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1.
Rev. bras. reprod. anim ; 47(2): 76-80, abr.-jun. 2023.
Artigo em Inglês | VETINDEX | ID: biblio-1434952

Resumo

The follicular growth waves are directly linked to the fluctuations in plasma gonadotrophins, which are controlled by the hypothalamic GnRH release pattern. Therefore, if the actions of the GnRH are inhibited or blocked, the final stages of the antral follicle growth are suppressed, resulting in an induced anestrus (a.k.a. waveless model). In the human medicine, GnRH agonists or antagonists are broadly used in the control of ovarian disfunctions, as well as in the preparation of women for assisted reproductive cycles. In cattle, a similar effect can be obtained by active immunization against GnRH. This was shown to be a viable strategy, for example, for the control of chronic cases of cystic ovarian disease in oocyte donors. However, on shall take into account the substantial individual variation on the immune response and, consequently, the lack of control of the duration of the anestrus induced. The waveless model is also very useful as a research model, once it controls the potential interference of the endogenous FSH and LH, improving the sensitivity of essays with exogenous hormones and consequently reducing the required number of replicas within studies.(AU)


O padrão de crescimento folicular em ondas está diretamente associado às flutuações nas concentrações plasmáticas de gonadotrofinas, controladas por sua vez pelo padrão de liberação de GnRH hipotalâmico. Desta forma, a inibição ou bloqueio da ação do GnRH suprime as etapas finais do crescimento folicular, resultando em anestro induzido (também chamado modelo waveless). Na medicina humana, agonistas ou antagonistas de GnRH são utilizados tanto no controle de disfunções ovarianas quanto na preparação de pacientes para procedimentos de reprodução assistida. Em bovinos, este efeito pode ser obtido pela imunização ativa contra GnRH, e mostrou-se estratégia viável, por exemplo, no controle de casos crônicos de doença ovariana cística em doadoras de oócitos. Contudo, é importante considerar a grande variação individual na resposta à imunização e consequente impossibilidade de controlar a duração do anestro induzido. O modelo waveless também é de grande utilidade na pesquisa, uma vez que elimina a potencial interferência do FSH e LH endógenos, aumentando a sensibilidade nos ensaios com hormônios exógenos e consequentemente reduzindo o número de réplicas necessárias nos estudos.(AU)


Assuntos
Animais , Bovinos/embriologia , Estruturas Embrionárias/crescimento & desenvolvimento , Fase Folicular , Gonadotropinas/análise
2.
Anim. Reprod. (Online) ; 20(2): e20230061, 2023.
Artigo em Inglês | VETINDEX | ID: biblio-1452314

Resumo

Animals that exhibited estrus had greater pregnancy success compared to animals that did not exhibit estrus before fixed-time AI (FTAI). Estradiol is synthesized in bovine ovarian follicles under gonadotropin regulation and can directly and indirectly regulate the uterine receptivity and luteal function. Estradiol concentrations at FTAI impacted oviductal gene expression and has been reported to play an important role in establishing the timing of uterine receptivity. These changes have been reported to impact uterine pH and sperm transport to the site of fertilization. After fertilization, preovulatory estradiol has been reported to improve embryo survival likely by mediating changes in uterine blood flow, endometrial thickness and changes in histotroph. Cows with greater estradiol concentrations at the time of GnRH-induced ovulation also had a larger dominant follicle size and greater circulating progesterone concentrations on day 7. Therefore, it is impossible to accurately determine the individual benefit of greater estradiol concentrations prior to ovulation and greater progesterone concentrations following ovulation to pregnancy establishment, as these two measurements are confounded. Research has indicated an importance in the occurrence and timing of increasing preovulatory concentrations of estradiol, but increasing estradiol concentrations by supplementation may not be sufficient to increase fertility. Increased production of estradiol by the preovulatory follicle may be required to enhance fertility through the regulation of sperm transport, fertilization, oviductal secretions, the uterine environment, and embryo survival.(AU)


Assuntos
Animais , Feminino , Gravidez , Bovinos/embriologia , Estradiol/efeitos adversos , Folículo Ovariano/química , Útero/química , Corpo Lúteo/química , Fase Folicular
3.
Rev. bras. reprod. anim ; 46(4): 364-368, out.-dez. 2022.
Artigo em Português | VETINDEX | ID: biblio-1415066

Resumo

Algunas especies de animales como equinos, pequeños rumiantes, algunos roedores y caninos y felinos silvestres presentan un período de reposo sexual estacional de duración e intensidad variable. Esta estacionalidad está directamente relacionada con las horas luz diarias (fotoperiodo) a las que se hallan sometidos los animales y se evidencia en las localizaciones geográficas en las que existen marcadas variaciones en la duración del día durante el año. En el felino doméstico (Felis silvestris catus) la estacionalidad ovulatoria y estral de la hembra se halla bien documentada y ocurre durante los días que presentan más de 12 horas de luz (primavera-verano). Sin embargo, la estacionalidad reproductiva del gato ha sido definida más tardíamente. Es así que se ha observado que, si bien los gatos producen espermatozoides todo el año, la producción espermática es mayor en la estación reproductiva acompañando la estacionalidad de la hembra. Estos hallazgos han permitido utilizar estos conocimientos para el control de la reproducción y aplicación de biotecnología en felinos domésticos. Asimismo, los conocimientos obtenidos pueden ser aplicados a felinos silvestres tomando al gato doméstico como modelo.(AU)


Algumas espécies de animais como equinos, pequenos ruminantes, alguns roedores e os caninos e felinos selvagens apresentam um período de repouso sexual sazonal de duração e intensidade variáveis. Essa sazonalidade está diretamente relacionada às horas diárias de luz (fotoperíodo) a que os animais são submetidos e é evidenciada em localizações geográficas onde ocorrem variações marcantes na duração do dia durante o ano. No felino doméstico (Felis silvestris catus) a sazonalidade ovulatória e estral da fêmea é bem documentada e ocorre em dias com mais de 12 horas de luz (primavera-verão). No entanto, a sazonalidade reprodutiva do gato foi definida posteriormente, quando foi observado que, embora os gatos produzam espermatozoides ao longo do ano, a produção de espermatozoides é maior na estação reprodutiva, acompanhando a sazonalidade da fêmea. A utilização desses conhecimentos possibilitaram o controle da reprodução e aplicação da biotecnologia em felinos domésticos. Da mesma forma, tomando o gato doméstico como modelo, os conhecimentos obtidos podem ser aplicados aos gatos selvagens.(AU)


Assuntos
Animais , Feminino , Gatos/fisiologia , Ciclo Estral/fisiologia , Fenômenos Reprodutivos Fisiológicos , Fase Folicular/fisiologia , Estações do Ano , Fotoperíodo
4.
Acta sci. vet. (Impr.) ; 50: Pub. 1876, 2022. ilus, tab
Artigo em Português | VETINDEX | ID: biblio-1400770

Resumo

Background: Fixed Time Artificial Insemination (FTAI) has achieved a significant evolution in the last 18 years, however, despite the progress achieved by modern FTAI programs, the conception rates obtained are still low. Therefore, this study aimed to evaluate the interrelation between progesterone levels in the periovulatory period and reproductive parameters of Nellore cows submitted to an FTAI protocol. Materials, Methods & Results: On a random day, called day 0 (D0), 57 cows received a P4 device associated with the intramuscular (IM) application of 2.0 mg of estradiol benzoate. On D9, the P4 devices were removed and then were administered 500 µg of cloprostenol sodium IM; 0.6 mg of estradiol cypionate IM and 300 IUI of Equine Chorionic Gonadotrophin IM. Blood samples were collected for the determination of serum P4 concentrations on D9 and D11 of the protocol. The evaluations of follicular diameter (DFOL), follicular wall area (AFOL) and the vascularization area of the follicle wall (VFOL) were carried out on D11 using B-mode ultrasonography examination and colour Doppler, and then the artificial inseminations were performed. The evaluation of the corpus luteum diameter (CLD), of the total corpus luteum area (CLA), of the area of corpus luteum vascularization (CLV) and blood sampling for determination of postovulatory P4 levels (Post-P4) were performed on D24. For the analysis of the P4 concentration the chemiluminescence method was used, with a sensitivity of 0.1 ng/mL. According to the P4 concentrations on D11, cows were divided into 2 groups, LOW LEVELS OF P4 and HIGH LEVELS OF P4. The diagnosis of pregnancy was performed using transrectal ultrasonography on D45, at this point the cows were divided into 2 groups, PREGNANT and NON-PREGNANT. The correlation between DFOL and P4 dosage on D11 was moderate, negative and significant and between the AFOL and the serum P4 levels on D9, was moderate, negative and significant. As for the other correlations between follicular and luteal parameters and serum P4 levels, these were low to moderate, negative and not significant. Cows in the LOW LEVELS OF P4 group had significantly larger diameter and follicular areas than the cows in the HIGH LEVELS OF P4 group, the other follicular and luteal parameters showed no statistical difference. Of the total 57 cows that were inseminated, 30 cows became pregnant. Cows in the PREGNANT group had serum P4 levels on D9 equivalent to that obtained by the NON-PREGNANT group. However, at D11 the cows that became pregnant presented significantly lower serum P4 levels than cows that did not become pregnant. Discussion: The results of the interrelation between follicular parameters and P4 levels obtained in the present study, pointed out that the lower the levels of P4, the higher the follicular parameters, corroborating with other authors. Thus, larger preovulatory follicles provided high ovulation rates. Periovulatory serum P4 levels did not significantly affect the morphofunctional parameters of the CL. Such findings may be justified by high periovulatory P4 levels resulting from less efficient luteolysis, exert a negative effect on the results of FTAI protocols, because progesterone inhibits the release of LH pulses. It is concluded that lower periovulatory P4 levels established a favourable condition for follicular development and fertility, however, morphofunctional parameters of the corpus luteum were not affected.


Assuntos
Animais , Feminino , Gravidez , Bovinos , Progesterona/análise , Monitorização Uterina/veterinária , Fase Folicular/fisiologia , Folículo Ovariano/crescimento & desenvolvimento , Inseminação Artificial/veterinária , Hemodinâmica
5.
Anim. Reprod. (Online) ; 18(3): e20210016, 2021. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461551

Resumo

We aimed to compare the effect of three estradiol benzoate (EB) doses on follicular wave emergence (FWE) and dominant follicle growth of suckled Nelore cows submitted to TAI (D0). On a random day of estrous cycle (D−10), multiparous (MULT; n=36) and primiparous (PRIM; n=20) suckled Nelore cows received an intravaginal progesterone (P4) device and were assigned in three groups. Cows in the EB-1 (n=20), EB-1.5 (n=15) or EB-2 (n=21) groups received, respectively, an im treatment with 1, 1.5 or 2 mg EB. A subgroup (n=10-13 cows/group) were subject to daily ovarian evaluations from D−10 to D0. On D−2, P4 devices were removed, and all cows received the same treatment: 1 mg estradiol cypionate, 0.53 mg sodium cloprostenol, and 300 IU eCG. Statistical analyses were performed considering only the main effects of treatment group and parity order. The proportion of cows with a synchronized FWE and the moment of the FWE did not differ (p>0.05) among the treatment groups (overall: 80% [28/35] and 4.1 ± 0.4 days); however, the FWE occurred earlier (p=0.007) in MULT (3.8 ± 0.2 days) than PRIM (5.1 ± 0.4) cows. The proportion of animals detected in estrus was greater (86% [31/36] vs. 70% [14/20]; p=0.02) and the dominant follicle was larger on D−2 (9.7 ± 0.3 mm vs. 7.8 ± 0.7 mm; p=0.006) and D0 (11.9 ± 0.4 mm vs. 10 ± 0.5 mm; p=0.008) in MULT than PRIM cows. In conclusion, the three EB doses presented similar efficiency to synchronize the FWE in suckled Nelore cows. Moreover, a delayed FWE and smaller dominant follicle is observed in PRIM cows, contributing to the reduced reproductive performance in this parity category when using similar TAI protocols of MULT cows.


Assuntos
Feminino , Animais , Bovinos , Benzoatos/administração & dosagem , Bovinos/embriologia , Estradiol/análogos & derivados , Fase Folicular
6.
Anim. Reprod. ; 18(3): e20210016, 2021. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-31934

Resumo

We aimed to compare the effect of three estradiol benzoate (EB) doses on follicular wave emergence (FWE) and dominant follicle growth of suckled Nelore cows submitted to TAI (D0). On a random day of estrous cycle (D−10), multiparous (MULT; n=36) and primiparous (PRIM; n=20) suckled Nelore cows received an intravaginal progesterone (P4) device and were assigned in three groups. Cows in the EB-1 (n=20), EB-1.5 (n=15) or EB-2 (n=21) groups received, respectively, an im treatment with 1, 1.5 or 2 mg EB. A subgroup (n=10-13 cows/group) were subject to daily ovarian evaluations from D−10 to D0. On D−2, P4 devices were removed, and all cows received the same treatment: 1 mg estradiol cypionate, 0.53 mg sodium cloprostenol, and 300 IU eCG. Statistical analyses were performed considering only the main effects of treatment group and parity order. The proportion of cows with a synchronized FWE and the moment of the FWE did not differ (p>0.05) among the treatment groups (overall: 80% [28/35] and 4.1 ± 0.4 days); however, the FWE occurred earlier (p=0.007) in MULT (3.8 ± 0.2 days) than PRIM (5.1 ± 0.4) cows. The proportion of animals detected in estrus was greater (86% [31/36] vs. 70% [14/20]; p=0.02) and the dominant follicle was larger on D−2 (9.7 ± 0.3 mm vs. 7.8 ± 0.7 mm; p=0.006) and D0 (11.9 ± 0.4 mm vs. 10 ± 0.5 mm; p=0.008) in MULT than PRIM cows. In conclusion, the three EB doses presented similar efficiency to synchronize the FWE in suckled Nelore cows. Moreover, a delayed FWE and smaller dominant follicle is observed in PRIM cows, contributing to the reduced reproductive performance in this parity category when using similar TAI protocols of MULT cows.(AU)


Assuntos
Animais , Feminino , Bovinos , Bovinos/embriologia , Benzoatos/administração & dosagem , Estradiol/análogos & derivados , Fase Folicular
7.
Anim. Reprod. (Online) ; 17(2): e20190115, 2020. ilus, tab
Artigo em Inglês | VETINDEX | ID: biblio-1461504

Resumo

Domestic and wild goats are very susceptible animals to predation, specially when pregnancy occurs. This study aimed to evaluate the use of goat fetal ovarian tissue for vitrification followed by xenotransplantation and fresh xenotransplantation in two immunosuppressed mice models (C57BL/6 SCID and Balb-C NUDE). Goat fetus ovaries were collected in slaughterhouses, divided into small cortical pieces and were destined for fresh xenotransplantation (FX) and cryopreservation followed by xenotransplantation (CX). Five recipients from each lineage were used for FX and 10 animals from each lineage for CX. The mice were euthanized after 65 postoperative days, and the transplants were collected for microscopic assessment. The blood plasma was collected for estradiol measurement. Independently of mice strain, all recipients presented complete estrus cycle in FX and 80% after CX groups. Follicles were observed at all development stages without morphological changes. The volume density and total vessel surface observed in the transplants were different (p <0.01) between groups. The estradiol levels in the recipients did not differ (p <0.05) among the treatments. Thus, it is possible to activate the preantral follicles in the ovaries of fetuses by optimizing germplasm utilization and conservation of domestic and endangered wild goats that are in predatory situations, undesirable drowning or accidental death, since provided conditions for xenotransplantation are performed.


Assuntos
Masculino , Feminino , Animais , Camundongos , Criopreservação , Fase Folicular , Ruminantes/anatomia & histologia , Ruminantes/fisiologia , Vitrificação , Transplante Heterólogo
8.
Anim. Reprod. ; 17(2): e20190115, 2020. ilus, tab
Artigo em Inglês | VETINDEX | ID: vti-29308

Resumo

Domestic and wild goats are very susceptible animals to predation, specially when pregnancy occurs. This study aimed to evaluate the use of goat fetal ovarian tissue for vitrification followed by xenotransplantation and fresh xenotransplantation in two immunosuppressed mice models (C57BL/6 SCID and Balb-C NUDE). Goat fetus ovaries were collected in slaughterhouses, divided into small cortical pieces and were destined for fresh xenotransplantation (FX) and cryopreservation followed by xenotransplantation (CX). Five recipients from each lineage were used for FX and 10 animals from each lineage for CX. The mice were euthanized after 65 postoperative days, and the transplants were collected for microscopic assessment. The blood plasma was collected for estradiol measurement. Independently of mice strain, all recipients presented complete estrus cycle in FX and 80% after CX groups. Follicles were observed at all development stages without morphological changes. The volume density and total vessel surface observed in the transplants were different (p <0.01) between groups. The estradiol levels in the recipients did not differ (p <0.05) among the treatments. Thus, it is possible to activate the preantral follicles in the ovaries of fetuses by optimizing germplasm utilization and conservation of domestic and endangered wild goats that are in predatory situations, undesirable drowning or accidental death, since provided conditions for xenotransplantation are performed.(AU)


Assuntos
Animais , Masculino , Feminino , Camundongos , Ruminantes/anatomia & histologia , Ruminantes/fisiologia , Vitrificação , Fase Folicular , Criopreservação , Transplante Heterólogo
9.
Anim. Reprod. (Online) ; 16(3): 440-448, 2019.
Artigo em Inglês | VETINDEX | ID: biblio-1461453

Resumo

Much of what we know about the involvement of the immune system in periovulatory follicle differentiation, ovulation and subsequent formation of the corpus luteum in cattle is drawn from the findings of studies in several mammalian livestock species. By integrating published histological data from cattle, sheep and pigs and referring back to the more comprehensive knowledge bank that exists for mouse and humans we can sketch out the key cells of the immune system and the cytokines and growth factors that they produce that are involved in follicle differentiation and luteinization, ovulation and early follicle development. These contributions are reviewed and the key findings, discussed.


Assuntos
Feminino , Animais , Bovinos , Fase Folicular/imunologia , Ovulação/imunologia , Sistema Imunitário/imunologia , Corpo Lúteo
10.
Anim. Reprod. ; 16(3): 440-448, 2019.
Artigo em Inglês | VETINDEX | ID: vti-22351

Resumo

Much of what we know about the involvement of the immune system in periovulatory follicle differentiation, ovulation and subsequent formation of the corpus luteum in cattle is drawn from the findings of studies in several mammalian livestock species. By integrating published histological data from cattle, sheep and pigs and referring back to the more comprehensive knowledge bank that exists for mouse and humans we can sketch out the key cells of the immune system and the cytokines and growth factors that they produce that are involved in follicle differentiation and luteinization, ovulation and early follicle development. These contributions are reviewed and the key findings, discussed.(AU)


Assuntos
Animais , Feminino , Bovinos , Fase Folicular/imunologia , Ovulação/imunologia , Sistema Imunitário/imunologia , Corpo Lúteo
11.
R. bras. Ci. Vet. ; 26(3): 111-115, jul.-set. 2019. ilus, tab
Artigo em Português | VETINDEX | ID: vti-24704

Resumo

O objetivo deste estudo foi obter o perfil eletroforético das proteínas séricas em éguas cíclicas e verificar as diferenças entre as fases folicular e luteal do ciclo estral nesta espécie. Foram utilizadas 18 éguas, totalizando 36 amostras de soro, sendo duas de cada égua. As amostras foram colhidas no estro e no diestro. As proteínas séricas totais foram obtidas pelo método do Biureto, a partir da utilização de Kits comerciais (LABTEST®) e, as diferentes subfrações proteicas, por eletroforese em gel de poliacrilamida (SDS-PAGE). O eletroforetograma das proteínas séricas colocou em evidência a presença de 17 a 25 frações proteicas, cujos pesos moleculares variaram de 22 a 254 kDa. Identificaram-se duas proteínas ainda não nomeadas oficialmente, de massas moleculares (MM) 23 kDa e 144 kDa. Os valores médios ± SEM obtidos para cada variável no estro e no diestro, respectivamente, foram: proteínas totais (g/dL) 7,11 ± 0,07 e 7,36 ± 0,07; albumina (mg/dL) 4790,83 ± 69,10 e 5027,19 ± 69,10; α1 glicoproteína ácida (mg/dL) 4,90 ± 0,31 e 4,93 ± 0,31; ceruloplasmina (mg/dL) 15,28 ± 1,31 e 10,65 ± 1,31; haptoglobina (mg/dL) 22,70 ± 1,16 e 27,06 ± 1,16; transferrina (mg/dL) 329,00 ± 9,78 e 350,16 ± 9,78; IgA (mg/dL) 119,91 ± 6,30 e 107,03 ± 6,30; IgG (mg/dL) 1525,07 ± 40,18 e 1517,25 ± 40,18; MM 23 (mg/dL) 204,44 ± 8,61 e 219,79 ± 8,61; MM 144 (mg/dL) 22,13 ± 0,55 e 21,49 ± 0,55. Não houve diferença significativa das proteínas totais e suas frações do estro para o diestro. Conclui-se que as modificações hormonais durante as fases do ciclo estral da égua não interferem no proteinograma sérico.(AU)


This study aimed to obtain the electrophoretic profile of serum proteins in cyclic mares and to verify the differences between the follicular and luteal phases of the estrous cycle in this species. Eighteen mares were used, totaling 36 serum samples, two of each mare. Samples were collected both in estrus and in diestrus. Total serum proteins were obtained by the Biureto method, by using commercial kits (LABTEST®), while the different protein subfractions by polyacrylamide gel electrophoresis (SDS-PAGE). The electroforetogram of serum proteins evidenced the presence of 17 to 25 protein fractions, whose molecular weights ranged from 22 to 254 kDa. Two proteins that were not yet officially named were identified, of molecular weights (MW) of 23 kDa and 144 kDa. The mean values (± SEM) obtained for each variable in estrus and diestrus were, respectively: total proteins (g/dL) 7.11 ± 0.07 and 7.36 ± 0.07; albumin (mg/dL) 4790.83 ± 69.10 and 5027.19 ± 69.10; α1 acid glycoprotein (mg/dL) 4.90 ± 0.31 and 4.93 ± 0.31; ceruloplasmine (mg/dL) 15.28 ± 1.31 and 10.65 ± 1.31; haptoglobine (mg/dL) 22.70 ± 1.16 and 27.06 ± 1.16; transferrin (mg/ dL) 329.00 ± 9.78 and 350.16 ± 9.78; IgA (mg/dL) 119.91 ± 6.30 and 107.03 ± 6.30; IgG (mg/dL) 1525.07 ± 40.18 and 1517.25 ± 40.18; MW 23 (mg/dL) 204.44 ± 8.61 and 219.79 ± 8.61; MW 144 (mg/dL) 22.13 ± 0.55 and 21.49 ± 0.55. No significant difference was verified in total proteins and its fractions in estrus and diestrus. The hormonal changes during the specific stages of the estrous cycle of the mare do not interfere with the serum proteinogram.(AU)


Assuntos
Animais , Feminino , Proteínas Sanguíneas/análise , Fase Folicular/sangue , Fase Luteal/sangue , Ciclo Estral , Cavalos , Eletroforese das Proteínas Sanguíneas/veterinária
12.
Rev. bras. ciênc. vet ; 26(3): 111-115, jul./set. 2019. ilus, tab
Artigo em Português | LILACS, VETINDEX | ID: biblio-1491645

Resumo

O objetivo deste estudo foi obter o perfil eletroforético das proteínas séricas em éguas cíclicas e verificar as diferenças entre as fases folicular e luteal do ciclo estral nesta espécie. Foram utilizadas 18 éguas, totalizando 36 amostras de soro, sendo duas de cada égua. As amostras foram colhidas no estro e no diestro. As proteínas séricas totais foram obtidas pelo método do Biureto, a partir da utilização de Kits comerciais (LABTEST®) e, as diferentes subfrações proteicas, por eletroforese em gel de poliacrilamida (SDS-PAGE). O eletroforetograma das proteínas séricas colocou em evidência a presença de 17 a 25 frações proteicas, cujos pesos moleculares variaram de 22 a 254 kDa. Identificaram-se duas proteínas ainda não nomeadas oficialmente, de massas moleculares (MM) 23 kDa e 144 kDa. Os valores médios ± SEM obtidos para cada variável no estro e no diestro, respectivamente, foram: proteínas totais (g/dL) 7,11 ± 0,07 e 7,36 ± 0,07; albumina (mg/dL) 4790,83 ± 69,10 e 5027,19 ± 69,10; α1 glicoproteína ácida (mg/dL) 4,90 ± 0,31 e 4,93 ± 0,31; ceruloplasmina (mg/dL) 15,28 ± 1,31 e 10,65 ± 1,31; haptoglobina (mg/dL) 22,70 ± 1,16 e 27,06 ± 1,16; transferrina (mg/dL) 329,00 ± 9,78 e 350,16 ± 9,78; IgA (mg/dL) 119,91 ± 6,30 e 107,03 ± 6,30; IgG (mg/dL) 1525,07 ± 40,18 e 1517,25 ± 40,18; MM 23 (mg/dL) 204,44 ± 8,61 e 219,79 ± 8,61; MM 144 (mg/dL) 22,13 ± 0,55 e 21,49 ± 0,55. Não houve diferença significativa das proteínas totais e suas frações do estro para o diestro. Conclui-se que as modificações hormonais durante as fases do ciclo estral da égua não interferem no proteinograma sérico.


This study aimed to obtain the electrophoretic profile of serum proteins in cyclic mares and to verify the differences between the follicular and luteal phases of the estrous cycle in this species. Eighteen mares were used, totaling 36 serum samples, two of each mare. Samples were collected both in estrus and in diestrus. Total serum proteins were obtained by the Biureto method, by using commercial kits (LABTEST®), while the different protein subfractions by polyacrylamide gel electrophoresis (SDS-PAGE). The electroforetogram of serum proteins evidenced the presence of 17 to 25 protein fractions, whose molecular weights ranged from 22 to 254 kDa. Two proteins that were not yet officially named were identified, of molecular weights (MW) of 23 kDa and 144 kDa. The mean values (± SEM) obtained for each variable in estrus and diestrus were, respectively: total proteins (g/dL) 7.11 ± 0.07 and 7.36 ± 0.07; albumin (mg/dL) 4790.83 ± 69.10 and 5027.19 ± 69.10; α1 acid glycoprotein (mg/dL) 4.90 ± 0.31 and 4.93 ± 0.31; ceruloplasmine (mg/dL) 15.28 ± 1.31 and 10.65 ± 1.31; haptoglobine (mg/dL) 22.70 ± 1.16 and 27.06 ± 1.16; transferrin (mg/ dL) 329.00 ± 9.78 and 350.16 ± 9.78; IgA (mg/dL) 119.91 ± 6.30 and 107.03 ± 6.30; IgG (mg/dL) 1525.07 ± 40.18 and 1517.25 ± 40.18; MW 23 (mg/dL) 204.44 ± 8.61 and 219.79 ± 8.61; MW 144 (mg/dL) 22.13 ± 0.55 and 21.49 ± 0.55. No significant difference was verified in total proteins and its fractions in estrus and diestrus. The hormonal changes during the specific stages of the estrous cycle of the mare do not interfere with the serum proteinogram.


Assuntos
Feminino , Animais , Cavalos , Ciclo Estral , Fase Folicular/sangue , Fase Luteal/sangue , Proteínas Sanguíneas/análise , Eletroforese das Proteínas Sanguíneas/veterinária
13.
Anim. Reprod. (Online) ; 16(4): 914-922, 2019. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461478

Resumo

The aim of this study was to evaluate the long-term effects of recombinant bovine somatotropin (rbST) on follicle population and ovulatory follicle development in non-lactating dairy cows. Twenty-one Jersey cows were allocated in rbST (n=11) or control (n=10) groups. On day -60, cows in rbST group received 500 mg of somatotropin (s.c. Lactotropin®, Elanco). On day 0, control and rbST cows received an intravaginal progesterone-releasing device (1.9 g, CIDR®, Zoetis) and GnRH (100 mg, IM, Factrel®, Zoetis). On day 8, cows received PGF2α (25 mg, IM, Lutalyse®, Zoetis) and the CIDR® was removed. Twelve hours after device removal (D8), serum, follicular fluid and granulosa cells samples were collected. Serum and follicular concentration of estradiol (E2) and progesterone (P4) were analyzed. Total RNA was extracted from granulosa cells to measure gene expression of LHCGR, STAR, HSD-3B1, CYP11A1, CYP19A1, CYP17A1, IGFR and PAPPA by real-time PCR. Ultrasonography was performed on days -60, -53, -46, -14, -7, 0 and 8 for antral follicle count. Results were analyzed by repeated measures ANOVA and t-test. There was no effect of rbST treatment on the number of follicles during the 60 days period, as well as no effect on serum and follicular fluid E2 and follicular fluid P4 at the moment of follicle aspiration. There was a reduction in PAPPA (P = 0.006), CYP11A1 (P = 0.04) and CYP19A1 (P = 0.002) mRNA levels in granulosa cells of the pre-ovulatory follicle of rbST treated cows. In conclusion, a single dose of rbST did not have long-term effects on antral follicle population, serum and follicular E2/P4 concentrations in non-lactating dairy cows. Despite that, rbST injection decreased granulosa cell expression of genes related to steroidogenesis in the pre-ovulatory follicle.


Assuntos
Feminino , Animais , Bovinos , Bovinos/genética , Expressão Gênica , Hormônio do Crescimento/análise , Hormônio do Crescimento/efeitos adversos , Fase Folicular
14.
Rev. bras. ciênc. vet ; 26(3): 104-110, jul./set. 2019. il.
Artigo em Português | LILACS, VETINDEX | ID: biblio-1391261

Resumo

A obtenção de oócitos de boa qualidade é essencial para o sucesso de diversas biotécnicas reprodutivas. Objetivou-se determinar o efeito de duas técnicas na recuperação de oócitos de boa qualidade em gatas e cadelas em diferentes estágios reprodutivos. Foram utilizados 43 pares de ovários de gata e 35 de cadela após realização da ovariosalpingohisterectomia eletiva. A fase do ciclo estral foi classificada em inativa, folicular ou luteal. Os ovários da fase folicular foram divididos em três grupos: PUN) punção dos folículos com agulha; PUN+FAT) fatiamento do mesmo ovário já puncionado; e FAT) fatiamento do segundo ovário. Os ovários das fêmeas em fase luteal e inativa foram submetidos ao FAT. Foram obtidos no total 974 oócitos (~23/animal) nas fêmeas felinas e 940 (~27/animal) nas caninas. O fatiamento recuperou número superior (P<0,05) de oócitos. Não houve diferença (P>0,05) entre as técnicas de coleta na qualidade de estruturas recuperadas. A quantidade de oócitos recuperados em cada fase foi similar (P>0,05). Contudo, a fase inativa foi superior à luteal (P<0,05) e semelhante à folicular na quantidade de oócitos de boa qualidade em gatas e não houve diferença em cadelas. Conclui-se que o fatiamento recupera maior quantidade de oócitos, não influenciando em sua qualidade. As fases inativa e folicular recuperam maior quantidade de oócitos de boa qualidade em gatas e não afetam a recuperação em cadelas. Portanto, para otimizar o uso das biotecnologias, deve-se levar em consideração o estágio do ciclo estral em fêmeas felinas e a técnica de coleta utilizada na recuperação de oócitos.


The recovery of good quality oocytes is essential for the success of various reproductive biotechniques. The aim of this study was to determine the effect of two techniques on the recovery of good quality oocytes in queens and bitches at different reproductive stages. A total of 43 pairs of ovaries of queens and 35 of bitches after elective ovariosalpingohisterectomy were performed. The estrous cycle phase was classified as inactive, follicular or luteal. The ovaries of the follicular phase were allocated into three groups: PUN) puncture of the follicles with a needle; PUN + SLI) slicing of the same ovary already punctured; and SLI) slicing of the second ovary. The ovaries of luteal and inactive females were submitted to SLI. A total of 974 oocytes (~23/animal) were obtained in feline females and 940 (~27/animal) in canines females. The SLI technique recovered superior number (P<0.05) of oocytes. There was no difference (P>0.05) between the collection techniques in the quality of recovered structures. The number of oocytes recovered in each phase was similar (P>0.05). However, the inactive phase was higher than luteal (P<0.05) and similar to the follicular phase in the quantity of good-quality oocytes in queens and there was no difference in bitches. In conclusion, it is preferable to perform the slicing technique to recover more oocytes in both species. Moreover, in queens it is possible to obtain good quality oocytes in the inactive phase and in bitches the estrous cycle phase does not influence the quality.


Assuntos
Animais , Gatos , Cães , Oócitos , Gatos/anatomia & histologia , Técnicas Reprodutivas/veterinária , Ciclo Estral , Cães/anatomia & histologia , Recuperação de Oócitos/veterinária , Ovário , Fase Folicular , Fase Luteal
15.
Anim. Reprod. ; 16(4): 914-922, 2019. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-24190

Resumo

The aim of this study was to evaluate the long-term effects of recombinant bovine somatotropin (rbST) on follicle population and ovulatory follicle development in non-lactating dairy cows. Twenty-one Jersey cows were allocated in rbST (n=11) or control (n=10) groups. On day -60, cows in rbST group received 500 mg of somatotropin (s.c. Lactotropin®, Elanco). On day 0, control and rbST cows received an intravaginal progesterone-releasing device (1.9 g, CIDR®, Zoetis) and GnRH (100 mg, IM, Factrel®, Zoetis). On day 8, cows received PGF2α (25 mg, IM, Lutalyse®, Zoetis) and the CIDR® was removed. Twelve hours after device removal (D8), serum, follicular fluid and granulosa cells samples were collected. Serum and follicular concentration of estradiol (E2) and progesterone (P4) were analyzed. Total RNA was extracted from granulosa cells to measure gene expression of LHCGR, STAR, HSD-3B1, CYP11A1, CYP19A1, CYP17A1, IGFR and PAPPA by real-time PCR. Ultrasonography was performed on days -60, -53, -46, -14, -7, 0 and 8 for antral follicle count. Results were analyzed by repeated measures ANOVA and t-test. There was no effect of rbST treatment on the number of follicles during the 60 days period, as well as no effect on serum and follicular fluid E2 and follicular fluid P4 at the moment of follicle aspiration. There was a reduction in PAPPA (P = 0.006), CYP11A1 (P = 0.04) and CYP19A1 (P = 0.002) mRNA levels in granulosa cells of the pre-ovulatory follicle of rbST treated cows. In conclusion, a single dose of rbST did not have long-term effects on antral follicle population, serum and follicular E2/P4 concentrations in non-lactating dairy cows. Despite that, rbST injection decreased granulosa cell expression of genes related to steroidogenesis in the pre-ovulatory follicle.(AU)


Assuntos
Animais , Feminino , Bovinos , Bovinos/genética , Hormônio do Crescimento/efeitos adversos , Hormônio do Crescimento/análise , Expressão Gênica , Fase Folicular
16.
Ars Vet. ; 35(1): 38-42, mar. 2019. tab
Artigo em Português | VETINDEX | ID: vti-21778

Resumo

O objetivo do presente estudo foi avaliar a ocorrência da ciclicidade, o diâmetro do folículo dominante e do grau de edema uterino de éguas criadas na zona da mata do Estado de Rondônia, durante o período de transição da estacionalidade reprodutiva. Para este propósito, durante os meses de agosto e setembro (transição de primavera), éguas da raça Quarto de Milha (n = 46), hígidas, criadas a campo ou estabuladas, com idades variando de 2 a 12 anos foram submetidas a dois exames ginecológicos com intervalo de 14 dias, a fim de se determinar e/ou monitorar a presença de corpo lúteo, o diâmetro do folículo dominante e o grau de edema uterino. Empregando um modelo de regressão logística, o efeito do sistema de criação (a campo ou estabuladas) e da faixa etária ( 5, de 6 a 9 e 10 anos) foram avaliados sobre a ocorrência da ciclicidade, bem como sobre o diâmetro do folículo dominante e o grau de edema uterino por meio do modelo linear generalizado misto. Uma maior proporção de éguas (p = 0,001) foi considerada acíclica em relação às cíclicas [68,9% (31/45) vs. 31,1% (14/45), respectivamente]. Dentre as éguas acíclicas, houve uma semelhança (p = 0,127) na proporção daquelas com folículo dominante ou com folículos pequenos. O sistema de criação não influenciou (p > 0,05) a ocorrência da ciclicidade, o diâmetro do folículo dominante e o grau de edema uterino. A faixa etária (≤ 5, de 6 a 9 e ≥ 10 anos) determinou influencia (p < 0,05) na ocorrência da ciclicidade [28,6 b; 18,2b e 66,7% a; respectivamente] e no grau de edema uterino (1,0 ± 0,1 b; 2,0 ± 0,1 a e 2,0 ± 0,2 a; respectivamente). Conclui-se que a maioria das éguas criadas na região da zona da mata rondoniense apresentam-se acíclicas no período de transição de primavera. No entanto, a ocorrência da ciclicidade e o grau de edema uterino demonstraram ser influenciados pela faixa etária das éguas avaliadas.(AU)


The objective of this study was to evaluate the occurrence of cyclicity, the diameter of the dominant follicle and the degree of uterine edema in mares from forest area of the Rondonia State, during the transitional period of reproductive season. For this purpose, during the months of August and September (spring transition), healthy mares, Quarter Horse breed (N = 46), field or housed and aged 2 to 12 years were submitted to two examinations gynecological examination with a 14-day interval. The exams were performed to determine and / or monitor the presence of corpus luteum, the diameter of the dominant follicle and the degree of uterine edema. Using a logistic regression model, the effect of the breeding system (field or stabled) and age group (≤ 5, from 6 to 9 and ≥ 10 years) were evaluated on the occurrence of cyclicity as well as on the diameter of the dominant follicle and the degree of uterine edema by generalized linear mixed model. A higher proportion of mares (P = 0.001) was considered acyclical in relation to the cyclical (68.9% (31/45) vs. 31.1% (14/45), respectively]. Among the acyclic mares, there was a similarity (P = 0.127) in proportion to those with dominant follicle or with small follicles. The breeding system did not influence (P > 0.05) the occurrence of cyclicity, the diameter of the dominant follicle and the degree of uterine edema. The age group (≤ 5, from 6 to 9 and ≥ 10 years) determined influence (P < 0.05) on the occurrence of cyclicity (28.6 b; 18.2 b and 66.7% a; respectively) and in the degree of uterine edema (1.0 ± 0.1 b, 2.0 ± 0.1 a and 2.0 ± 0.2 a, respectively). It is concluded that the majority of the mares created in the forest zone of Rondônia region are acyclical in the period of transition of spring. However, the occurrence of cyclicity and the degree of uterine edema were influenced by the age range of the mares evaluated.(AU)


Assuntos
Animais , Feminino , Gravidez , Cavalos/fisiologia , Fase Folicular , Prenhez/imunologia , Prenhez/metabolismo , Ovulação/genética
17.
Ars vet ; 35(1): 38-42, mar. 2019. tab
Artigo em Português | VETINDEX | ID: biblio-1463487

Resumo

O objetivo do presente estudo foi avaliar a ocorrência da ciclicidade, o diâmetro do folículo dominante e do grau de edema uterino de éguas criadas na zona da mata do Estado de Rondônia, durante o período de transição da estacionalidade reprodutiva. Para este propósito, durante os meses de agosto e setembro (transição de primavera), éguas da raça Quarto de Milha (n = 46), hígidas, criadas a campo ou estabuladas, com idades variando de 2 a 12 anos foram submetidas a dois exames ginecológicos com intervalo de 14 dias, a fim de se determinar e/ou monitorar a presença de corpo lúteo, o diâmetro do folículo dominante e o grau de edema uterino. Empregando um modelo de regressão logística, o efeito do sistema de criação (a campo ou estabuladas) e da faixa etária ( 5, de 6 a 9 e 10 anos) foram avaliados sobre a ocorrência da ciclicidade, bem como sobre o diâmetro do folículo dominante e o grau de edema uterino por meio do modelo linear generalizado misto. Uma maior proporção de éguas (p = 0,001) foi considerada acíclica em relação às cíclicas [68,9% (31/45) vs. 31,1% (14/45), respectivamente]. Dentre as éguas acíclicas, houve uma semelhança (p = 0,127) na proporção daquelas com folículo dominante ou com folículos pequenos. O sistema de criação não influenciou (p > 0,05) a ocorrência da ciclicidade, o diâmetro do folículo dominante e o grau de edema uterino. A faixa etária (≤ 5, de 6 a 9 e ≥ 10 anos) determinou influencia (p < 0,05) na ocorrência da ciclicidade [28,6 b; 18,2b e 66,7% a; respectivamente] e no grau de edema uterino (1,0 ± 0,1 b; 2,0 ± 0,1 a e 2,0 ± 0,2 a; respectivamente). Conclui-se que a maioria das éguas criadas na região da zona da mata rondoniense apresentam-se acíclicas no período de transição de primavera. No entanto, a ocorrência da ciclicidade e o grau de edema uterino demonstraram ser influenciados pela faixa etária das éguas avaliadas.


The objective of this study was to evaluate the occurrence of cyclicity, the diameter of the dominant follicle and the degree of uterine edema in mares from forest area of the Rondonia State, during the transitional period of reproductive season. For this purpose, during the months of August and September (spring transition), healthy mares, Quarter Horse breed (N = 46), field or housed and aged 2 to 12 years were submitted to two examinations gynecological examination with a 14-day interval. The exams were performed to determine and / or monitor the presence of corpus luteum, the diameter of the dominant follicle and the degree of uterine edema. Using a logistic regression model, the effect of the breeding system (field or stabled) and age group (≤ 5, from 6 to 9 and ≥ 10 years) were evaluated on the occurrence of cyclicity as well as on the diameter of the dominant follicle and the degree of uterine edema by generalized linear mixed model. A higher proportion of mares (P = 0.001) was considered acyclical in relation to the cyclical (68.9% (31/45) vs. 31.1% (14/45), respectively]. Among the acyclic mares, there was a similarity (P = 0.127) in proportion to those with dominant follicle or with small follicles. The breeding system did not influence (P > 0.05) the occurrence of cyclicity, the diameter of the dominant follicle and the degree of uterine edema. The age group (≤ 5, from 6 to 9 and ≥ 10 years) determined influence (P < 0.05) on the occurrence of cyclicity (28.6 b; 18.2 b and 66.7% a; respectively) and in the degree of uterine edema (1.0 ± 0.1 b, 2.0 ± 0.1 a and 2.0 ± 0.2 a, respectively). It is concluded that the majority of the mares created in the forest zone of Rondônia region are acyclical in the period of transition of spring. However, the occurrence of cyclicity and the degree of uterine edema were influenced by the age range of the mares evaluated.


Assuntos
Feminino , Animais , Gravidez , Cavalos/fisiologia , Fase Folicular , Ovulação/genética , Prenhez/imunologia , Prenhez/metabolismo
18.
Rev. bras. ciênc. vet ; 26(3): 111-115, jul./set. 2019. il.
Artigo em Português | LILACS, VETINDEX | ID: biblio-1391268

Resumo

O objetivo deste estudo foi obter o perfil eletroforético das proteínas séricas em éguas cíclicas e verificar as diferenças entre as fases folicular e luteal do ciclo estral nesta espécie. Foram utilizadas 18 éguas, totalizando 36 amostras de soro, sendo duas de cada égua. As amostras foram colhidas no estro e no diestro. As proteínas séricas totais foram obtidas pelo método do Biureto, a partir da utilização de Kits comerciais (LABTEST) e, as diferentes subfrações proteicas, por eletroforese em gel de poliacrilamida (SDS-PAGE). O eletroforetograma das proteínas séricas colocou em evidência a presença de 17 a 25 frações proteicas, cujos pesos moleculares variaram de 22 a 254 kDa. Identificaram-se duas proteínas ainda não nomeadas oficialmente, de massas moleculares (MM) 23 kDa e 144 kDa. Os valores médios ± SEM obtidos para cada variável no estro e no diestro, respectivamente, foram: proteínas totais (g/dL) 7,11 ± 0,07 e 7,36 ± 0,07; albumina (mg/dL) 4790,83 ± 69,10 e 5027,19 ± 69,10; α1 glicoproteína ácida (mg/dL) 4,90 ± 0,31 e 4,93 ± 0,31; ceruloplasmina (mg/dL) 15,28 ± 1,31 e 10,65 ± 1,31; haptoglobina (mg/dL) 22,70 ± 1,16 e 27,06 ± 1,16; transferrina (mg/dL) 329,00 ± 9,78 e 350,16 ± 9,78; IgA (mg/dL) 119,91 ± 6,30 e 107,03 ± 6,30; IgG (mg/dL) 1525,07 ± 40,18 e 1517,25 ± 40,18; MM 23 (mg/dL) 204,44 ± 8,61 e 219,79 ± 8,61; MM 144 (mg/dL) 22,13 ± 0,55 e 21,49 ± 0,55. Não houve diferença significativa das proteínas totais e suas frações do estro para o diestro. Conclui-se que as modificações hormonais durante as fases do ciclo estral da égua não interferem no proteinograma sérico.


This study aimed to obtain the electrophoretic profile of serum proteins in cyclic mares and to verify the differences between the follicular and luteal phases of the estrous cycle in this species. Eighteen mares were used, totaling 36 serum samples, two of each mare. Samples were collected both in estrus and in diestrus. Total serum proteins were obtained by the Biureto method, by using commercial kits (LABTEST), while the different protein subfractions by polyacrylamide gel electrophoresis (SDS-PAGE). The electroforetogram of serum proteins evidenced the presence of 17 to 25 protein fractions, whose molecular weights ranged from 22 to 254 kDa. Two proteins that were not yet officially named were identified, of molecular weights (MW) of 23 kDa and 144 kDa. The mean values (± SEM) obtained for each variable in estrus and diestrus were, respectively: total proteins (g/dL) 7.11 ± 0.07 and 7.36 ± 0.07; albumin (mg/dL) 4790.83 ± 69.10 and 5027.19 ± 69.10; α1 acid glycoprotein (mg/dL) 4.90 ± 0.31 and 4.93 ± 0.31; ceruloplasmine (mg/dL) 15.28 ± 1.31 and 10.65 ± 1.31; haptoglobine (mg/dL) 22.70 ± 1.16 and 27.06 ± 1.16; transferrin (mg/dL) 329.00 ± 9.78 and 350.16 ± 9.78; IgA (mg/dL) 119.91 ± 6.30 and 107.03 ± 6.30; IgG (mg/dL) 1525.07 ± 40.18 and 1517.25 ± 40.18; MW 23 (mg/dL) 204.44 ± 8.61 and 219.79 ± 8.61; MW 144 (mg/dL) 22.13 ± 0.55 and 21.49 ± 0.55. No significant difference was verified in total proteins and its fractions in estrus and diestrus. The hormonal changes during the specific stages of the estrous cycle of the mare do not interfere with the serum proteinogram.


Assuntos
Animais , Proteínas Sanguíneas/análise , Ciclo Estral , Eletroforese/veterinária , Fase Folicular , Cavalos/anatomia & histologia , Fase Luteal
19.
Anim. Reprod. (Online) ; 16(3): 449-454, 2019. graf
Artigo em Inglês | VETINDEX | ID: biblio-1461454

Resumo

IVF success depends on hundreds of factors and details but the oocyte quality remains the most important and problematic issue. All antral follicles contain oocytes and all of them have that have reached their full size, can be aspirated, can mature and can be fertilized in vitro. But only a few will make it to embryo unless harvested at a very specific time/status. The conditions impacting the oocyte competence are essentially dependant on the follicular status. Growing follicles contains oocytes that have not completed their preparation, as they are still writing information (RNA), later, dominant follicles or follicles at the plateau phase, stop transcription and become candidates for development. Once in transcriptional arrest, the oocytes, if not ovulated in a short amount of time, do not always make good embryos. This window is affected by time and follicle size and looks like a bell curve. The following review further explain the physiological and molecular evidences that we have to illustrate the competence window and provides clues on how to optimize ovarian stimulation to maximise oocyte quality.


Assuntos
Feminino , Animais , Bovinos , Bovinos/embriologia , Fase Folicular/genética , Fertilização in vitro/métodos , Oócitos/crescimento & desenvolvimento
20.
Anim. Reprod. ; 16(3): 449-454, 2019. graf
Artigo em Inglês | VETINDEX | ID: vti-22355

Resumo

IVF success depends on hundreds of factors and details but the oocyte quality remains the most important and problematic issue. All antral follicles contain oocytes and all of them have that have reached their full size, can be aspirated, can mature and can be fertilized in vitro. But only a few will make it to embryo unless harvested at a very specific time/status. The conditions impacting the oocyte competence are essentially dependant on the follicular status. Growing follicles contains oocytes that have not completed their preparation, as they are still writing information (RNA), later, dominant follicles or follicles at the plateau phase, stop transcription and become candidates for development. Once in transcriptional arrest, the oocytes, if not ovulated in a short amount of time, do not always make good embryos. This window is affected by time and follicle size and looks like a bell curve. The following review further explain the physiological and molecular evidences that we have to illustrate the competence window and provides clues on how to optimize ovarian stimulation to maximise oocyte quality.(AU)


Assuntos
Animais , Feminino , Bovinos , Bovinos/embriologia , Fase Folicular/genética , Oócitos/crescimento & desenvolvimento , Fertilização in vitro/métodos
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