Insights into soy lecithin and egg yolk-based extenders for chilling canine spermatozoa.

SummaryThe aim of this study was to compare different concentrations of soy lecithin (LEC0.01%, LEC0.05% and LEC0.1%) with egg yolk (Control) in cooling extenders during the storage of semen at 5ºC for 5 days. Twelve dogs (n = 12) were selected, and semen was cooled and assessed after 2, 24, 48, 72, 96 or 120 h. At each time point, sperm were analyzed for kinetic patterns (using computer-assisted sperm analysis), mitochondrial activity (3'3- diaminobenzidine assay), lipid peroxidation (TBARS assay), DNA fragmentation (SCSA®) and plasma and acrosome membrane integrity (eosin/nigrosin and fast green/rose Bengal stains, respectively). The Control group (1814.4 ± 197.2) presented the highest rates of lipid peroxidation at 120 h. Conversely, progressive motility (42.8 ± 4%), linearity (45.4 ± 1%), and VAP (88 ± 3%) were higher in the Control group. In addition, there was lower mitochondrial activity in the Control group at 72 h. Therefore, our data show that lecithin used at these concentrations was not able to maintain sperm viability at as high qualities as would egg yolk. Moreover, the decrease in high mitochondrial activity and the persistence of sperm motility may indicate a compensatory mechanism in canine spermatozoa (i.e., glycolytic pathway). Furthermore, these higher lipid peroxidation indexes could indicate the necessity for future therapy using extenders and antioxidants over a long cooling time for dog sperm.

Análise morfológica de amostras de útero de cães fixados em formaldeído em diferentes concentrações e durações de tempo / Comparative morphological analysis of uterus of dogs in fixed buffered formaldehyde in different concentrations and time of fixation

Objetivo - A fixação é um dos pontos críticos para a realização de diversas técnicas laboratoriais, sendo o formaldeído amplamente utilizado na rotina e pesquisa científica com a finalidade de preservar a integridade morfológica das amostras especialmente para análise pela microscopia de luz. Mesmo com sua ampla utilização, não existem dados na literatura com protocolos padronizados que assegurem o tempo mínimo de fixação assim como a concentração ideal de formaldeído a ser utilizada. Métodos - Neste estudo foram analisadas amostras de útero de 20 cadelas hígidas, sem raça definida (SRD) e em idade reprodutiva (1-5 anos), fragmentados a uma espessura de 0,5 cm3. As amostras foram coletadas por cirurgia eletiva de ovário-salpingo-histerectomia (OSH), fixadas em formaldeído tamponado nas concentrações de 4%, 8% e 10% e retiradas nos tempos de 12, 24, 48 e 72 horas pós-fixação. Seguiu-se o processamento histológico de rotina e a análise pela microscopia de luz na coloração de hematoxilina e eosina (H/E) em diferentes aumentos. Resultados - A análise morfológica qualitativa pela microscopia de luz não demonstrou alterações morfológicas significativas entre as amostras fixadas em diferentes tempos e concentrações de formaldeído. Conclusão - Estes resultados corroboram com a rotina e a pesquisa permitindo a padronização do tempo mínimo de fixação de amostras (com espessura de até 0,5 cm3) em formaldeído para 12 horas quando utilizado nas concentrações 4%, 8% e 10%.

Objective - Fixation is a critical point for the performance of several diagnostic techniques, such as immunohistochemistry and DNA analysis. Formaldehyde is the fixation solution most commonly used in laboratories to preserve the morphological integrity of tissue. However, there are no reports of a protocol that ensures the minimum time and optimal concentration of formaldehyde. Methods - Analysis of the preservation of the uterus of 20 dogs with a fixed solution of buffered formaldehyde 4%, 8%, and 10%, during 12, 24, 48 and 72 hours of fixation. The uterus, included in paraffin and stained with hematoxylin-eosin, was evaluated in light microscopy. Results - The fixation with formaldehyde doesnït showed significant alteration in the tissue architecture of the uterus. Conclusion - This comparison indicates that the formaldehyde is a suitable solution for the system preservation, allowing its use in routine laboratory asa good protocol of fixation.