Espermatogenese em Passer domesticus (Aves, passeriformes) : estudo estrututral e ultra-estrutural

Orientador: Mary Anne Heide DolderDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: O presente estudo teve como objetivo principal analisar os processos envolvidos na espermatogênese de um passeriforme, passer domesticus. Para isso foram empregadas técnicas de microscopia eletrônica de transmissão, técnicas de microscopia de luz, bem como dois testes citoquímicos. A análise dos cortes em parafina revelam que o testículo deste animal é constituído de uma massa de túbulos semíniferos anastomosados contendo epitélio germinativo com células em diversos graus de maturação. As espermatogônias formam uma camada basal neste epitélio e acima delas são encontrados grupos de espermatócitos primários. Ao nível ultra-estrutural podemos verificar que os espermatócitos primários apresentam aspectos nucleares muito peculiares, permitindo sua classificação nas subfases da prófase I. Os estágios da prófase I, neste animal, são marcados pela presença de uma estrutura intra-nuclear esférica, o corpo redondo, e massas citoplasmáticas densas. O posicionamento das espermátides no interior do túbulo seminífero, e seu modo de interação com a célula de Sertoli, são resposáveis pela formação de feixes, organizados radialmente no interior do túbulo. Com bases em diversas características morfológicas, principalmente o grau de compactação cromatínica, as espermátides foram classificadas em 6 estádios de diferenciação: E1, E2, E3, E4, E5 e E6. Em cortes ultrafinos verifica-se que o grau de compactação cromatínica resulta no surgimento de padrões nucleares regulares e o tratamento pelo Azul de Toluidina á ph 4.0 indica a substituição das proteínas histônicas por protaminas. O desenvolvimento do flagelo e acrossomo iniciam-se muito cedo durante a espermiogênese, em regiões adjacentes e em associação com um complexo de Golgi bem desenvolvido, e o feixe de microtúbulos se relaciona a diversas mudanças celulares, contribuindo para obtenção da forma final dos espermatozóides. Os espermátozoides testiculares são células helicoidais, com acrossomo cônico bastante alongado, onde se destaca a crista espiral. Eles apresentam núcleo compacto e cauda extremamente longa. Na cauda, além do axonema com fibras densas acessórias, destaca-se a mitocôndria muito longa, fazendo um trajeto helicoidal ao redor do axonema. As células de Sertoli desempenham um papel fundamental na espermatogênese do pardal, fornecendo um suporte mecânico e estando envolvidas no processo de espermiaçãoAbstract: This study was undertaken to analyse the process of spermatogenesis of a passerine bird, Passe r domesticus. Preparation were made for cytochemical analysis and morphological studies with light and electron microscopes. Sections of parattin embedded testes revealed its or ganization in the form of branching seminiferous tubules, with cells of the germinative epithelium in various maturation stages. Spermatogoniae form a basal layer, overlaid by primary spermatocytes. Typical nuclei permitted the classification of these cells into sub-stages of prophase I, in which the intra nuclear round body and dense cytoplasmic masses were also observed. The positioning of the spermatids in the seminiferous tubules and their envelopment by the Sertoli cell, lead to their organization into bundles. Spermatids were grouped into six stages (E1, E2, E3, E4, E5 e E6) according their morphological characteristics, especially the degree of chromatin condensation. Ultrathin sections revealed that nuclear compactation occur in regular, well established patterns, while the Toluidine Blue staining at pH 4.0 indicated the substitution of histones by protamines. Early development of the acrosome and the flagellum occur adjacent to a well developed Golgi complex. The microtubular sheath is related to various cellular events, probably contibuting to the final shape of the spermatozoa. Spermatozoa are helically coiled cells with a long conical acrosome, marked by a spiral keel, a compact nucleus and a very long tail. In the tail, the axoneme is surrounded by dense accessory fibers, and a long helically arranged mitochondrion. Sertoli cells are of fundamental importance in spermiogenesis. They contribute with the mechanical support, metabolic processes such as reabsortion and partipate in spermiationMestradoBiologia CelularMestre em Ciências Biológica

Acid Phosphatase Activity In Gerbil Prostate: Comparative Study In Male And Female During Postnatal Development.

The prostate is present in both male and female mammals. It is composed of secretory epithelium, connective stroma, smooth muscle and neuroendocrine cells, which are under hormonal regulation. Acid phosphatases catalyze the hydrolysis of orthophosphate monoesters. We have compared the expression of acid phosphatases in gerbil (Meriones unguiculatus) prostate glands in both sexes using young, adult and old animals. Eighteen prostates were isolated, frozen, sectioned, fixed, incubated with sodium beta-glycerophosphate sodium, washed with acetate buffer solution, treated with ammonium sulfide and counterstained with Methyl-Green aqueous solution. Ultracytochemical analyses were also conducted. This substrate revealed total acid phosphatase activity. The expression of the enzyme was heterogeneous, occurring in all ages during postnatal development. The data revealed that the female prostate matured before the male prostate. In addition, acid phosphatase activity in both sexes was regulated by androgen variation concomitant with development.28335-4

Modulação androgênica e estrogênica na próstata: uma abordagem em modelos experimentais de roedores com enfoque na biologia estrutural

O sistema endócrino é uma complexa rede de glândulas e hormônios que regulam muitas das funções do corpo, incluindo crescimento, desenvolvimento e maturação, como as vias de ação de muitos órgãos. A próstata é um importante alvo dos hormônios e sua maturidade funcional e seu desenvolvimento são influenciados pelos níveis de esteroides. O presente grupo de pesquisa tem estudado os potenciais efeitos dos agentes esteroides sobre a próstata masculina e feminina do gerbilo da Mongólia (Meriones unguiculatus), utilizando métodos morfológicos e imuno-histoquímicos. Os resultados têm revelado a próstata do gerbilo da Mongólia como uma importante ferramenta para estudos da ação dos hormônios esteroides e seus antagonistas.The endocrine system is a complex network of glands and hormones that regulates many of the body's functions; including growth, development and maturation, as well as the way several organs operate. The prostate is an important target of hormones and its functional maturity and development are influenced by steroids levels. Our research group has been evaluating the potential effects of the steroidal agents on the Mongolian gerbil (Meriones unguiculatus) male and female prostate using different morphological and immunohistochemical methods. Our results have revealed the Mongolian gerbil prostate as an important tool for the morphofunctional studies of steroid hormones and its antagonist actions.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Biological Behavior Of The Gerbil Ventral Prostate In Three Phases Of Postnatal Development.

In this study, we characterized the gerbil's ventral prostate histology ultrastructurally and quantitatively throughout three phases of postnatal development (young, adult, and old) in order to comprehend its biological behavior and propensity to developing spontaneous lesions with aging. The gerbil prostate is composed of alveoli and ducts immersed in a stroma composed of smooth muscle, fibroblasts, collagen and elastic fibers and vessels. The prostate tissue components present morphological and quantitative aspects that vary according to age. Young animals have an immature gland with modest secretory activity. Synthetic activity remained stable in adult and old gerbil. However, prostatic morphology was altered in the aging, showing an increased epithelium and stromal fibrosis. The nuclei of the secretory cells increased with aging, whereas nucleoli presented few alterations during postnatal development. The epithelial proliferation and stromal remodeling noted in this study indicate that the gerbil prostate may respond to the androgen declines typical of senescence through epithelial proliferation and stromal remodeling.288723-3

A high-fat diet fed during different periods of life impairs steroidogenesis of rat Leydig cells

This study evaluated the impact of a high-fat diet (HFD) during different stages of rat life, associated or not with maternal obesity, on the content of sex steroid hormones and morphophysiology of Leydig cells. The following periods of development were examined: gestation (O1), gestation and lactation (O2), from weaning to adulthood (O3), from lactation to adulthood (O4), gestation to adulthood (O5), and after sexual maturation (O6). The HFD contained 20% unsaturated fat, whereas the control diet had 4% fat. Maternal obesity was induced by feeding HFD 15 weeks before mating. All HFD groups presented increased body weight, hyperinsulinemia and reduced insulin sensitivity. Except for O1, all HFD groups exhibited a higher adiposity index, hyperleptinemia, reduced testosterone and estradiol testicular levels, and decreased testicular 17β-HSD enzyme . Morphometrical analyses indicated atrophy of Leydig cells in the O2 group. Myelin vesicles were observed in the mitochondrial matrix of Leydig cells in O3, O4, O5 and O6, and autophagosomes containing mitochondria were found in O5 and O6. In conclusion, HFD feeding, before or after sexual maturation, reduces the functional capacity of rat Leydig cells. Maternal obesity associated with HFD during pregnancy/lactation prejudices Leydig cell steroidogenesis and induces its atrophy in adulthood, even if it is replaced by a conventional diet at later stages of life. Regardless of the life period of exposure to HFD, deregulation of leptin is the main factor related to steroidogenic impairment of Leydig cells, and, in groups exposed for longer periods (O3, O4, O5 and O6), this is worsened by structural damage and mitochondrial degeneration of these cells.1526795808CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP306258/20112011/01612-4; 2009/16071-9; 2011/03596-6; 2013/18011-

Differentiation Of Leydig Cells In The Mongolian Gerbil.

Information on postnatal Leydig cell (LC) differentiation in the Mongolian gerbil has been unavailable. Therefore, current investigation was designed to examine LC lineage differentiationin this rodent, from birth to adulthood. Gerbil testes at 1 day, 1-7 weeks (w), 2 and 3 months of age were conventionally processed by light and transmission electron microscopy. Immunocytochemistry for specific markers of steroidogenic enzymes, namely 3beta-hydroxysteroid dehydrogenase (3beta-HSD) and 11beta-hydroxysteroid steroid dehydrogenase 1 (11beta-HSD1) and also for androgen receptor (AR) was performed. The establishment of adult Leydig cell populations (ALC) during testis maturation in the gerbil follows the pattern previously described in other mammalian species, with the four progressive stages of differentiation. The LC progenitors were identified at second w by 3beta-HSD expression; the first newly formed ALC were recognized at fourth w whereas immature ALC appeared at fifth w. The latter were recognized by abundance of cytoplasmic lipid, in addition to expression of 11beta-HSD1 and intense nuclear AR immunoreaction. Mature ALC in gerbil exhibited irregular eccentric nuclei and a cytoplasmic canaliculus in the perinuclear area. Only one third of mature ALC in adult gerbils showed a high expression of 11beta-HSD1 and AR expression was highly variable among them. In conclusion, the process of differentiation of ALC population in gerbil follows the pattern previously established for other rodents. However, the resulting mature ALC are strikingly different due their singular asymmetric morphology and presence of a cytoplasmic canaliculus and as well as their functional heterogeneity.73119-2

Differential Ontogenetic Exposure To Obesogenic Environment Induces Hyperproliferative Status And Nuclear Receptors Imbalance In The Rat Prostate At Adulthood.

Experimental data indicate that high-fat diet (HFD) may alter proliferative activity and prostate health. However, the consequences of HFD exposure during different periods of ontogenetic development on prostate histophysiology remain to be elucidated. Herein, we compare the influence of obesogenic environment (OE) due to maternal obesity and HFD at different periods of life on proliferative activity and nuclear receptors frequency in the rat ventral prostate and a possible relationship with metabolic and hormonal alterations. Male Wistar rats (19 weeks old), treated with balanced chow (Control group-C; 3% high-fat, 3.5 Kcal/g), were compared with those exposed to HFD (20% high-fat, 4.9 kcal/g) during gestation (G-maternal obesity), gestation and lactation (GL), from post-weaning to adulthood (WA), from lactation to adulthood (LA) and from gestation to adulthood (GA). After the experimental period, the ventral prostate lobes were removed and analyzed with different methods. Metabolic data indicated that G and GL rats became insulin resistant and WA, LA, and GA became insulin resistant and obese. There was a strong inverse correlation between serum testosterone (∼133% lower) and leptin levels (∼467% higher) in WA, LA, and GA groups. Estrogen serum levels increased in GA, and insulin levels increased in all groups, especially in WA (64.8×). OE-groups exhibited prostatic hypertrophy, since prostate weight increased ∼40% in G, GL, LA, and GA and 31% in WA. As indicated by immunohistochemistry, all HFD-groups except G exhibited an increase in epithelial cell proliferation (PCNA-positive) and a decrease in frequency of AR- and ERβ-positive epithelial cells; there was also an increment of ERα-positive stromal cells in comparison with control. Cells containing PPARγ increased in both epithelium and stroma of all OE groups and those expressing LXRα decreased, particularly in groups OE-exposed during gestation (G, GL and GA). OE leads to prostate hypertrophy regardless of the period of development and, except when restricted to gestation, leads to a hyperproliferative status which was correlated to downregulation of AR and LXRα and upregulation of ERα and PPARγ signaling. Prostate © 2016 Wiley Periodicals, Inc.7

Estrogen receptors alpha and beta in male and female gerbil prostates

The Mongolian gerbil (Meriones unguiculatus, Gerbilinae: Muridae) is useful for prostate studies, because both males and females spontaneously develop prostatic disorders with age. Estrogens regulate prostate homeostasis via two estrogen receptors, ER alpha (ESR1) and ER beta (ESR2), but the cellular distribution and regulation of these receptors in the gerbil prostate has not been described. Both receptors were localized by immunohistochemistry in the ventral prostate of intact male and female gerbils, in males 7 and 21 days after castration, and in females treated with testosterone for 7 and 21 days. In male and female adult gerbils, ER alpha was detected mainly in prostatic stromal cells, whereas ER beta was present mostly in secretory and basal cells. More ER alpha-positive stromal cells were found in females than in males, as was a reduction toward the male value in females treated with testosterone. Castration did not alter ER alpha expression. Testosterone was necessary for maintenance of ER beta in the male prostate epithelium: ER beta expression declined markedly in prostates of males older than 1 yr, and castration of 4-mo-old males caused a reduction in ER beta to levels seen in 1-yr-old males. Because ER beta is an antiproliferative receptor, its loss with age may predispose the aging gerbil to proliferative diseases of the prostate. © 2013 by the Society for the Study of Reproduction, Inc