RESUMO
The present study evaluated the effect of the addition of antioxidants anethole (AN) and robinin (RO) in the vitrification solution, and the in vitro incubation (IVI) medium of ovine ovarian tissue. Ovarian fragments were vitrified without antioxidant (VWA) or with different concentrations of AN (30, 300 and 2000⯵g/mL) or RO (0.125, 0.25 and 0.50â¯mg/mL), followed by IVI (24â¯h). Histological analyses showed that the percentage of morphologically normal preantral follicles (MNPF) in AN 2000 did not differ from RO 0.125 or fresh ovarian tissue (CTR). Subsequently, ovarian fragments were vitrified in the presence of AN 2000 and RO 0.125 followed by IVI without or with (AN 2000+ and RO 0.125+) the same antioxidants. The follicular activation in all treatments was significantly increased as compared to the CTR. The stroma cell density (SCD) in all the vitrified fragments was significantly lower than the CTR. However, in the AN 2000 and RO 0.125 this parameter was significantly higher when compared to the VWA. The reactive oxygen species (ROS) in the ovarian cortex of the AN 2000 or AN 2000+ were significantly reduced in comparison with the CTR while the intracellular ROS levels of AN 2000 and CTR were similar. The total antioxidant capacity (TAC) in RO 0.125 was significantly higher than that of VWA, AN 2000 and AN 2000+. According to the results, the use of antioxidants (AN or RO) only in the vitrification solution of ovine ovarian tissue is recommended, due to their better preservation of the SCD. Moreover, AN 2000 best maintains the follicular morphology, while RO 0.125 has a high TAC.
Assuntos
Antioxidantes/metabolismo , Criopreservação/veterinária , Ovário/efeitos dos fármacos , Ovinos , Preservação de Tecido/veterinária , Animais , Criopreservação/métodos , Meios de Cultura , Feminino , Espécies Reativas de Oxigênio/metabolismo , VitrificaçãoRESUMO
This study investigated the levels of messenger ribonucleic acids (mRNA) for inhibin-ßA subunit in goat primordial, primary and secondary follicles, as well as in cumulus-oocyte complexes (COCs) and mural granulosa / theca cells of antral follicles. The effects of activin-A (100ng mL-1) and/or follicle stimulating hormone (FSH, 50ng mL-1) on growth and expression of mRNA for activin-A and FSH receptor (FSH-R) in secondary follicles cultured for six days were evaluated. The data showed that the expression of inhibin-ßA is lower in secondary follicles than in primary follicles and is higher in large antral follicles than in small antral follicles. After culture, activin-A and/or FSH promoted growth of secondary follicles, while FSH increased the levels of mRNA for inhibin-ßA, and activin-A increased the levels of FSH-R mRNA. In conclusion, mRNA for inhibin-ßA is expressed at different levels in pre-antral and antral follicles and activin-A acts as a stimulator of the FSH-R expression in goat follicles. On its turn, the expression of inhibin-ßA is stimulated by FSH, which together with activin-A promotes secondary follicle growth in-vitro.
Este estudo investigou os níveis de ácidos ribonucleicos (RNAm) para a subunidade ßA da inibina em folículos primordiais, primários e secundários caprinos, bem como em complexos cumulus-oócitos (CCOs) e células da granulosa mural/teca de folículos antrais. Além disso, avaliaram-se os efeitos da ativina-A (100ng mL-1) e/ou hormônio folículo estimulante (FSH, 50ng mL-1) sobre o crescimento e a expressão do RNAm para inibina-ßA e receptores de FSH (FSH-R) em folículos secundários cultivados por seis dias. Os dados mostraram que a expressão de inibina-ßA é menor em folículos secundários do que em folículos primários e é maior em grandes folículos antrais que nos pequenos folículos antrais. Após o cultivo, ativina-A e/ou FSH promoveram o crescimento de folículos secundários. Enquanto o FSH aumentou os níveis de RNAm para inibina-ßA, a ativina-A aumentou os níveis de RNAm para FSH-R. Em conclusão, a inibina-ßA é expressa em diferentes níveis em folículos pré-antrais e antrais e a ativina-A atua como um estimulador da expressão de FSH-R em folículos caprinos. Por sua vez, a expressão de inibina-ßA é estimulada pelo FSH, que, juntamente com ativina, promove o crescimento de folículos secundários in vitro.
RESUMO
This study investigated the levels of messenger ribonucleic acids (mRNA) for inhibin-ßA subunit in goat primordial, primary and secondary follicles, as well as in cumulus-oocyte complexes (COCs) and mural granulosa / theca cells of antral follicles. The effects of activin-A (100ng mL-1) and/or follicle stimulating hormone (FSH, 50ng mL-1) on growth and expression of mRNA for activin-A and FSH receptor (FSH-R) in secondary follicles cultured for six days were evaluated. The data showed that the expression of inhibin-ßA is lower in secondary follicles than in primary follicles and is higher in large antral follicles than in small antral follicles. After culture, activin-A and/or FSH promoted growth of secondary follicles, while FSH increased the levels of mRNA for inhibin-ßA, and activin-A increased the levels of FSH-R mRNA. In conclusion, mRNA for inhibin-ßA is expressed at different levels in pre-antral and antral follicles and activin-A acts as a stimulator of the FSH-R expression in goat follicles. On its turn, the expression of inhibin-ßA is stimulated by FSH, which together with activin-A promotes secondary follicle growth in-vitro.
Este estudo investigou os níveis de ácidos ribonucleicos (RNAm) para a subunidade ßA da inibina em folículos primordiais, primários e secundários caprinos, bem como em complexos cumulus-oócitos (CCOs) e células da granulosa mural/teca de folículos antrais. Além disso, avaliaram-se os efeitos da ativina-A (100ng mL-1) e/ou hormônio folículo estimulante (FSH, 50ng mL-1) sobre o crescimento e a expressão do RNAm para inibina-ßA e receptores de FSH (FSH-R) em folículos secundários cultivados por seis dias. Os dados mostraram que a expressão de inibina-ßA é menor em folículos secundários do que em folículos primários e é maior em grandes folículos antrais que nos pequenos folículos antrais. Após o cultivo, ativina-A e/ou FSH promoveram o crescimento de folículos secundários. Enquanto o FSH aumentou os níveis de RNAm para inibina-ßA, a ativina-A aumentou os níveis de RNAm para FSH-R. Em conclusão, a inibina-ßA é expressa em diferentes níveis em folículos pré-antrais e antrais e a ativina-A atua como um estimulador da expressão de FSH-R em folículos caprinos. Por sua vez, a expressão de inibina-ßA é estimulada pelo FSH, que, juntamente com ativina, promove o crescimento de folículos secundários in vitro.
RESUMO
This study investigated the levels of messenger ribonucleic acids (mRNA) for inhibin-ßA subunit in goat primordial, primary and secondary follicles, as well as in cumulus-oocyte complexes (COCs) and mural granulosa / theca cells of antral follicles. The effects of activin-A (100ng mL-1) and/or follicle stimulating hormone (FSH, 50ng mL-1) on growth and expression of mRNA for activin-A and FSH receptor (FSH-R) in secondary follicles cultured for six days were evaluated. The data showed that the expression of inhibin-ßA is lower in secondary follicles than in primary follicles and is higher in large antral follicles than in small antral follicles. After culture, activin-A and/or FSH promoted growth of secondary follicles, while FSH increased the levels of mRNA for inhibin-ßA, and activin-A increased the levels of FSH-R mRNA. In conclusion, mRNA for inhibin-ßA is expressed at different levels in pre-antral and antral follicles and activin-A acts as a stimulator of the FSH-R expression in goat follicles. On its turn, the expression of inhibin-ßA is stimulated by FSH, which together with activin-A promotes secondary follicle growth in-vitro.
Este estudo investigou os níveis de ácidos ribonucleicos (RNAm) para a subunidade ßA da inibina em folículos primordiais, primários e secundários caprinos, bem como em complexos cumulus-oócitos (CCOs) e células da granulosa mural/teca de folículos antrais. Além disso, avaliaram-se os efeitos da ativina-A (100ng mL-1) e/ou hormônio folículo estimulante (FSH, 50ng mL-1) sobre o crescimento e a expressão do RNAm para inibina-ßA e receptores de FSH (FSH-R) em folículos secundários cultivados por seis dias. Os dados mostraram que a expressão de inibina-ßA é menor em folículos secundários do que em folículos primários e é maior em grandes folículos antrais que nos pequenos folículos antrais. Após o cultivo, ativina-A e/ou FSH promoveram o crescimento de folículos secundários. Enquanto o FSH aumentou os níveis de RNAm para inibina-ßA, a ativina-A aumentou os níveis de RNAm para FSH-R. Em conclusão, a inibina-ßA é expressa em diferentes níveis em folículos pré-antrais e antrais e a ativina-A atua como um estimulador da expressão de FSH-R em folículos caprinos. Por sua vez, a expressão de inibina-ßA é estimulada pelo FSH, que, juntamente com ativina, promove o crescimento de folículos secundários in vitro.
RESUMO
Background: The anti-müllerian hormone (AMH) is a member of the transforming growth factor (TGF-) superfamily, which exerts important functions on local regulation of folliculogenesis. Although in vivo and in vitro studies suggest that AMH affects the primordial follicle assembly and activation, as well as the responsiveness of growing follicles to folliclestimulating hormone (FSH), the physiological mechanisms involved in these actions remain to be fully elucidated. Given the relevance of AMH in the folliculogenesis, this review aimed to describe the structural features, expression and the main biological effects of AMH on the follicular development. Review: Originally identified as a testicular product, AMH is responsible for regression of the Müllerian ducts during sexual differentiation of male embryos. In females, AMH is produced almost exclusively by granulosa cells of ovarian growing follicles, whose serum levels are positively related to the number of ovarian follicles, making AMH an excellent clinic marker of ovarian reserve. Along this work, it was shown aspects related to the structural characterization of AMH and its specific type II receptor (AMHRII). AMH is a glycoprotein dimer linked by disulfide bonds. The mature protein comprises two unequal domains: a long N-terminal domain (110-kDa) and a short C-terminal domain (25-kDa), responsible for the biological act
A foliculogênese é um evento complexo que envolve o recrutamento do pool de folículos primordiais, seguido por uma fase de crescimento e diferenciação. É sabido que para a sua regulação são requeridos vários fatores parácrinos e autócrinos, especialmente fatores de crescimento produzidos pelas células da granulosa. Dentre esses fatores, destaca-se o hormônio anti-mülleriano (AMH), uma glicoproteína membro da superfamília de fatores de crescimento transformantes (TGF-). Inicialmente o AMH foi estudado por seu papel regulatório no processo de diferenciação sexual masculina. [...]
RESUMO
Background: The anti-müllerian hormone (AMH) is a member of the transforming growth factor (TGF-) superfamily, which exerts important functions on local regulation of folliculogenesis. Although in vivo and in vitro studies suggest that AMH affects the primordial follicle assembly and activation, as well as the responsiveness of growing follicles to folliclestimulating hormone (FSH), the physiological mechanisms involved in these actions remain to be fully elucidated. Given the relevance of AMH in the folliculogenesis, this review aimed to describe the structural features, expression and the main biological effects of AMH on the follicular development. Review: Originally identified as a testicular product, AMH is responsible for regression of the Müllerian ducts during sexual differentiation of male embryos. In females, AMH is produced almost exclusively by granulosa cells of ovarian growing follicles, whose serum levels are positively related to the number of ovarian follicles, making AMH an excellent clinic marker of ovarian reserve. Along this work, it was shown aspects related to the structural characterization of AMH and its specific type II receptor (AMHRII). AMH is a glycoprotein dimer linked by disulfide bonds. The mature protein comprises two unequal domains: a long N-terminal domain (110-kDa) and a short C-terminal domain (25-kDa), responsible for the biological act
A foliculogênese é um evento complexo que envolve o recrutamento do pool de folículos primordiais, seguido por uma fase de crescimento e diferenciação. É sabido que para a sua regulação são requeridos vários fatores parácrinos e autócrinos, especialmente fatores de crescimento produzidos pelas células da granulosa. Dentre esses fatores, destaca-se o hormônio anti-mülleriano (AMH), uma glicoproteína membro da superfamília de fatores de crescimento transformantes (TGF-). Inicialmente o AMH foi estudado por seu papel regulatório no processo de diferenciação sexual masculina. [...]
RESUMO
Background: The development of animal reproductive biotechniques is intended of raising reproduction effi ciency. The mammal ovary contains thousands of follicles, of which approximately 99.9% are eliminated by means of the atresia, apoptosis and cellular necrosis process. In other to reduce this follicular loss, the methods for isolation and characterization of preantral follicles have been studied as a premise to culture systems of these structures. The purpose of this study was quantify and evaluate the quality of preantral ovarian follicles from prepubertal gilts after mechanical isolation procedure. Furthermore, it aims to analyze the preantral follicular histological morphology.Materials, Methods & Results: Ovaries (n = 20) from prepubertal gilts were divided in two halves and used each one for isolation and histological processes. The tissue chopper previously regulated for the performance of seriated cuts at 200 m intervals was used for mechanical isolation. The marker bisbenzimidine Hoechst 33342 was added to the follicular pool for evidence the presence of granulosa cells around oocyte and the viability of the isolated PAF was evaluated by using propidium iodide. For the histological evaluation, the ovarian halves were fi xed, dehydrated and diaphanized, and after enclosed in paraffi n, each of them was divided into 5 blocks and sectioned in series 6 m thick. At
Background: The development of animal reproductive biotechniques is intended of raising reproduction effi ciency. The mammal ovary contains thousands of follicles, of which approximately 99.9% are eliminated by means of the atresia, apoptosis and cellular necrosis process. In other to reduce this follicular loss, the methods for isolation and characterization of preantral follicles have been studied as a premise to culture systems of these structures. The purpose of this study was quantify and evaluate the quality of preantral ovarian follicles from prepubertal gilts after mechanical isolation procedure. Furthermore, it aims to analyze the preantral follicular histological morphology.Materials, Methods & Results: Ovaries (n = 20) from prepubertal gilts were divided in two halves and used each one for isolation and histological processes. The tissue chopper previously regulated for the performance of seriated cuts at 200 m intervals was used for mechanical isolation. The marker bisbenzimidine Hoechst 33342 was added to the follicular pool for evidence the presence of granulosa cells around oocyte and the viability of the isolated PAF was evaluated by using propidium iodide. For the histological evaluation, the ovarian halves were fi xed, dehydrated and diaphanized, and after enclosed in paraffi n, each of them was divided into 5 blocks and sectioned in series 6 m thick. At
RESUMO
Background: The development of animal reproductive biotechniques is intended of raising reproduction effi ciency. The mammal ovary contains thousands of follicles, of which approximately 99.9% are eliminated by means of the atresia, apoptosis and cellular necrosis process. In other to reduce this follicular loss, the methods for isolation and characterization of preantral follicles have been studied as a premise to culture systems of these structures. The purpose of this study was quantify and evaluate the quality of preantral ovarian follicles from prepubertal gilts after mechanical isolation procedure. Furthermore, it aims to analyze the preantral follicular histological morphology.Materials, Methods & Results: Ovaries (n = 20) from prepubertal gilts were divided in two halves and used each one for isolation and histological processes. The tissue chopper previously regulated for the performance of seriated cuts at 200 m intervals was used for mechanical isolation. The marker bisbenzimidine Hoechst 33342 was added to the follicular pool for evidence the presence of granulosa cells around oocyte and the viability of the isolated PAF was evaluated by using propidium iodide. For the histological evaluation, the ovarian halves were fi xed, dehydrated and diaphanized, and after enclosed in paraffi n, each of them was divided into 5 blocks and sectioned in series 6 m thick. At
Background: The development of animal reproductive biotechniques is intended of raising reproduction effi ciency. The mammal ovary contains thousands of follicles, of which approximately 99.9% are eliminated by means of the atresia, apoptosis and cellular necrosis process. In other to reduce this follicular loss, the methods for isolation and characterization of preantral follicles have been studied as a premise to culture systems of these structures. The purpose of this study was quantify and evaluate the quality of preantral ovarian follicles from prepubertal gilts after mechanical isolation procedure. Furthermore, it aims to analyze the preantral follicular histological morphology.Materials, Methods & Results: Ovaries (n = 20) from prepubertal gilts were divided in two halves and used each one for isolation and histological processes. The tissue chopper previously regulated for the performance of seriated cuts at 200 m intervals was used for mechanical isolation. The marker bisbenzimidine Hoechst 33342 was added to the follicular pool for evidence the presence of granulosa cells around oocyte and the viability of the isolated PAF was evaluated by using propidium iodide. For the histological evaluation, the ovarian halves were fi xed, dehydrated and diaphanized, and after enclosed in paraffi n, each of them was divided into 5 blocks and sectioned in series 6 m thick. At
RESUMO
The leukemia inhibitory factor (LIF) is a glycoprotein from the interleukin-6 family, which acts with its membrane receptors in different biological processes. Studies have reported the involvement of this factor during ovarian follicular development and embryo implantation in mammals. This review summarizes the main aspects of the molecular structure of LIF, its receptors, mechanisms of action, expression and functions focusing on ovarian physiology and embryo implantation. Keywords: LIF, ovarian, follicle, endometrium, mammalian.
O fator inibidor de leucemia (LIF) é uma glicoproteína pertencente à família das interleucinas-6 que ligados aos seus receptores intermembranários atua em diferentes processos biológicos. Estudos têm relatado a participação deste fator durante o desenvolvimento folicular ovariano e implantação em mamíferos. Esta revisão resume os principais aspectos da estrutura molecular do LIF, seus receptores, mecanismos de ação, expressão e funções focando na fisiologia ovariana e implantação embrionária. Palavras-Chave: LIF, ovário, folículo, endométrio, mamífero.