Efeito de antioxidantes aos espermatozoides de equinos submetidos ao estresse térmico
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Data
2010-10-05
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Universidade Federal de Goiás
Resumo
The evaluation of the reproductive capacity of stallions and semen cooled and / or frozen is of fundamental importance in practical breeding of horses. Whereas predicting the fertility of a stallion is still a subjective decision, the present study was conducted to evaluate different staining techniques, as well as tests that assess the viability of semen in horses, studying the effects of the addition of cysteine and glutathione in plasma membrane integrity of sperm DNA, and to evaluate the effects of the addition of these antioxidants in preserving the viability of sperm undergoing incubation and refrigerated for short periods. We used semen from six stallions, wich were split into seven samples, one kept as control (Control Group) and other antioxidants cysteine was added at a ratio of 1.0 mM (Group C1, 0), 1.5 mM (Group C1, 5), 2.5 mM (Group C 2.5) and glutathione also at 1.0 mM (Group G1, 0), 1.5 mM (Group G1, 5) and 2.5 mM (Group G 2.5 ). All tests were performed every six hours. The three treatments were: Treatment I cooled to 12 ⁰C for 12 hours (zero hour; 6h resf.; 12h resf.) Chilled in boxes; Treatment II incubated in a water bath at 37 ⁰C for 12 hours (6h 37 ° C, 12h 37 ° C) and Treatment III - cooled and subsequently incubated, according to the treatments I and II. The evaluation of plasma membrane integrity was performed by staining eosine-nigrosin; acrosomal membrane by FITC-PSA and DNA by acridine orange. For each analysis were numbered 200 to 500 cells. The evaluation of the viability of sperm was performed by MTT assay according to Mosmann (1983).
The results showed that antioxidants were effective (p <0.05) in keeping the DNA intact chromatin, especially glutathione. In the acrosome of antioxidants were protective, at times 18 and 24 hours, and in other treatments, no significant difference (p> 0.05) between control and treated groups. The MTT test showed that groups treated with antioxidants had absorbance values similar to those of control, showing positive effect (p <0.05) only when cooled by six o'clock in the cysteine group 2.5. In relation to the plasma membrane of spermatozoa stained with eosin-nigrosin, no protective effect of antioxidants in the samples. The values of their averages were close to the control group (p> 0.05). One factor was estimated that cooling per se, independent of the addition of antioxidants used, has been effective in protecting the sperm. And the incubation at 37 ⁰ C causes these cells, and the addition of cysteine and glutathione were efficient, if not protect, but to maintain the integrity of the factors evaluated, not causing more damage to sperm.
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Palavras-chave
cisteina , glutationa , MTT , FITC-PSA , laranja de acridina , espermatozóide , garanhão , cystein , glutathione , MTT , FITC-PSA , acridine orange , spermatozoa , stallion
Citação
OLIVEIRA, Aline França Dias. Effect of antioxidants on stallion spermatozoa under heat stress. 2010. 76 f. Dissertação (Mestrado em Ciências Biolóicas) - Universidade Federal de Goiás, Goiânia, 2010.