Navegando por Autor "BRITO, Danielle Cristina Calado de"

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Acesso aberto (Open Access)
Expressão gênica e viabilidade de folículos ovarianos pré-antrais de Sapajus apella congelados e cultivados in vitro
(Universidade Federal do Pará, 2012-03-05) BRITO, Danielle Cristina Calado de; DOMINGUES, Sheyla Farhayldes Souza; http://lattes.cnpq.br/2794753357251149; SANTOS, Regiane Rodrigues dos; http://lattes.cnpq.br/0500967766886604
The aim of the present study was to develop a freezing protocol for the preservation of preantral follicles from Sapajus apella (capuchin monkeys). To this end, ovarian fragments were exposed to different cryoprotectant solutions, added or not by antioxidants (selenium or trolox), frozen and in vitro cultived by 24 hours. Morphology, ultrastructure, viability, oxidative stress and mocelular analyses were performed. The collection site was in the Primates Nacional Center (CENP) and nine healthy mature female capuchin monkeys were used. Ovarian biopsies of 1 mm3 were collected by laparoscopy. The follicles were classified accordingly to their developmental phase in primordial, primary or secondary. Follicular viability scored using fluorescent markers (propidium iodide and Hoechst). qRT-PCR was used to evaluate the expression of hormones and growth factors. TEAC was used to measure oxidative stress in the tissue. In cryoprotectant solution containing trolox did not affected follicular morphology and gene expression. Cryopreservation resulted in higher rates of follicular viability when trolox was present in the solution, but expression of genes encoding BMP4 and KL was negatively affected. Our findings show a favorable effect of adding trolox to a cryopreservation solution. However, follicular viability and gene expression was affected after in vitro culture.
Acesso aberto (Open Access)
Vitrificação de tecido ovariano de gata doméstica (Felis catus): um modelo para a preservação da fertilidade em felinos silvestres
(Universidade Federal do Pará, 2016-09-06) BRITO, Danielle Cristina Calado de; SANTOS, Regiane Rodrigues dos; http://lattes.cnpq.br/0500967766886604; PIECZARKA, Julio Cesar; http://lattes.cnpq.br/6644368250823351
The aim o the present thesis was to develop an efficient vitrification protocol of the ovarian tissue from domestic cat (Felis catus). This study was divided: Phase I: Effect of different basis media during the vitrification of cat ovarian tissue; Phase II: Effect of different sugars (extracellular cryoprotectants) and the vitrification technique for the vitrification of feline ovarian tissue. In phase I, the morphology of preantral follicles was similar (p > 0.05) to fresh control when RPMI-1640 was used as basis medium for vitrification. RPMI-1640 does not contain phenol red, which was found to enhance ethylene glycol (EG) toxicity during vitrification. In phase 2, the percentage of morphologically normal preantral follicles was similar (p > 0.05) to fresh control only when the vitrification medium contained 0.1 or 0.5 M trehalose, instead of sucrose or raffinose at same concentrations. Furthermore, based on parameters such as morphology, cell proliferation and thickness of collagen fibers, it is possibe to assume that efficient vitrification of feline ovarian tissue can be performed by combining trehalose with EG, with or without dimethylsulfoxide (DMSO), applying the solid-surface vitrification (SSV) or ovarian tissue cryosystem (OTC) method. Although vitrification with OTC in the presence of EG did not differ from the other treatments, this protocol presented the highest percentages of preserved preantral follicles (56%), being similar to control (64%). Additionally, no effect on gene regulation was observed after vitrification when apoptosis markers (BAX – protein X associated to Bcl-2), endoplasmic reticulum (ER) stress (ER protein 29 – ERP29), water channels proteins like aquaporins 3 and 9 (AQP3 and AQP9), the membrane ABC transporters ABCB1 and ABCG2, except when the SSV method was applied using only EG as cryoprotectant followed by seven days in vitro culture, where ERP29 up-regulation (ER stress) and AQP9 down-regulation (impaired water transport) were observed. Based on this, it can be concluded that to efficiently preserve feline ovarian tissue, is is necessary the use of a vitrification protocol free of phenol red, supplemented with trehalose, as extracellular cryoprotectant, and EG alone or in combination with DMSO, as intracellular cryoprotectants. Both open (SSV) and closed (OTC) systems are equaly efficient to maintain follicular survival during the vitrification procedure.