Resumo. O processo de criopreservação acarreta estresse oxidativo à célula espermática e a adição de antioxidantes aos meios de refrigeração e congelação de sêmen pode auxiliar na proteção dos espermatozoides contra o dano induzido pelas espécies reativas de oxigênio (ROS), incluindo perda da motilidade de forma irreversível, peroxidação lipídica e fragmentação do DNA, interferindo na capacidade de fertilização do espermatozoide. Objetivou-se com este trabalho avaliar o efeito da adição dos antioxidantes Hidroxi butiltolueno (BHT) e extrato etanólico da casca do caule do mufumbo (Cumbretum leprosum) ao diluidor Botucrio ® para criopreservação de sêmen equino. Foram coletados quatro ejaculados de quatro garanhões da raça Quarto de Milha, por meio de vagina artificial. Após a descongelação das amostras em banho Maria a 37º C foi estimada a taxa de peroxidação lipídica dos espermatozoides pela medida do nível de malonaldeído. O delineamento experimental foi em bloco ao acaso. As variáveis de volume seminal, motilidade espermática, vigor espermático e HOST foram submetidas a análise de variância (ANOVA) utilizando-se o procedimento modelos lineares gerais (Proc GLM) e para comparação de média foi utilizado o teste Tukey, na probabilidade de 5%. A quantificação malonaldeído foi submetida à análise de variância (ANOVA), seguido de Tukey como post hoc teste, na probabilidade de 5%. Não houve diferença, pois o acréscimo de BHT nas concentrações 5, 1,0 e 2,0 mM de BHT e de extrato de mufumbo nas doses de 30, 60 e 120 mg ao diluidor Botucrio não melhorou a qualidade espermática. Desta forma, os antioxidantes não foram determinantes em promover a diminuição formação de radicais livres e, consequentemente, não melhoraram a qualidade do sêmen após a crioinjúria provocando danos à membrana dos espermatozoides. A suplementação nas doses 0,5, 1,0 e 2,0 mM de BHT e de extrato de mufumbo nas doses de 30, 60 e 120 mg ao diluidor botucrio ® não influenciou na qualidade seminal pós criopreservação.
Background: Canine sperm is a very delicate cell that is quite susceptible to oxidative stress since the cytoplasm is restricted and features little antioxidant reserves. Furthermore, the sperm membrane has some polyunsaturated fatty acids sensitive to lipid peroxidation, which makes it important to addition antioxidant substances to the diluter aiming at decreasing such stress to the sperm cell, particularly during seminal cryopreservation. Several antioxidants have been used in this process in some domestic animal’s species, however, the use of palmitic acid has been little reported in works on cryopreservation of semen of the canine species. Hence, this study aimed to assess the effect of addition antioxidants palmitic acid and vitamin E to the Tris-egg yolk diluter on the semen quality of dogs after thawing.Materials, Methods & Results: Samples were collected from the ejaculates of 4 adult dogs, apparently healthy, of the American Pit Bull Terrier breed of kennels in the city of Teresina, PI, places where the pre-freezing procedures of the dog’s semen were performed. The samples were diluted in Tris citric acid fructose (3.28 g Tris-hydroxymethyl-aminomethane, 1.78 g citric acid monohydrate and 1.25 g D-fructose), dissolved in 100 mL distilled water, and added 20% egg yolk and 6% glycerol, at the concentration of 100x106 sptz/mL. The semen samples were divided into 3 mL aliquots to form 3 experimental groups: G1 - Only Tris-egg yolk (Control group); G2 - Tris-egg yolk + 100 µM palmitic acid; and G3 - Tris-egg yolk + 116 µM vitamin E. Semen was collected weekly over a period of little over 2 months. After thawing, thermorresistance test (TTR) was carried out at 0, 30, 60, and 90 min to assess spermatics motility and vigor, in addition to analysis of integrity of plasma membrane, acrosomal membrane and mitochondrial activity of the sperm, using fluorescent probes. These assessments were performed out at the Animal Reproduction Biotechnology Laboratory (LBRA/UFPI). In the TTR, G2 and G3 didn´t exhibit significant results for spermatics motility or vigor when compared with the control group. The palmitic acid and vitamin E also had no significant effects on the parameters of acrosomal membrane integrity or mitochondrial activity. However, sperm cryopreserved with the addition of palmitic acid exhibited significant differences for plasma membrane integrity, providing greater protection to the sperm cells in G2.Discussion: The palmitic acid is one of the most saturated fatty acids in human semen, with reports of great proportions also in the seminal plasma of dogs. Its main role is to protect the plasma membrane from external damage, improving viability and fertility of the sperm after cryopreservation. Data is scarce in the literature on the composition of fatty acids in canine semen and regarding the use of palmitic acid as a seminal antioxidant in that species, which grants further studies aiming to investigate such valuable information for canine reproduction. It is concluded that addition palmitic acid at 100 µM concentration to the Tris-egg yolk diluter was able to preserve the integrity of the plasma membrane during the process of cryopreservation of canine semen.Keywords: dog, semen, antioxidants, cryopreservation.Descritores: cão, sêmen, antioxidantes, criopreservação.
Resumo. O testículo é um órgão que possui função exócrina, relacionada com a produç ão, armazenamento e transporte de espermatozoides, e função endócrina relacionada com a produção de hormônios andrógenos. Na busca por bons reprodutores tem sido avaliado características morfometrias testiculares como a circunferência escrotal, no entanto, somente a mesma não é suficiente para realizar uma seleção, utilizando-se assim de outros parâmetros como volume e comprimento testicular. Tendo isso em vista, o objetivo desta revisão é abordar a influência dos fatores biométricos e sua relação com a morfologia testicular e a qualidade seminal, para a avaliação de reprodutores.
The goal of the present study was to assess the viability of the cryopreserved ovine (Ovis aries) semen, upon supplementation with α-terpineol and rosemary essential oil (Rosmarinus officinalis). The collection of the semen from six rams formed the pool, collected once a week during 7 weeks. The diluted semen was packed into straws (0.25 ml) and frozen in a TK 3000® device. Both α-terpineol and rosemary essential oil were added in the concentrations of 6.25, 12.5 and 25 μg/ml to the TRIS-yolk extender forming six experimental groups; the control group received only the TRIS-yolk extender. The samples were analyzed after thawing regarding motility and vigor, integrity of the plasmatic membrane, thermoresistance test (TT), mitochondrial membrane potential, acrosomal integrity and computer-assisted semen analysis (CASA). The levels of the thiobarbituric acid reactive substances were also analyzed. According to the results obtained with the addition of the concentrations of 6.25, 12.5 and 25 μg/ ml of α-terpineol, significantly reduced the parameters assessed through CASA (VSL, LIN and WOB) and TT. Rosemary essential oil did not have deleterious effects on the spermatozoa and did not reduce the oxidative stress in the concentrations studied, although it presented absolute values higher than those of the control in several parameters. Alpha-terpineol in the concentrations studied was not able to reduce the oxidative stress and had toxic effect over the ovine spermatozoa.
A possible explanation for the lower values after the addition of 50μM arachidonic acid would be its excessive concentration, considering that arachidonic acid, through the action of cyclooxygenase, is converted into prostaglandin E2, inducing the influx of Ca2+ through the membrane. sperm, leading to membrane fusion. It is concluded that the addition of 0.5μM and 5μM of arachidonic acid to the extender maintained the integrity of the acrosome membrane in post-cryopreservation goat semen, being beneficial for the process of sperm capacitation and fertilization, suggesting further research aiming at a greater accuracy of the concentrations of arachidonic acid as an antioxidant.
Investigação e medidas de diagnóstico da brucelose bovina no alto sertão sergipanoResearch and diagnostic measures for bovine brucellosis in the high sertão sergipano
It was aimed to compare semen and ions concentrations in the prostatic fluid of German Shepherd and Rottweiler dogs. Five semen collections of each dog (5 / breed) were accomplished and the following parameters assessed: total motility, vigor, concentration, percentage of sperm alive and normal morphology, as well as pH, cellular distribution and concentrations of calcium, phosphorus and magnesium were evaluated in prostatic fluid. Rottweiler dogs presented higher motility, concentration, percentage of alive Sperm and normal morphology than GS dogs. However, these dogs demonstrated higher levels of calcium in prostatic fluid. Differences were not observed concerning sperm vigor, phosphorus and magnesium concentrations, pH or the cellular distribution in prostatic fluid. It was concluded that Rottweiler dogs present better semen quality than German Shepherd dogs and a smaller liberation of calcium in prostatic fluid.
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