Efeito da centrifugação e filtragem do sêmen bovino sobre a criopreservação espermática
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Data
2016-02-26
Autores
Campanholi, Suzane Peres [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
O plasma seminal já foi descrito como benéfico e ao mesmo tempo prejudicial aos espermatozoides e isto está relacionado com o tempo de contato entre eles. A criopreservação do sêmen em touros pode ser prejudicada por exposição contínua dos espermatozoides ao plasma seminal (PS) e a aplicação de métodos para sua remoção pode aumentar a qualidade dos espermatozoides recuperados pós-descongelação, melhorando os resultados de biotécnicas que utilizam sêmen criopreservado. Pelo fato da centrifugação causar muitos danos aos espermatozoides, a filtragem com Sperm Filter® apresenta-se como um novo método de remoção do PS que almeja melhores resultados, porém não foi encontrado relato da sua aplicação em bovinos até o momento. O objetivo deste estudo foi avaliar o efeito dos métodos de centrifugação e filtragem com Sperm Filter® sobre a criopreservação do sêmen bovino. Para isso, o sêmen de 38 touros Nelore foi colhido por eletroejaculação. Após a colheita, o sêmen foi fracionado em três alíquotas iguais para divisão em três grupos: controle (N), em que o sêmen foi diluído com PS; centrifugação (C), em que o PS foi removido por centrifugação a 600 X g por 10 minutos; e filtragem (F), em que o PS foi removido por filtragem com Sperm Filter®. As amostras foram criopreservadas, e pós-descongelação foram avaliados a cinética espermática, a integridade das membranas plasmática e acrossomal, o potencial mitocondrial, o estresse oxidativo e a capacidade fecundante através da produção in vitro de embriões (PIVE). Com relação à cinética, maiores valores da velocidade do trajeto (P = 0,0005) e velocidade progressiva (P = <0,0001) foram observados nos grupos C e F e os parâmetros frequência de batimento, retilinearidade e linearidade foram superiores no grupo F (P = 0,0008; P = 0,0133 e P = 0,0005, respectivamente). A integridade de membrana foi prejudicada pela centrifugação e filtragem do sêmen (P < 0,0001), porém o estresse oxidativo foi reduzido com esses métodos de remoção do PS (P < 0,0001). Na PIVE, as maiores taxas de desenvolvimento embrionário (blastocistos e blastocistos eclodidos) foram observadas no grupo N e F (P = 0,008 e P = 0,0042, respectivamente). Portanto, a remoção do plasma seminal pelo método da centrifugação reduz a qualidade do sêmen criopreservado bovino, interferindo negativamente na fertilidade dos espermatozoides e o método da filtragem com Sperm Filter® apresentou melhor resultado evidenciado por altas taxas de desenvolvimento embrionário.
Seminal plasma has been described as beneficial and harmful to the spermatozoa and this is related to the contact time between them. Cryopreservation of semen from bulls can be undermined by continuous exposure of sperm to the seminal plasma (SP) and the application of methods for removal can increase the quality of post-thaw sperm recovered, improving fertilization. Centrifugation cause a lot of damage to sperm and filtering with Sperm Filter® is a new SP removal method that aims to better results, but has not yet been applied in bull. The aim of this study was to evaluate the effect of the methods of centrifugation and filtering with Sperm Filter® on cryopreservation of bovine semen. For this, semen of 38 Nelore bulls was collected by electroejaculation. Semen was fractioned into three equal aliquots for division into three groups: control (N), wherein the semen was diluted with SP; centrifugation (C), in which the SP was removed by centrifugation at 600 X g for 10 minutes; and filtration (F), in which the SP was removed by filtration with Sperm Filter®. Samples were cryopreserved and was evaluated after thawing sperm kinetics, the integrity of plasma and acrosomal membranes, mitochondrial potential, oxidative stress and the fertilizing capacity by in vitro embryo production (IVP). The kinetics, higher values of the path velocity (P = 0.0005) and progressive speed (P = <0.0001) were observed in the groups C and F and the beat frequency parameters, straightness and linearity were higher in group F (P = 0.0008, P = 0.0133 and P = 0.0005, respectively). Membrane integrity was impaired by centrifugation and filtration of semen (P <0.0001), though oxidative stress was reduced with these SP removal methods (P <0.0001). In IVP, the highest rate of embryonic development (blastocyst and hatched blastocyst) were observed in the C and F group (P = 0.008 and P = 0.0042, respectively). Therefore, removal of seminal plasma by the method of centrifugation reduces the quality of semen cryopreserved cattle, a negative effect on fertility of sperm, and the method of filtering with Sperm Filter® showed better results evidenced by high rates of embryonic development.
Seminal plasma has been described as beneficial and harmful to the spermatozoa and this is related to the contact time between them. Cryopreservation of semen from bulls can be undermined by continuous exposure of sperm to the seminal plasma (SP) and the application of methods for removal can increase the quality of post-thaw sperm recovered, improving fertilization. Centrifugation cause a lot of damage to sperm and filtering with Sperm Filter® is a new SP removal method that aims to better results, but has not yet been applied in bull. The aim of this study was to evaluate the effect of the methods of centrifugation and filtering with Sperm Filter® on cryopreservation of bovine semen. For this, semen of 38 Nelore bulls was collected by electroejaculation. Semen was fractioned into three equal aliquots for division into three groups: control (N), wherein the semen was diluted with SP; centrifugation (C), in which the SP was removed by centrifugation at 600 X g for 10 minutes; and filtration (F), in which the SP was removed by filtration with Sperm Filter®. Samples were cryopreserved and was evaluated after thawing sperm kinetics, the integrity of plasma and acrosomal membranes, mitochondrial potential, oxidative stress and the fertilizing capacity by in vitro embryo production (IVP). The kinetics, higher values of the path velocity (P = 0.0005) and progressive speed (P = <0.0001) were observed in the groups C and F and the beat frequency parameters, straightness and linearity were higher in group F (P = 0.0008, P = 0.0133 and P = 0.0005, respectively). Membrane integrity was impaired by centrifugation and filtration of semen (P <0.0001), though oxidative stress was reduced with these SP removal methods (P <0.0001). In IVP, the highest rate of embryonic development (blastocyst and hatched blastocyst) were observed in the C and F group (P = 0.008 and P = 0.0042, respectively). Therefore, removal of seminal plasma by the method of centrifugation reduces the quality of semen cryopreserved cattle, a negative effect on fertility of sperm, and the method of filtering with Sperm Filter® showed better results evidenced by high rates of embryonic development.
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Palavras-chave
Centrifugação, Criopreservação, Filtragem, Plasma seminal, Sêmen, Touro, Centrifugation, Cryopreservation, Filtration, Seminal plasma, Semen, Bull