Análise comparativa do perfil lipídico do plasma seminal associado ao estresse oxidativo

O estresse oxidativo é um dos fatores mais importantes na diminuição da qualidade do sêmen, pois leva a perda da integridade da membrana dos espermatozóides e de danos estruturais ao DNA através da cascata de lipoperoxidação. Os danos funcionais relacionados ao estresse oxidativo como a diminuição da motilidade e da viabilidade do espermatozóide são algumas das principais causas de infertilidade masculina. Ainda, os lipídios que compõe a membrana plasmática são macromoléculas que, além de estarem envolvidas em complexos sistemas biológicos e processos metabólicos da célula, são altamente susceptíveis ao processo de lipoperoxidação desencadeado pelo estresse oxidativo. Neste contexto, este projeto propôs o estudo das alterações no perfil lipídico do plasma seminal que pudessem estar relacionadas ao estresse oxidativo e posterior comparação destes perfis em busca de biomarcadores de infertilidade. Para isso, foram coletadas amostras de sêmen de 116 pacientes que procuraram o setor de Reprodução Humana da Universidade Federal de São Paulo. Estas amostras foram submetidas a técnica de TBARS para quantificação dos produtos finais do estresse oxidativo e separação dos grupos, e em seguida, ao protocolo de extração de lipídios para obtenção dos espectros de massa através da técnica de MALDI (Matrix Assisted Laser Disorption Ionization). Com esta análise foi possível a identificação de 31 lipídeos super representados nos diferentes grupos e que, futuramente, poderão vir a ser utilizados na avaliação da qualidade seminal

Expressão e localização de aquaporinas no epidídimo do gerbilo, Meriones unguiculatus, em diferentes fases do desenvolvimento pós-natal

Aquaporins (AQPs) are membrane protein channels expressed in a wide variety of cells and tissues that are involved in the transport of water. In the male excurrent duct, the movement of fluid and solutes across the epithelium is essential for establishing the proper luminal environment in which sperm mature and are stored. The initial aim of the present propose is to localize AQP 2 e 9 in epithelium of the initial segment, caput, corpus and cauda epididymidis in Mongolian gerbil during postnatal development (10, 30, 60, 120 and 300 days), observing possible changes in the pattern of expression of the AQPs along the postnatal development. The animals will be killed by inhalation of carbon dioxide. After euthanasia, the epididymis will be removed and the different regions will be isolated. The biological material obtained will be processed in accordance with the routines of histological techniques and imunnohistochemistry to study the expression and localization of the AQPs 2 and 9. The results obtained will be analyzed and photographed in conventional light microscope

Análise químico-farmacêutica de cápsulas de glimepirida

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Principais marcadores utilizados na avaliação da qualidade do sêmen de animais domésticos

Methods of semen cryopreservation allow changes in spermatic cells, such as damage in plasma and acrossomal membrane and modifications in mitochondrial function due to a disorder in the lipidic bilayer. For effective oocyte fertilization, spermatozoa require functional competent membranes, and intact organelles, acrosome and DNA. However, most laboratory methods used to evaluate semen quality are not highly correlated with fertilizing capacity. The discovery of a variety of fluorochromes and compounds conjugated to fluorescent probes has enabled an accurate assessment of the viability, integrity and function of spermatozoa. Among the most used probes that label the various compartments of the sperm cell there are the membrane impermeable fluorescent dyes to test the membrane integrity, as well as acylated dyes that pass the intact membrane. For the acrossomal integrity the most commonly used method is lectins labeled by a fluorescent probe. The acrosome reaction and spermatic capacitation is detected by the evaluation of membrane architecture and disorder of lipids in plasma membrane. Mitochondrial function can be determined using markers for their aerobic activity. The DNA status of spermatozoa has been determined using the metachromatic properties of Acridine Orange, and the DNA fragmentation can also be assessed by TUNEL assay. Finally, DNA condensation is analyzed using a single cell DNA gel electrophoresis assay that indicates DNA compactation. This monograph aims to compile the various tests used to detect damaged spermatozoa under cryopreservation methods, searching for improve the predictive value of semen analysis with the intention of a successful conception

Efeito da ingestão aguda de taurina sobre o desempenho físico e metabolismo lipídico de nadadores de elite de Ribeirão Preto

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Receptor tipo quinase de tomate: caracterização molecular e implicações nas infecções virais

Pós-graduação em Ciências Biológicas (Genética) - IBB

Características seminais e resfriamento de sêmen de tamanduá (Myrmecophaga tridactyla) de vida livre

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Impaired protamination and sperm DNA damage in a Nellore bull with high percentages of morphological sperm defects in comparison to normospermic bulls

The routine semen evaluation assessing sperm concentration, motility and morphology, does not identify subtle defects in sperm chromatin architecture. Bulls appear to have stable chromatin, with low levels of DNA fragmentation. However, the nature of fragmentation and its impact on fertility remain unclear and there are no detailed reports characterizing the DNA organization and damage in this species. The intensive genetic selection, the use of artificial insemination and in vitro embryo production associated to the cryopreservation process can contribute to the chromatin damage and highlights the importance of sperm DNA integrity for the success of these technologies. Frozen-thawed semen samples from three ejaculates from a Nellore bull showed high levels of morphological sperm abnormalities (55.8±5.1%), and were selected for complementary tests. Damage of acrosomal (76.9±8.9%) and plasma membranes (75.7±9.3%) as well as sperm DNA strand breaks (13.8±9.5%) and protamination deficiency (3.7±0.6%) were significantly higher compared to the values measured in the semen of five Nellore bulls with normospermia (24.3±3.3%; 24.5±6.1%; 0.6±0.5%; 0.4±0.6% for acrosome, plasma membrane, DNA breaks and protamine deficiency, respectively) (P<0.05). Motility and percentage of spermatozoa with low mitochondrial potential showed no differences between groups. This study shows how routine semen analyses (in this case morphology) may point to the length and complexity of sperm cell damage emphasizing the importance of sperm function testing.

Avaliação da fosforilação da Akt e do conteúdo da proteína transportadora de glicose GLUT4 em músculo esquelético de ratos adultos com lesão periapical

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Aplicação da equação de Fokker-Planck no estudo de canais iônicos

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Características físicas e morfológicas do sêmen de Bos taurus e Bos indicus antes e após a criopreservação

Pós-graduação em Medicina Veterinária - FCAV

Utilização do choque osmótico na avaliação da viabilidade de sêmen criopreservado de ovinos

This study evaluated the use of hipoosmotic swelling test (HOST) with deionized water (0 mOsmol), as a method of post thaw ram semen evaluation and correlate their findings with different techniques of semen evaluation. Therefore, twenty semen samples of 20 different adult rams were assessed as for kinetic sperm parameters through computerized system (IVOS 12, Hamiton Thorn Biosciences, Beverly, MA, EUA) and subjective analysis. The sperm membranes viability was carried out by the association of fluorescent probes (propidium iodide, JC-1 and FITC-PSA). The structural integrity of the plasma membrane was also studied through supravital test with eosin and the functional integrity of membrane evaluated by doing the hipoosmotic swelling test with deionized water (0 mOsmol), in the following proportions: One part of semen for 10 (HOST 10), 50 (HOST 50) and 100 (HOST 100) parts of water. After semen dilution in the different proportions it was fixed in formalin-buffered saline and analyzed with regard to percentage of HOST reactive sperm (bent/coiled). The percentage of reaction obtained for HOST 10 (33,1%); HOST 50 (32,8%) and HOST 100 (31,8%) did not differ significantly. HOST 10 presented positive correlation with the plasma membrane integrity by the EOS (r = 0,80; p < 0,05). Positive correlations between HOST 50 and HOST 100 with sperm subpopulation with membrane integrity by fluorescence were observed (r = 0,83 and r = 0,85; p < 0,01). The findings suggest that the HOST with deionized water can provide additional information for post thawing ram sperm viability evaluation.

Criopreservación de semen bovino de la cola del epididimo utilizando los métodos convencional y automatizado

Cryopreservation of sperm is important to preserve the germplasm from animals of genetic value, which can die unexpectedly. This study compares conventional and automated methods of cryopreservation of spermatozoa obtained from the epididymis of bulls post-mortem. Twenty-two epididymides were obtained from a commercial slaughterhouse. Spermatozoa were collected from the tail of the epididymis using the retrograde flow technique. Thus, the samples, which were diluted in 10 ml of extender without glycerol (Botubov® I, Botupharma, Botucatu, SP, Brazil), were evaluated on motility, sperm vigor, structural and functional (swelling hypoosmotic test) membrane integrity, mitochondrial activity, sperm viability and ADN fragmentation. The samples were divided into two aliquots and diluted in extender with glycerol (Botubov® II, Botupharma, Botucatu, SP, Brazil) at a concentration of 50x106 motile sperm/0.5 French straws. One sample was frozen by the conventional method (4 hours at 5°C, in a refrigerator and 20 min in nitrogen vapor) and the other by the automated method (Cryogen® Dualflex, Neovet, Uberaba, MG, Brazil). The parameters were higher in all the tests of fresh sperm samples, with the exception of the swelling hypoosmotic test, which showed no significant difference when the results were compared with sperm frozen by the conventional method. The average motility of fresh spermatozoa was 74%, and conventional and automated averages were 29 and 25%, respectively. Therefore, although cryopreservation techniques reduce sperm quality parameters, the viability of the sperm is maintained, and these methods can be used to preserve sperm.

Trehalose and a calcium chelator for ram semen cryopreservation

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeito da adição de butil-hidroxitolueno nos meios de refrigeração e congelação sobre a viabilidade espermática de equinos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)