Adição do complexo ciclodextrina-colesterol na criopreservação do sêmen caprino

• Autor(a) principal: Santos, Madriano Christilis da Rocha
• Data de Publicação: 2013
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/1869
• topic: Colesterol; Caprino; Sêmen; Ciclodextrina; Cholesterol; Goats; Semen; Cyclodextrin; CNPQ::CIENCIAS AGRARIAS::ZOOTECNIA

The objective of this study was to analyze by spermatic motility and vigor the accuracy of six different of Ciclodextrin-Cholesterol Compound (CCC) concentrations on the spermatic motility and vigor effect after goat semen cryopreservation process and to evaluate the plasma membrane function and integrity of thawed goat semen. Chapter 2 The objective of this study was to evaluate the functionality and integrity of plasma membrane of thawed goat semen after addition of Ciclodextrin-Cholesterol Compound (CCC) diluted in tris glycerol or tris ethylene glycol + yolk hen s egg diluent to goat semen. And further, to evaluate the way of inclusion of the CCC (together with diluent or preincubated for 15 minutes). Chapter 1 Five Billy goats were used and five samples were collected from each and then fractionated in seven aliquots. One aliquot did not receive the CCC been considered a negative control (Tcontrol), the others received the CCC in the following concentrations: 0.5; 1.0; 1.5; 2.0; 2.5; 3.0 mg for spermatic concentration of [ ] = 200 * 106/mL, being called treatments: T0.5; T1.,0; T1.5; T2.0; T2.5; and T3.0, respectively. The spermatic motility and vigor and the supravital and hypoosmotic test were applied to the post-thaw semen. Chapter 2 Four billy goats were used and three samples were collected from each with a total of twelve ejaculates. Each ejaculate was evaluated and the ones good to be frozen by the minimum requirements of motility and vigor by the Brazilian College of Animal Reproduction (CBRA, 1998) were fractionated in six aliquots of 350 * 106 spermatozoa (enough for 7 doses with 50 * 106 spermatozoa/semen straw). Each aliquot received 1.0 mg of CCC in the following treatment: TGlycerol negative control for Tris glycerol diluent without CCC; TGlycerol + CCC Tris glycerol diluent + CCC; TGlycerol + CCC + preincubated CCC diluted in isosmotic solution (normal saline) 15 minutes before addittion of the Tris glycerol diluent; TEthyleno diluent without CCC;TEthylene glycol negative control for Tris ethyleno glycol glycol + CCC Tris ethylene glycol + yolk hen s egg diluent + CCC; TEthylene glycol + CCC + preincubated CCC diluted in isosmotic solution (normal saline) 15 minutes before addition of the Tris ethylene glycol + yolk hen s egg diluent. Chapter 1 Treatments were considered to be different if (P < 0.05). The spermatic motility was affected by all treatments (P<0.05), T1.0 e T0.5 values were higher (33.45% ± 1.07 and 32.59 ± 0.98, respectively) than 28.62 ± 1.05 for the Tcontrol and above the minimum value of 30%, required by CBRA (1998) for thawed semen. The motility for treatments: T1.5; T2.0; T2.5 and T3.0 were 27.41 ± 0.96; 26.55% ± 0.94; 25.89% ± 0.99 e 25.17% ± 1.06, respectively. The sperm vigor was not affected by the treatments(P>0.05), and their mean values were:Tcontrol(2.1 ± 0.07), T0.5(2.24 ±0.07), T1.0(2.4 ±0.08), T1.5(2.28 ±0.07), T2.0(2.13 ±0.07), T2.5(2.19 ±0.07), T3.0(2.26±0.09). The percentage of cells with functional membrane in the supravital test in T0.5 (41.5 ± 0.85) was higher (P<0.05) than Tcontrol (37.7 ± 0.94). The percentage of plasma membrane functionality did not change (P>0.05) among the treatments T2.0; T2.5; T1.0; T3.0 (38.06 ± 0.91; 38.06 ± 1.0; 36.33 ± 0.81; 35.54 ± 0.81; 34.77 ± 1.13, respectively) as compared to Tcontrol ones. The percentage of plasma membrane functionality for treatment T1.5 (30.65 ± 0.98) was lower (P<0.05) than all the others. No difference was found (P>0.05) in the integrity of plasma membrane by hypoosmotic test among Tcontrol (28.55 ± 1.01), T2.5 (24.47 ± 0.76), T0.5 (24.21 ± 0.82), T1.0 (23.74 ± 0.85), T1.5 (21.92 ± 1.18), T2.0 (21.24 ± 0.87), T3.0 (19.68 ± 0.88). Chapter 2 It was found difference among the averages of all treatments for the spermatic motility (P<0.05) TGlycerol preincubated (37.08 ± 2.17), TGlycerol TEthylene glycol + CCC + preincubated TEthylene glycol + CCC + CCC + (35.00 ± 1.51), TGlycerol (29.55 ± 2.81), (29.17 ± 4.52), TEthylene glycol + CCC (25.83 ± 3.98), (23.64 ± 4.10). The vigor of thawed semen was not affected by treatments used (P>0.05), whose averages are: TEthyleno glycol + CCC + preincubated (2.96± 0.31), TEthyleno glycoll + CCC (2.91 ± 0.12), TGlycerol + CCC + preincubated (2.75 ± 0.2), T Ethyleno glycol (2.59 ± 0.35), TGlycerol + CCC (2.29 ± 0.18), TGlycerol (2.22 ± 0.18). No difference was found (P>0.05) on the percentage plasma membrane integrity by hypoosmotic test on treatments TGlycerol CCC/preincubated (49.65 ± 5.9), TGlycerol + CCC/preincubated + CCC (52.0 ± 4), TEthylene glycol+ (45.05 ± 4.83), TGlycerol (44.8 ± 2.31), TEthylene glycol (44.39 ± 4.66), TEthylene glycol + CCC (39.6 ± 4.96). The functionality of the plasma membrane by supravital test was not affected by the treatments (P>0.05). The treatments averages were: TEthylene 3.82), TEthylene glycol + CCC (26.96 ± 4.73), TGlycerol glycol+ CCC/preincubated + CCC/preincubated (29.0 ± (24.5 ± 4.2), TGlycerol + CCC (23.54 ± 4.49), TGlycerol (20.79 ± 3.35), TEthylene glycol (19.45 ± 4.08). The ATP production by mitochondrial sheath was affected by treatmens (P<0.05) and theirs means were: TEthylene glycol + CCC (62.92 ± 8.34), TEthylene glycol+ CCC/preincubated (62.72 ± 8.79), TEthylene glycol (59.19 ± 9.57), TGlycerol + CCC/preincubated (11.92 ± 6.89), TGlycerol (11.64 ± 5.43), TGlycerol + CCC (7.76 ± 3.31). The acrosomal glycol (78.19 ± 5.82), glycol + CCC (70.68 ± 5.77), membrane integrity means by treatments were: TEthylene TEthylene glycol+ CCC/preincubated (74.43 ± 6.31), TEthylene TGlycerol (47.06 ± 4.31), TGlycerol + CCC (46.88 ± 6.58), TGlycerol + CCC/preincubated (44.6 ± 8.57) and were influenced by the treatments (P<0.05). The plasma membrane integrity by fluorescent probes (Propidium iodide + diethyl carboxyfluorescein) were not affected by treatments (P>0.05) and theirs means were: TGlycerol (63.98 ± 8.4), TGlycerol + CCC (63.08 ± 9.05), TGlycerol + CCC/preincubated (52.24 ± 9.47), T5 (52.09 ± 10.88), TEthylene glycol+ CCC/preincubated (44.37 ± 8.96), TEthylene glycol (37.19 ± 10.81).Chapter 1 It is concluded that the addition of 0.5 mg of CCC at spermatic concentration of 200 * 106/mL reduced the loss of integrity and functionality of the plasma membrane. However, the addition of 1.0 mg of CCC at the spermatic concentration 200 * 106/ mL showed better results for sperm cell motility in post-thawed semen. Chapter 2 It is concluded based on the results that the addittion of 1.0 mg of CCC on the spermatic concentration of 200 * 106/ mL did not affected the integrity neither the functionality of plasma membrane of goat sperm. The Tris + Ethyleno glycol + yolk egg hen s yolk diluent did not alter the functionality of plasma membrane or vigor.. Although it has a positive influence on the acrossomal membrane integrity, negatively affects the motility of the thawed sperm compared to solvent-based Tris- glycerol. The preincubation of sperm with CCC does not alter the characteristics of sperm vigor, and integrity and functionality of the plasma membrane, but positively affect the motility of the thawed sperm.