisa relatou anteriormente a presença e funcionalidade do receptor de glicocorticoide (GR) em epidídimo de ratos adultos. O conhecimento dos mecanismos pelos quais o GR e sua sinalização afetam o desenvolvimento embrionário do trato reprodutor masculino, no entanto, é ainda escasso. No presente projeto, caracterizamos a expressão espacial e temporal do GR e sua sinalização celular no desenvolvimento embrionário e pós-natal do epidídimo de ratos Wistar. No período pré-natal focamos os estudos no ducto mesonéfrico (também chamado ducto de Wolff), o precursor embrionário do epidídimo. Especificamente avaliamos: a) a concentração plasmática de testosterona e corticosterona de fetos machos nas idades embrionárias e17,5 e e20,5; b) o perfil de expressão de transcritos de Gr e do receptor de androgênio (Ar) em ductos mesonéfricos de fetos machos nas idades e12,5, e17,5 e e20,5 e, c) o perfil de expressão de transcritos do receptor de mineralocorticoide (Mr) e da enzima 11β-HSD1 (Hsd11b1) em ductos mesonéfricos de fetos machos nas idades e17,5 e e20,5; d) a imuno-distribuição do GR, AR e 11β-HSD1 em ductos mesonéfricos de fetos machos nas idades e14,5, e16,5, e18,5 e e20,5 e e) o perfil de expressão (mRNA) e a imunodistribuição do GR e 11β-HSD1 em epidídimo de ratos nas idades pós-natais dpn1 a dpn120. Os dados em conjunto corroboram com a hipótese de que androgênios e glicocorticoides atuem conjuntamente durante a morfogênese do ducto mesonéfrico e desenvolvimento pós-natal epididimário, implicando que desbalanços na sinalização de androgênios/glicocorticoides e seus respectivos receptores durante o período embrionário possam contribuir para desordens reprodutivas masculinas na vida adulta. A presença de um receptor GR funcional no ducto mesonefrico em cultura organotipica ex vivo está em avaliação na continuidade deste trabalho.
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    Morfogênese epididimária: relevância funcional de androgênios e glicocorticoides
    (Universidade Federal de São Paulo (UNIFESP), 2020-07-30) Sousa, Maria Eleticia De [UNIFESP]; Winston, Maria Christina Werneck De Avellar [UNIFESP]; Universidade Federal de São Paulo
    The epididymis is a single and convoluted tubule, which connects the efferent ductules to the vas deferens. It is an important organ for sperm maturation, transport, storage, and their protection before ejaculation. The epididymal morphogenesis, that occurs from the mesonephric duct (its embryonic precursor), is primarily dependent on androgens by mechanisms that are not well defined yet. Recently, our research group has identified a differential expression pattern of glucocorticoids receptors (GR) on epithelial and mesenchymal cells of the developing rat mesonephric duct. The biological role of GR/glucocorticoid signaling, as well as its potential interaction with androgens during pre- and post-natal development of the epididymis is not well established yet. In the present study, we used a preclinical model of dexamethasone (synthetic glucocorticoid) antenatal treatment in rat dams (Wistar) as a tool to help to elucidate the role and function of glucocorticoids during the mesonephric duct development, with a focus on epididymal morphogenesis. Initially, we tested the daily DEX (1 mg/kg, s.c.) treament of dams during the gestational period that, in the rat, correlates with epididymal morphogenesis (e17.5-e19.5; group DEX-LW versus control, SAL-LW; euthanasia at e20.5). The specific aims, using this preclinical model, were: a) to evaluate the effects of the DEX antenatal treatment on (i) biometric and hormonal parameters in the dams, and (ii) biometric and hormonal parameters in the male fetuses, b) to characterize the effects of the DEX antenatal treatment on histomorphological parameters and cellular proliferation rate of the mesonephric duct of the male offsprings, c) to characterize the effects of the DEX antenatal treatment in the immunodistribution of AR and GR in the mesonephric duct of the male offsprings, d) to characterize the effect of the DEX antenatal treatments in the transcriptional levels of a subset of genes that are target of GR and AR regulation. Based on the results obtained, we subsequently conducted studies by exposing rat dams to daily DEX treatment during the gestacional period that involves masculinization and differntation windows (e13.5-e19.5; DEX-FL versus control SAL-FL; euthanasia at e20.5). Our readouts were: biometrical parameters (dams and fetuses), hormonal dosage (corticosterone and testosterone; dams and fetuses), histopathology, immunofluorescence studies and quantification of gene expression by RT-qPCR on isolated mesonephric duct. Althogether the results confirmed the hypothesis that glucocorticoids play a role in morphogenetic events of the mesonephric duct, including epididymal morphogenesis; also, the data contributed to the better understanding that changes in maternal plasma concentration of glucocorticoids impact the expression and function profile of GR and AR in the duct. From a translational point of view, the present study corroborates to a better understand of how corticotherapy or changes in the endougenous glucocorticoids profile during gestational period may influence reproductive health of male offsprings. (Research Ethics Committee, CEUA/UNIFESP, #8759031215).
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    Proteínas dissulfeto isomerase (PDIs): relação com hormônios esteroidais sexuais e função epididimária
    (Universidade Federal de São Paulo (UNIFESP), 2019-07-25) Fernandes, Samuel Guilerme [UNIFESP]; Winston, Maria Christina Werneck De Avellar [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
    Protein disulfide isomerase (PDI) family is a member of the thioredoxin protein superfamily. There are at least 21 genes encoding PDIs that greatly vary in their protein structure, tissue expression, and cellular distribution. PDIs are multifunctional chaperones primarily involved in the catalysis of thiol-disulfide exchange reactions for protein folding, which are rate-limiting steps for protein maturation in the endoplasmic reticulum. In addition to their roles in different physiological processes, including sperm maturation and male fertility, PDIs have also been associated with several pathologies. The mechanisms by which PDIs regulate the male reproductive function in health and disease, however, are still poorly understood. Surprisingly, we observed in pilot studies that transcripts encoded by at least 20 Pdi genes are present in the caput epididymis from adult rats. Considering the crucial role of the epididymis to sperm function, we set to uncover in the present study the expression pattern and androgen regulation of a subset of Pdi genes (P4hb, Pdia3, Pdia5, Pdia6, Pdilt and Erp29) in the epididymis of developing rats. Firstly, we detected Pdi mRNAs in the Wolffian duct (embryonic epididymal precursor) from fetuses at embryonic ages (e) e17.5 and e20.5, a temporal window when epididymal morphogenesis occurs; in contrast to similar mRNA levels of Pdilt and Erp29 genes, upregulation of P4hb, Pdia3, Pdia5 and Pdia6 mRNA was observed in WD from e17.5 to e20.5. Next, we observed that these Pdi genes were differentially expressed during postnatal in epididymis from neonatal rats (pnd1, pnd5, pnd10 and pnd20; whole epididymis) and in epididymal regions from sexually immature (pnd40) and mature rats (adult, pnd120); in these studies, only Pdia3, Pdia5 and Pdia6 genes displayed significant age-related changes in their transcript levels. Surgical castration, followed or not by testosterone replacement and efferent ductules ligation were then performed in adult rats to evaluate Pdi gene modulation by androgens and/or testicular factors, respectively. The downregulation of Pdia3, Pdia5, Pdia6 and Erp29 transcripts in androgen-deprived caput epididymis was prevented by testosterone replacement, indicating their positive regulation by androgens. Conversely, the castration-induced decrease in caput epididymis P4hb and Pdilt mRNA levels were only partially or not restored, respectively, by the androgen treatment. P4hb was the only Pdi gene to be downregulated following efferent ductules ligation, suggesting its modulation by both androgens and testicular factors. The bioinformatic-based analysis revealed potential role for androgen/AR signaling pathways in the regulatory modulation of some, but not all Pdi genes. Lastly, immunohistochemistry was conducted with antibodies against two PDIs, i.e. PDILT and P4HB; both PDIs were immunolocalized in epithelial and non-epithelial cells along the adult epididymis and also in the luminal spermatozoa. Altogether, our results give support for PDIs as novel players in the developing and adult epididymis.
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    Regulação Androgênica, Atividade Biológica e Sinalização Celular de Beta-Defensinas na Morfogênese do Epidídimo
    (Universidade Federal de São Paulo (UNIFESP), 2019-07-25) Ferreira, Lucas Garcia Alves [UNIFESP]; Winston, Maria Christina Werneck De Avellar [UNIFESP]; Ribeiro, Camilla Moreira [UNIFESP]; http://lattes.cnpq.br/9498472516886944; http://lattes.cnpq.br/5534065920525009; http://lattes.cnpq.br/2251799531659813; Universidade Federal de São Paulo (UNIFESP)
    Reproductive disorders in adult men, a topic of public health interest, may be related to develop-mental and masculinization abnormalities of the male reproductive tract. The mesonephric duct (or Wolffian duct) is an embryonic structure that undergoes a process of tubular-epithelial morphogenesis primarily induced by androgens that originates the epididymis, an essential organ for sperm maturation and function. Disruption of the development of this structure may result in infertility at adulthood, jus-tifying the need for a better understanding of the mechanisms and mediators driving epididymal mor-phogenesis. Our research group has recently identified the β-defensin SPAG11C (sperm associated an-tigen 11 C), a component of innate immunity, as an androgen-dependent mesenchymal factor that mod-ulates the rat mesonephric duct morphogenesis. We have also detected the expression of other β-defen-sins in this tissue. Members of this protein family can promote their effects via Toll-like receptor (TLR) cellular signaling. Taken together, the present aims of the study were to characterize the expression profile and the androgen regulation of a subset of β-defensins in the rat mesonephric duct, to investigate the functional aspects of the DEFB2 (defensin beta 2) and its potential involvement with TLR4 signaling pathway during the prenatal epididymal development, as well as to validate an experimental approach for the detection of recombinant β-defensin processing and secretion in a cell culture model. Our results revealed that the tested β-defensins presented differential gene expression, androgen regulation and cel-lular distribution profile during prenatal epididymal morphogenesis in rat. The β-defensin Defb4 tran-script, the homologue of human DEFB2 (hDEFB2), was negatively regulated between the embryonic days (e) 17.5 and 20.5, a period when the mesonephric duct morphological differentiation occurs. In ducts (e17.5) cultured ex vivo for 96 h, recombinant hDEFB2 reduced duct elongation and coiling as a result of increased cell death in both epithelial and mesenchymal cells. We have also observed that TLR4-induced signaling pathway is present and functional in the mesonephric duct, and its activation by LPS from E. coli can be negatively regulated by hDEFB2. Furthermore, we have validated a Western blot protocol for the detection of SPAG11C protein forms on cell lysate and supernatant from cultured human cells transiently transfected to express this β-defensin. Collectively, these results reveal the par-ticipation of innate immunity components on the scenario of mesonephric duct development, contrib-uting to increased knowledge of the functional repertoire and role of β-defensins in the epididymis and in events related to male fertility.