Steady-state levels of vasoactive intestinal peptide (VIP) mRNA in goat ovaries and the effect of VIP on the in vitro development of isolated preantral follicles

The aims of this study were to verify the steady-state level of vasoactive intestinal peptide (VIP) mRNA in goat follicles at various developmental stages and to investigate the influence of VIP on the survival, antrum formation and growth of secondary follicles cultured for 6 days. Primordial, primary and secondary goat follicles and small and large antral follicles were obtained to quantify VIP mRNA by real-time reverse transcription with the polymerase chain reaction. The influence of VIP and the presence or absence of follicle-stimulating hormone (FSH) on the development of secondary follicles and on mRNA expression for VIP and FSH receptor (FSHR) were determined after 6 days of culture. Survival, antrum formation and follicular diameter were evaluated every other day of culture. The levels of VIP mRNA in primary and secondary follicles were significantly higher than in primordial follicles. Cumulus–oocyte complexes (COCs) from both small and large antral follicles had significantly higher levels ofVIP mRNA than their respective granulosa/theca cells. During culture, the addition of VIP and/or FSH had o effect on follicular development. However, the presence of FSH and/or VIP in the culture medium significantly reduced VIP mRNA levels, but did not alter FSHR mRNA levels. In conclusion, VIP mRNA was detected in all goat follicular categories and cellular types, VIP and/or FSH did not affect the development of secondary follicles and reduce the expression of VIP mRNA levels.(AU)

Steady-state levels of vasoactive intestinal peptide (VIP) mRNA in goat ovaries and the effect of VIP on the in vitro development of isolated preantral follicles

The aims of this study were to verify the steady-state level of vasoactive intestinal peptide (VIP) mRNA in goat follicles at various developmental stages and to investigate the influence of VIP on the survival, antrum formation and growth of secondary follicles cultured for 6 days. Primordial, primary and secondary goat follicles and small and large antral follicles were obtained to quantify VIP mRNA by real-time reverse transcription with the polymerase chain reaction. The influence of VIP and the presence or absence of follicle-stimulating hormone (FSH) on the development of secondary follicles and on mRNA expression for VIP and FSH receptor (FSHR) were determined after 6 days of culture. Survival, antrum formation and follicular diameter were evaluated every other day of culture. The levels of VIP mRNA in primary and secondary follicles were significantly higher than in primordial follicles. Cumulus–oocyte complexes (COCs) from both small and large antral follicles had significantly higher levels ofVIP mRNA than their respective granulosa/theca cells. During culture, the addition of VIP and/or FSH had o effect on follicular development. However, the presence of FSH and/or VIP in the culture medium significantly reduced VIP mRNA levels, but did not alter FSHR mRNA levels. In conclusion, VIP mRNA was detected in all goat follicular categories and cellular types, VIP and/or FSH did not affect the development of secondary follicles and reduce the expression of VIP mRNA levels.

Papel dos antioxidantes no cultivo in vitro de células ovarianas / The role of antioxidants in the in vitro culture of ovarian cells

Radicais livres são moléculas que contêm elétrons desemparelhados na camada de valência atuando na defesa do organismo contra agentes estranhos, podendo ser importantes marcadores da remodelação dos tecidos. Entretanto, um desequilíbrio entre sua produção e neutralização pode levar a danos celulares, denominados estresse oxidativo. Os agentes neutralizantes dessas moléculas são conhecidos como antioxidantes e podem ser de natureza enzimática ou não. Tendo em vista que o metabolismo oxidativo é essencial para a produção de energia em gametas e embriões, a produção de radicais livres é inevitável. Alguns agentes antioxidantes têm sido adicionados aos meios de cultivo de células ovarianas a fim de reduzir o estresse oxidativo. Sendo assim, a presente revisão objetiva descrever os principais antioxidantes e suas funções em compartimentos celulares com ênfase nas células ovarianas.(AU)

Free radical is any molecule containing unpaired electrons in the valence shell, which protects the body against foreign agents and can act as important markers in tissue remodelation. Nevertheless, an imbalance between production and neutralization of these factors can damage cells in a process named oxidative stress. Neutralizing agents for these molecules are known as antioxidants, which can be enzymatic in nature or not. Since oxidative metabolism is essential for energy production in gametes and embryos, free radicals generation is unavoidable. Some antioxidant agents have been employed in culture media for ovarian cells. In this context, the aim of present review is to describe the main antioxidants and their functions within cellular compartments, with focus on ovarian cells.(AU)

Papel dos antioxidantes no cultivo in vitro de células ovarianas / The role of antioxidants in the in vitro culture of ovarian cells

Radicais livres são moléculas que contêm elétrons desemparelhados na camada de valência atuando na defesa do organismo contra agentes estranhos, podendo ser importantes marcadores da remodelação dos tecidos. Entretanto, um desequilíbrio entre sua produção e neutralização pode levar a danos celulares, denominados estresse oxidativo. Os agentes neutralizantes dessas moléculas são conhecidos como antioxidantes e podem ser de natureza enzimática ou não. Tendo em vista que o metabolismo oxidativo é essencial para a produção de energia em gametas e embriões, a produção de radicais livres é inevitável. Alguns agentes antioxidantes têm sido adicionados aos meios de cultivo de células ovarianas a fim de reduzir o estresse oxidativo. Sendo assim, a presente revisão objetiva descrever os principais antioxidantes e suas funções em compartimentos celulares com ênfase nas células ovarianas.

Free radical is any molecule containing unpaired electrons in the valence shell, which protects the body against foreign agents and can act as important markers in tissue remodelation. Nevertheless, an imbalance between production and neutralization of these factors can damage cells in a process named oxidative stress. Neutralizing agents for these molecules are known as antioxidants, which can be enzymatic in nature or not. Since oxidative metabolism is essential for energy production in gametes and embryos, free radicals generation is unavoidable. Some antioxidant agents have been employed in culture media for ovarian cells. In this context, the aim of present review is to describe the main antioxidants and their functions within cellular compartments, with focus on ovarian cells.

Ascorbic acid improve the survival and in vitro growth of isolated caprine preantral follicles

The present study aims to investigate the influence of two concentrations of ascorbic acid on the survival, growth , antral formation and m RNA expression of the matrix metalloproteinases - 9 (MMP - 9) and their tissue inhibitor - 2 (TIMP - 2) on caprine preantral follicles during long - term in vitro culture. Isolated preantral follicles were individ ually cultured without or with ascorbic acid at 50 μ g/ml (AA50) or 100 μ g/ml (AA100) during 18 days. The parameters evaluated were follicular viability, growth, antrum formation and extruded oocytes. The genes MMP - 9 and TIMP - 2 were quantified by real - time polymer ase chain reaction (qPCR) after 18 days of culture in the control medium (MEM + ) or ascorbic acid (50 or 100 μ g/ml) and in fresh control (non cultured) . At the end of culture, AA50 significantly increased the percentage of viable follicles compared w ith other treatments . Moreover, mean daily increase in follicular diameter (μm/day) was significantly higher in the presence of both concentrations of ascorbic acid than in MEM + alone. Higher rates of antral formation and lower percentages of extruded oocy tes were observed in medium containing AA50 compared with control medium. Real Time RT - PCR assays showed that AA50 increase s MMP - 9 expression significantly compared with fresh control and MEM + alone. In conclusion, ascorbic acid at 50 μ g/ml was very import ant for the maintenance of caprine preantral follicle viability and development after in vitro culture and influences in vitro the enzymes involved with basement membrane remodeling.

Ascorbic acid improve the survival and in vitro growth of isolated caprine preantral follicles

The present study aims to investigate the influence of two concentrations of ascorbic acid on the survival, growth , antral formation and m RNA expression of the matrix metalloproteinases - 9 (MMP - 9) and their tissue inhibitor - 2 (TIMP - 2) on caprine preantral follicles during long - term in vitro culture. Isolated preantral follicles were individ ually cultured without or with ascorbic acid at 50 μ g/ml (AA50) or 100 μ g/ml (AA100) during 18 days. The parameters evaluated were follicular viability, growth, antrum formation and extruded oocytes. The genes MMP - 9 and TIMP - 2 were quantified by real - time polymer ase chain reaction (qPCR) after 18 days of culture in the control medium (MEM + ) or ascorbic acid (50 or 100 μ g/ml) and in fresh control (non cultured) . At the end of culture, AA50 significantly increased the percentage of viable follicles compared w ith other treatments . Moreover, mean daily increase in follicular diameter (μm/day) was significantly higher in the presence of both concentrations of ascorbic acid than in MEM + alone. Higher rates of antral formation and lower percentages of extruded oocy tes were observed in medium containing AA50 compared with control medium. Real Time RT - PCR assays showed that AA50 increase s MMP - 9 expression significantly compared with fresh control and MEM + alone. In conclusion, ascorbic acid at 50 μ g/ml was very import ant for the maintenance of caprine preantral follicle viability and development after in vitro culture and influences in vitro the enzymes involved with basement membrane remodeling.(AU)

Sistemas de cultivo in vitro para o desenvolvimento de oócitos imaturos de mamíferos / Systems in vitro development of immature oocytes of mammalians

O ovário mamífero possui um grande estoque de folículos primordiais, que é uma fonte potencial de oócitos para a produção in vitro de embriões. Sendo assim, o desenvolvimento de sistemas de cultivo in vitro que explorem o grande número de oócitos imaturos oriundos de folículos pré-antrais de ovários mamíferos torna-se importante para maximizar o potencial reprodutivo das fêmeas. A presente revisão aborda os aspectos relacionados aos sistemas de cultivo in vitro para desenvolvimento de oócitos imaturos oriundos de folículos pré-antrais, bem como às técnicas para analisar sua eficiência. (AU)

The mammalian ovary has a large population of primordial follicles, which is a potential source of oocytes useful for embryo in vitro production. Due to this potential, the development of in vitro culture systems to exploit the large number of immature oocytes from the preantral follicles in the mammalian ovary becomes very important to maximize the female reproductive potential. The present review approaches related aspects of in vitro culture systems for the development of immature oocytes originated from the preantral follicles, as well the techniques to analyze their efficiency.(AU)

Sistemas de cultivo in vitro para o desenvolvimento de oócitos imaturos de mamíferos / Systems in vitro development of immature oocytes of mammalians

O ovário mamífero possui um grande estoque de folículos primordiais, que é uma fonte potencial de oócitos para a produção in vitro de embriões. Sendo assim, o desenvolvimento de sistemas de cultivo in vitro que explorem o grande número de oócitos imaturos oriundos de folículos pré-antrais de ovários mamíferos torna-se importante para maximizar o potencial reprodutivo das fêmeas. A presente revisão aborda os aspectos relacionados aos sistemas de cultivo in vitro para desenvolvimento de oócitos imaturos oriundos de folículos pré-antrais, bem como às técnicas para analisar sua eficiência.

The mammalian ovary has a large population of primordial follicles, which is a potential source of oocytes useful for embryo in vitro production. Due to this potential, the development of in vitro culture systems to exploit the large number of immature oocytes from the preantral follicles in the mammalian ovary becomes very important to maximize the female reproductive potential. The present review approaches related aspects of in vitro culture systems for the development of immature oocytes originated from the preantral follicles, as well the techniques to analyze their efficiency.