Medium and time conservation affect follicular morphology and superoxide dismutase enzyme activity of bovine preantral follicles during storage at 4ºC / Meio e tempo de conservação afetam a morfologia folicular e atividade da enzima superóxido dismutase de folículos pré-antrais bovinos durante a conservação a 4 ºC

The aim of this study was to evaluate the morphology and superoxide dismutase enzyme (SOD) activity of bovine preantral follicles (PFs) preserved in TCM 199, saline solution or PBS at different conservation periods. Cow ovaries (n=6) were divided into 7 fragments. One small piece of each ovarian fragment was randomly removed to evaluate SOD activity, while the remainder was immediately fixed for morphological evaluation as a control group. The other 6 fragments were randomly distributed in tubes containing TCM 199, saline solution, or PBS and maintained at 4ºC for 6 or 24 h. For histological evaluation, the fragments were fixed in Carnoy and stained with PAS-hematoxylin, following being classified PFs in relation to their follicular morphology in normal or degenerated. Determination of SOD activity was based on the ability to inhibit autoxidation of adrenaline in adrenochrome. Evaluation of follicular morphology showed that follicles preserved in TCM 199 for 6 h did not differ from the control (P > 0.05). In contrast, preservation in saline solution and PBS for 6 or 24 h and TCM 199 for 24 h decreased normal PFs compared to the control (P < 0.05). SOD showed a lower activity in ovarian cortical tissue kept in TCM 199 for 6 h and saline solution for 24 h than in the other groups. Our study shows that incubation using TCM 199 at 4°C for 6 h can be used to efficiently conserve female bovine PFs in situ.(AU)

O objetivo desse estudo foi avaliar a morfologia e atividade da enzima superóxido dismutase (SOD) de folículos pré-antrais bovinos (PFs) preservados em TCM 199, solução salina ou PBS por diferentes períodos de conservação. Ovários de vacas (n=6) foram divididos em 7 fragmentos. Um pequeno pedaço de cada fragmento ovariano foi removido para avaliar a atividade da SOD enquanto que o restante foi imediatamente fixado para avaliação morfológica como grupo controle. Os outros 6 fragmentos foram distribuídos aleatoriamente em tubos contendo TCM 199, solução salina ou PBS e mantidos a 4ºC por 6 ou 24 h. Para avaliação histológica, os fragmentos foram fixados em Carnoy e corados com PAShematoxilina, sendo classificados em seguida os PFs em relação à sua morfologia folicular em normal ou degenerados. A determinação da atividade da SOD foi baseada na capacidade de inibir a autooxidação da adrenalina no adrenocromo. A avaliação da morfologia folicular mostrou que os folículos preservados em TCM199 por 6 h não diferiram do controle (P > 0,05). Em contraste, a preservação em solução salina e PBS por 6 ou 24 h e TCM 199 por 24 h diminuiu os PFs normais em comparação com o controle (P < 0,05). A SOD mostrou uma menor atividade no tecido cortical ovariano mantido em TCM 199 por 6 h e solução salina por 24 h do que nos outros grupos. Nosso estudo mostra que a incubação usando TCM 199 a 4° C por 6 h pode ser usada eficientemente para conservar PFs de fêmeas bovinas in situ.(AU)

Protective role of Syzygium cumini leaf extracts against paraquat-induced oxidative stress in superoxide-dismutase-deficient Saccharomyces cerevisiae strains

The aim of this study was to evaluate the protective effect of three different extracts prepared from Syzygium cumini leaves against paraquat-induced toxicity in Saccharomyces cerevisiaestrains deficient in superoxide dismutase (SOD). Additionally, the extracts phenolic and flavonoid contents, in vitro antioxidant activity, and phytochemical composition (using high-pressure liquid chromatography) were determined. Bioactive compounds from S. cumini leaves were extracted with infusion (traditional method) or ultrasound (aqueous or hydroalcoholic). Compared to the infusion extract, the ultrasound extracts exhibited a greater protective capacity against paraquat toxicity in the yeast cells as well as higher antioxidant activity. These results may be directly related to the higher phenolic and flavonoid contents in these extracts, since they are recognized as having high antioxidant actions.

Powdered coconut water (ACP 406®) as an alternative base culture medium for in vitro culture of goat preantral follicles enclosed in ovarian tissue

This study evaluated a powdered coconut water solution (ACP 406®) as a base culture medium on the in vitro survival and development of in situ goat preantral follicles. The ovarian fragments were either immediately fixed in Carnoy solution (non-cultured control) or individually cultured for 2 or 6 days. The following culture media (all containing 100 μg/mL penicillin and 100 μg/mL streptomycin) were evaluated: α-MEM (α-MEM alone, without additional supplementation); α-MEM+ (supplemented α-MEM); ACP (ACP®406 alone); or ACP+ (supplemented ACP®406). Additional supplementation includes: 1.25 mg/mL bovine serum albumin, 10 μg/mL insulin, 5.5 μg/mL transferrin, 5 ng/mL selenium, 2 mM glutamine, and 2 mM hypoxanthine. The endpoints (i) follicular morphology; (ii) development; (iii) estradiol production; and (iv) reactive oxygen species (ROS) were recorded. Data were analyzed using chi-square, Turkey, t-test or One-Way ANOVA. Differences were considered significant when P < 0.05. At day 2 of culture, a greater (P < 0.05) percentage of morphologically normal follicles was observed between ACP+ and ACP treatments. Moreover, at day 2 of culture, no hormonal difference (P < 0.05) was observed between ACP+ and both α-MEM treatments. At day 6 of culture when ACP and α-MEM treatments were compared the percentage of healthy follicles were similar (P > 0.05) among treatments. Overall, all treatments had lower primordial follicles (P < 0.05) accompany by greater developing follicles (P < 0.05) percentages than non-cultured control treatment, indicating primordial follicle activation. However, at day 6 of culture, the percentage of primordial follicle development were similar (P > 0.05) among the treatments. Likewise, no differences (P > 0.05) were observed for ROS production and follicular and oocyte diameters among treatments. Therefore, ACP+ has the equivalent efficiency to MEM+ in maintaining the survival and development of goat preantral follicles, representing an alternative plant-based low-cost culture medium for in vitro culture.(AU)

Powdered coconut water (ACP 406®) as an alternative base culture medium for in vitro culture of goat preantral follicles enclosed in ovarian tissue

This study evaluated a powdered coconut water solution (ACP 406®) as a base culture medium on the in vitro survival and development of in situ goat preantral follicles. The ovarian fragments were either immediately fixed in Carnoy solution (non-cultured control) or individually cultured for 2 or 6 days. The following culture media (all containing 100 μg/mL penicillin and 100 μg/mL streptomycin) were evaluated: α-MEM (α-MEM alone, without additional supplementation); α-MEM+ (supplemented α-MEM); ACP (ACP®406 alone); or ACP+ (supplemented ACP®406). Additional supplementation includes: 1.25 mg/mL bovine serum albumin, 10 μg/mL insulin, 5.5 μg/mL transferrin, 5 ng/mL selenium, 2 mM glutamine, and 2 mM hypoxanthine. The endpoints (i) follicular morphology; (ii) development; (iii) estradiol production; and (iv) reactive oxygen species (ROS) were recorded. Data were analyzed using chi-square, Turkey, t-test or One-Way ANOVA. Differences were considered significant when P 0.05) among treatments. Overall, all treatments had lower primordial follicles (P 0.05) among the treatments. Likewise, no differences (P > 0.05) were observed for ROS production and follicular and oocyte diameters among treatments. Therefore, ACP+ has the equivalent efficiency to MEM+ in maintaining the survival and development of goat preantral follicles, representing an alternative plant-based low-cost culture medium for in vitro culture.

Platelet-derived growth factor-BB (PDGF-BB) improves follicular survival, oocyte and follicular diameters, in a dose-dependent manner, after the in vitro culture of goat preantral follicles enclosed in ovarian tissue fragments

The aims of this study were to investigate the effects of different concentrations of Platelet-derived growth factor-BB (PDGF-BB) on the survival, activation, levels of ROS, and growth of goat preantral follicles enclosed in ovarian tissue. For this, ovarian fragments were cultured for 7 days in Alpha Minimum Essential Medium (α-MEM+ ) with or without PDGF-BB (0, 25, 50 and 100 ng/ml). The results showed that both the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments maintained the percentage of morphologically normal follicles from day 1 to day 7. In addition, the 25 ng/ml PDGF treatment showed a significantly higher percentage of morphologically normal follicles when compared to the other treatments. At day 7, greater (P < 0.05) follicular and oocyte diameters were observed in the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments when compared to the cultured control treatment. On day 7 of culture, all the treatments tested had a significant increase in the percentage of developing follicles when compared to the non-cultured control. However, the percentage of follicle activation, as well as ROS production, were similar (P < 0.05) among the treatments, irrespective of culture time. In conclusion, PDGF-BB improved, in a concentration-dependent manner, follicular survival as well as oocyte and follicular diameter after in vitro culture of goat preantral follicle-enclosed in ovarian tissue fragments.

Platelet-derived growth factor-BB (PDGF-BB) improves follicular survival, oocyte and follicular diameters, in a dose-dependent manner, after the in vitro culture of goat preantral follicles enclosed in ovarian tissue fragments

The aims of this study were to investigate the effects of different concentrations of Platelet-derived growth factor-BB (PDGF-BB) on the survival, activation, levels of ROS, and growth of goat preantral follicles enclosed in ovarian tissue. For this, ovarian fragments were cultured for 7 days in Alpha Minimum Essential Medium (α-MEM+ ) with or without PDGF-BB (0, 25, 50 and 100 ng/ml). The results showed that both the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments maintained the percentage of morphologically normal follicles from day 1 to day 7. In addition, the 25 ng/ml PDGF treatment showed a significantly higher percentage of morphologically normal follicles when compared to the other treatments. At day 7, greater (P < 0.05) follicular and oocyte diameters were observed in the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments when compared to the cultured control treatment. On day 7 of culture, all the treatments tested had a significant increase in the percentage of developing follicles when compared to the non-cultured control. However, the percentage of follicle activation, as well as ROS production, were similar (P < 0.05) among the treatments, irrespective of culture time. In conclusion, PDGF-BB improved, in a concentration-dependent manner, follicular survival as well as oocyte and follicular diameter after in vitro culture of goat preantral follicle-enclosed in ovarian tissue fragments.(AU)