The aim of this study was to evaluate the effect of different concentrations of trans-resveratrol or quercetin on the ability of goat sperm to withstand being frozen. Six pools of semen obtained from six male goats were processed with different concentrations of resveratrol or quercetin (Experiment 1: 0, 15, 25, 50, 75 or 100µM resveratrol; Experiment 2: 0, 15, 25, 50, 75 or 100µM quercetin) and frozen. After thawing, the semen was evaluated for sperm kinematics, plasma membrane and acrosome integrity, morphology and oxidative stress following 0 and 1h of incubation. Immediately after thawing (0h), wobble (oscillation index) in the groups treated with 100µM of quercetin or resveratrol was lower (P<0.05) than in those treated with 0 and 25µM resveratrol and 0µM quercetin, respectively. After 1h of incubation, the total motility in treatments with 15, 50 and 75µM quercetin, as well as the plasma membrane integrity in all quercetin concentrations were lower (P<0.05) than at 0h. In opposition, the linearity of semen samples treated with 100µM quercetin and the straightness of those treated with 75 and 100µM quercetin were lower (P<0.05) at 0h than at 1h after thawing. Thus, it can be concluded that resveratrol and quercetin at high concentrations (100µM) transiently reduce the wobble of goat sperm submitted to frozen storage, and that quercetin (75 and 100µM) increases the linearity and straightness over time, which can be favorable for fertility.
l-Carnitine (LC) plays a key role in sperm metabolism, easily providing energy through b-oxidation, which positively affects motility. The objective of this study was to investigate the association between blood plasma and seminal plasma LC levels, as well as the effect of LC as an additive in a skimmed milk-based extender during sperm storage at 5°C. In the first experiment, semen and blood samples from 14 Quarter Horse stallions were used. The LC content in blood plasma and seminal plasma was determined by spectrophotometry and their relationships with seminal parameters were evaluated. In the second experiment, ejaculates (n = 16) from four Quarter Horses were used. Each ejaculate was split into four treatment groups with different LC concentrations: 0 (control), 0.5, 1.0, and 2.0 mM. Sperm motility, integrity of plasma and acrosomal membranes, intracellular reactive oxygen species content, and plasma membrane stability were evaluated immediately after samples reached 5°C (0 hour) and after 24, 48, and 72 hours. There was a positive correlation ( p < 0.05) between LC levels in seminal plasma with both sperm concentration and plasma and acrosomal membrane integrity. Furthermore, the addition of LC (1 and 2 mM) preserved the motility of equine sperm stored at 5°C. It was concluded that the concentrations of LC with seminal plasma present correlate to semen parameters and the addition of LC to skimmed milk-based extender preserves the motility of equine sperm stored at 5°C for up to 48 hours.
The addition of antioxidants to semen cryopreservation extenders has been employed for combating oxidative damage. This work aimed to evaluate the addition of carotenoid canthaxanthin to a cryopreservation extender of ram semen. Three breeder rams were used and, after semen collection, with 48-hour intervals between collection, the samples were included in the pool formation (n = 6). The experimental groups comprised 0 (control), 0.1, 1, 10, and 25 μM of canthaxanthin. After thawing (37°C/30 s) and incubation at 37°C for 2 hours, semen aliquots from each group were evaluated for sperm kinetics (CASA), the integrity of the plasma and acrosomal membranes (iPAM), intracellular reactive oxygen species (ROS) production, and lipid peroxidation (LPO) by flow cytometry associated with the image. The control group and canthaxanthin 1 μM after incubation at 37°C for 2 hours showed increases of curvilinear velocity and amplitude of lateral head displacement with decreases of linearity, straightness, and wobble (p < 0.05), which were not observed for the canthaxanthin 10 and 25 μM. The supplementation of a Tris-egg yolk extender with canthaxanthin had no effect on the iPAM, intracellular ROS production in viable spermatozoa, or LPO. In conclusion, supplementation with 10 and 25 μM of canthaxanthin in a Tris-egg yolk extender used for ram semen cryopreservation is able to protect ovine sperm from kinetic changes after incubation at 37°C for 2 hours post-thawing.
The aim of this study was to evaluate the effects of different concentrations of (+)-catechin or (-)-epigallocatechin gallate (EGCG) on goat semen freezability. Poolsof semen were processed (Experiment 1: 0, 15, 25, 50, 75, or 100µM (+)-catechin; Experiment 2: 0, 15, 25, 50, 75, or 100µM EGCG) and frozen. After thawing, the samples were evaluated for kinematics, plasma membrane (PMi) and acrosome integrity, morphology, and oxidative stress, at 0 and 1h. In Experiment 1, at 0h, VSL and VAP were greater (P<0.05) with 15µM than with 50 and 100; WOB was lower (P<0.05) with 100µM than with 0, 15, and 25; and BCF was higher (P<0.05) with 75 and 100µM than with 0. In turn, in Experiment 2, progressive motility was higher (P<0.05) with0 and 15µM than with50 and 75; LIN was lower (P<0.05) with75 and100µM than with0 and 15; WOB was higher (P<0.05) with0 and 15µM; and PMi was greater (P<0.05) with100µM than 0. Thus, (+)-catechin or EGCG at higher concentrations inhibits the kinematics of frozen goat sperm, in a transitory way, and 100µM of EGCG preserves the PMi.
The objective of this study was to investigate the need of seminal plasma removal for short-term cooling of buck semen in soybean lecithin (SL) based extender. Each pool was divided equally, and one half was subjected to centrifugation to remove seminal plasma (SP-), while the other half remained with seminal plasma (SP+). Then, both SP+ and SP-samples were diluted in two SL extenders (extender A = 1% SL; extender B = 2% SL), cooled to 5ºC and stored for 48 hours. The sperm kinetics, evaluated by CASA, and plasma membrane integrity (PMI), acrosomal integrity (ACI) and high mitochondrial membrane potential (HMMP), evaluated by epifluorescence microscopy, were determined within five minutes after reaching 5°C (T0), as well as after 24 (T24) and 48 (T48) hours of storage. Interactions (seminal plasma vs. extender vs. time;) were observed for all variables assessed. Total and progressive motility and other variables of sperm kinetics decreased after 24 hours of cooling in the SP+ group, and after 48 hours of storage, these same variables were lower in SP+/B compared to SP-/B groups. Furthermore, SP+ reduced PMI (extender B, T48), HMMP (A and B extenders, T48) and ACI (extender A, T0) compared to SP-samples. The interactions between seminal plasma and soybean lecithin phospholipids seemed to occur in a time-dependent manner. It was concluded that the removal of seminal plasma improves the quality of goat semen that was cooled in a soybean lecithin-based extender, especially when using 2% soybean lecithin.
RESUMO O objetivo deste estudo foi avaliar o efeito da suplementação do diluidor de congelação de sêmen ovino com o flavonoide miricetina contra os danos ocasionados aos espermatozoides. Oito pools de sêmen, obtidos de quatro reprodutores ovinos, foram congelados com diferentes concentrações de miricetina (0, 1, 10, 100 e 1000nM). Após o descongelamento, o sêmen foi avaliado quanto à cinética espermática, à integridade das membranas plasmática e acrossomal, ao potencial de membrana mitocondrial, aos níveis de ROS intracelular, à peroxidação lipídica e à estabilidade de membrana. Amostras tratadas com miricetina 10nM apresentaram menor percentual de células rápidas (P≤0,05), quando comparadas ao grupo miricetina 1000nM. Amostras do grupo controle apresentaram maior (P≤0,05) VAP que o grupo 10nM de miricetina, enquanto amostras criopreservadas com miricetina (10, 100 e 1000nM) evidenciaram maior (P<0,05) BCF, quando comparadas ao grupo controle. O grupo tratado com miricetina 1000nM apresentou maior percentual (P<0,05) de células com peroxidação lipídica, quando comparado ao grupo controle. Em conclusão, a suplementação do diluidor de criopreservação de sêmen ovino com 10 e 100nM de miricetina afeta a cinética espermática sem provocar alterações na estrutura geral do gameta, enquanto 1000nM de miricetina provoca mudanças na cinética associadas à danos peroxidativos.
The aim of this study was to evaluate the effect of green tea extract (GTE) on the spermatic parameters of Wistar rats, submitted or not to testicular heat shock (HS). For this, 48 animals were treated according to the experimental groups (G1: not exposed to HS and untreated; G2: exposed to HS and untreated; G3: not exposed to HS and treated with GTE; G4: exposed to HS and treated with GTE). Subgroups of rats were euthanized on days 15, 30, and 60 to recover the spermatozoa. The total motility (TM), vigor, spermatic morphology and concentration, mitochondrial membrane potential, plasma membrane integrity, and acrosome integrity (ACi) were analyzed. The TM was higher in G1 and G3 than in G2 and G4 on day 30, and higher in G4 on day 60. The overall means of TM and vigor were higher in G1 and G3 than in G2 and G4, as well as TM on day 60. For the morphology, G2 and G4 were lower than G1 and G3 on day 15, and G4 was lower than G1 and G3 on day 30. Moreover, in G1 and G3 morphology was higher on days 15 and 30, and in G4 it was lower on day 30, with the overall means being higher in G1 and G3 than in G2 and G4, as well as on days 15 and 60 compared to day 30. The overall mean of ACi, on day 30, was lower than on days 15 and 60 for all the groups. Therefore, HS is shown to be widely deleterious to the gametes, and the daily administration of 100 mg/kg green tea extract does not improve the spermatic parameters of Wistar rats, submitted or not to testicular HS, although it leads to better recovery of spermatic motility and morphology at 60 days.
O esôfago é responsável pelo simples e rápido deslocamento da ingesta desde a faringe até o estômago. Sua distribuição anatômica começa dorsalmente à laringe e segue sob a traqueia, desviando-se à esquerda para entrar na cavidade torácica. Nesta região, o esôfago passa pelo mediastino à direita da aorta, dorsalmente à base do coração, penetrando na cavidade abdominal através do hiato diafragmático presente no músculo diafragma (Fubini e Pease, 2004). Inúmeras são as enfermidades que podem comprometer a integridade desse segmento anatômico e alterar a dinâmica do sistema digestivo em ruminantes. São raras as descrições de megaesôfago, mas sabe-se que é o termo utilizado para designar a dilatação esofágica resultante de um esôfago com hipomotilidade ou aperistáltico, secundário a distúrbios neuromusculares, e com obstrução da luz esofagiana. A dilatação é resultante do acúmulo de alimento na luz do esôfago, sendo uma causa comum de regurgitação (Parish et al., 1996).Qualquer enfermidade que comprometa o reflexo nervoso controlador da deglutição ou que obstrua a luz esofagiana pode resultar em dilatação esofágica. Pode ser resultante de anormalidades no desenvolvimento da inervação esofágica ou pela persistência do quarto arco aórtico direito, ou ainda pela ausência de motilidade esofágica gerada por disfunção do nervo vago, que pode ocorrer devido a diversas enfermidades (Ramadan, 1993; Fubini e Pease, 2004).Com relação ao diagnóstico, as esofagiopatias podem ser diagnosticadas por meio de exames radiográficos, esofagoscopia e ultrassonografia, além da observação dos sinais clínicos. Técnicas como a fluoroscopia também podem ser úteis para esclarecer os distúrbios da motilidade. Segundo Ramadan (1993) e Fubini e Pease (2004), a esofagografia, simples e contrastada, é o exame de escolha para a avaliação de lesões ou desordens esofágicas em animais de produção.Poucos são os relatos na literatura sobre essa enfermidade. Já foram identificados alguns casos em caprinos (Ramadan, 1993;Parish et al., 1996), e até o momento, não existe descrição desse tipo de enfermidade em caprinos no Brasil. Relevante considerar que relatos de casos dessa enfermidade são importantes para se conhecer efetivamente a fisiopatologia e as consequências sobre a saúde animal, além da possibilidade de diagnóstico, considerando-se os recursos fornecidos pela imagem. Assim sendo, este trabalho teve o objetivo de descrever aspectos clínicos e procedimentos de diagnóstico de megaesôfago em caprinos por meio do esofagograma simples e contrastado.Foi atendido em Hospital Veterinário Escola um caprino, sem raça definida, fêmea, com
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