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1.
Reprod Sci ; 2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33721296

RESUMO

The development of culture systems capable of maintaining follicular growth since the preantral stage has been the target of investigations. Mesenchymal stem cells (MSC) present potential for use in a wide range of applications, including research aimed at preserving fertility. Therefore, this study investigated the use of caprine Wharton's Jelly Mesenchymal Stem Cells (WJMSC) on the survival and in vitro development of goat preantral follicles enclosed in ovarian fragments cultured for 1 or 7 days. Fragments of the ovarian cortex were immediately fixed (non-cultured control) or distributed in four treatments: ovarian tissue cultured in control medium (α-MEM+); ovarian tissue cultured in α-MEM+ supplemented with FBS (α-MEM+ + FBS); ovarian tissue co-cultured with stem cells in α-MEM+ (α-MEM+ + SC); and ovarian tissue co-cultured with stem cell in α-MEM+ + FBS (α-MEM+ + SC + FBS). The rates of cell proliferation, follicular survival, and activation, as well as follicular diameter, were evaluated. After 7 days, the treatment co-cultured with stem cells showed a higher (P < 0.05) percentage of morphologically normal preantral follicles compared to the other treatments, as well as a higher (P < 0.05) activation rate compared to cultured control. Moreover, the follicular diameter was higher (P < 0.05) in the treatment co-cultured with stem cells compared to co-cultured with stem cells plus FBS. This study demonstrates for the first time that in vitro co-culture of caprine WJMSC with preantral follicles enclosed in goat ovarian tissue improves activation and early follicular development.

2.
Reprod Sci ; 27(8): 1602-1608, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32436196

RESUMO

Oxidative stress is one of the most detrimental factors that affect oocyte developmental competence and embryo development in vitro. The impact of anethole supplementation to in vitro maturation (IVM) media on oocyte maturation and further bovine in vitro embryo production was investigated. Oocytes of slaughterhouse-derived bovine ovaries were placed in IVM with anethole at different concentrations of 30 (AN30), 300 (AN300), and 2000 µg/mL (AN2000), or without (control treatment). The oocytes were assessed for maturation rates, and for reactive oxygen species (ROS) and ferric reducing antioxidant power (FRAP) levels, and mitochondrial membrane potential. Embryo development was assessed by cleavage and blastocyst rates, and embryo cell number. The percentage of metaphase II oocytes were similar among the treatments (range, 77%-96%). Anethole at 300 µg/mL was the only treatment that yielded higher cleavage and embryo development (morula and blastocyst) rates compared to the control treatment. The ROS production in the oocytes after maturation did not differ among treatments. However, oocytes treated with anethole at 300 µg/mL had higher (P < .05) FRAP and mitochondrial membrane potential compared to the control treatment. Furthermore, AN300 treatment increased (P < .05) the average number of total cells in blastocysts compared to the control and AN30 treatments. The use of anethole at 300 µg/mL during IVM is suggested to improve the quantity and quality of bovine embryos produced in vitro. The beneficial effects of anethole on embryonic developmental competence in vitro seems to be related to its capacity to regulate the redox balance and improve mitochondrial function in oocytes and embryos.

3.
Anim Reprod Sci ; 215: 106310, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32216933

RESUMO

An appropriate implantation site favors angiogenesis and avoids ovarian tissue damage after tissue grafting. The objective of this study was to evaluate the effects of intramuscular (IM) and subcutaneous (SC) sites for ovarian grafts in goats by evaluating follicular morphology and activation, preantral follicle and stromal cell densities, tissue DNA fragmentation, collagen types I and III depositions, and graft revascularizations. Ovarian cortical tissue was transplanted in IM or SC sites and recovered 7 or 15 days post-transplantation. There was a greater percentage of developing follicles and lesser follicular and stromal cell densities in all grafted tissues as compared to ovarian tissues of the control group. The stromal cell density and percentage of normal follicles were positively associated. At 15 days post-transplantation, tissues at the SC and IM sites had similar amounts of DNA fragmentation and type III collagen content. In contrast, tissues at the SC, as compared with IM site, had greater abundances of collagen type I. Furthermore, there was a positive association between collagen type I and percentage of morphologically normal follicles post-transplantation. In addition to a marked decrease in follicular density 15 days post-transplantation in ovarian grafts at the SC and IM sites, low percentages of normal follicles and follicular activation were observed similarly in both transplantation sites. There were also positive associations of stromal cell density and abundance of type I collagen fibers with the percentage of intact follicles in grafted ovarian tissues.

4.
Anim Reprod Sci ; 205: 126-133, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31047761

RESUMO

The aim of the present study was to establish a protocol for solid surface vitrification of peccary ovarian tissue by using different cryoprotectants. Ovarian pairs from five adult females were fragmented and two fragments (fresh control group) were immediately subjected to morphological evaluation using classical histology, transmission electron microscopy, and viability analysis using fluorescent probes. The remaining fragments (n = 18) were vitrified using a solid surface method with different concentrations (3 or 6 M) of ethylene glycol (EG), dimethyl sulfoxide (DMSO) or dimethyl formamide (DMF). After 2 weeks, samples were re-warmed and evaluated. A decrease in the percentage of morphologically normal preantral follicles (PFs) was verified for all the groups in comparison to the fresh control (92.0 ± 2.8%); however, if only the primordial follicles are considered, the most effective preservation (P < 0.05) was achieved with the use of EG at 3 M (74.2±7.3%) or DMSO at 6 M (75.0 ± 4.2%). Ultrastructural analysis indicated there were well-preserved PFs in all the groups evaluated, having well-defined membranes, a few vacuoles, and organelles that were uniformly distributed throughout the cytoplasm, mainly round and elongated mitochondria in close association with lipid droplets. Viability was preserved (P < 0.05) with the use of EG at 3 (97%) or 6 (97%) M, DMSO at 3 (100%), and DMF at 6 (97%) M. Solid surface vitrification, therefore, is an effective method for conservation of peccary female germplasm, especially with the use of EG at 3 M, which was highly effective for preservation of both the morphology and viability of PFs.


Assuntos
Artiodáctilos/fisiologia , Crioprotetores/farmacologia , Ovário/fisiologia , Preservação de Tecido/veterinária , Vitrificação/efeitos dos fármacos , Animais , Sobrevivência Celular , Feminino
5.
Reprod Sci ; : 1933719119831783, 2019 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808260

RESUMO

Oxidative stress is one of the most detrimental factors that affect oocyte developmental competence and embryo development in vitro. The impact of anethole supplementation to in vitro maturation (IVM) media on oocyte maturation and further bovine in vitro embryo production was investigated. Oocytes of slaughterhouse-derived bovine ovaries were placed in IVM with anethole at different concentrations of 30 (AN30), 300 (AN300), and 2000 µg/mL (AN2000), or without (control treatment). The oocytes were assessed for maturation rates, and for reactive oxygen species (ROS) and ferric reducing antioxidant power (FRAP) levels, and mitochondrial membrane potential. Embryo development was assessed by cleavage and blastocyst rates, and embryo cell number. The percentage of metaphase II oocytes were similar among the treatments (range, 77%-96%). Anethole at 300 µg/mL was the only treatment that yielded higher cleavage and embryo development (morula and blastocyst) rates compared to the control treatment. The ROS production in the oocytes after maturation did not differ among treatments. However, oocytes treated with anethole at 300 µg/mL had higher ( P < .05) FRAP and mitochondrial membrane potential compared to the control treatment. Furthermore, AN300 treatment increased ( P < .05) the average number of total cells in blastocysts compared to the control and AN30 treatments. The use of anethole at 300 µg/mL during IVM is suggested to improve the quantity and quality of bovine embryos produced in vitro. The beneficial effects of anethole on embryonic developmental competence in vitro seems to be related to its capacity to regulate the redox balance and improve mitochondrial function in oocytes and embryos.

6.
Anim Reprod Sci ; 196: 120-129, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30049427

RESUMO

The present study aimed to investigate a concentration-response curve of human recombinant FSH (hrFSH) for in vitro culture of isolated preantral and early antral follicles of goats. Isolated follicles were cultured for 18 days using the following treatments: basic culture medium (control); or control medium supplemented with 10, 50, and 100 mIU/mL of hrFSH. At the end of the culture, cumulus-oocyte complexes were recovered and subjected to in vitro maturation. The following endpoints were evaluated: follicle morphology, growth rate and antrum formation, oocyte viability and meiotic stage, and estradiol production, as well as relative expression of FSH receptor (FSHR), and steroidogenic enzyme (3ß-HSD, CYP17, and CYP19A1) genes. In antral follicles, the FSH addition at 50 mIU/mL increased follicular diameter and growth rate, percentage of fully developed oocytes, and oocyte diameter (P < 0.05), and tended to increase the percentage of MII oocytes when compared to the control (P = 0.07). With preantral follicles, FSH addition at 100 mIU/mL increased relative abundance of mRNA for CYP19A1 when compared to the control (P < 0.05). At the same FSH concentrations of 100 and 50 mIU/mL, there was a greater relatively abundance of mRNA for 3ß-HSD and CYP17 in preantral than in antral follicles (P < 0.05). For preantral and antral follicle comparisons when the same treatments were imposed, there were greater concentrations of estradiol for antral follicles (P < 0.05). In conclusion, hrFSH enhanced in a concentration-dependent manner the in vitro development of caprine antral follicles; however, there was no positive effect in the culture of preantral follicles.


Assuntos
Hormônio Foliculoestimulante Humano/farmacologia , Cabras , Folículo Ovariano/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Feminino , Hormônio Foliculoestimulante , Humanos , Oócitos , Folículo Ovariano/fisiologia
7.
Cryobiology ; 83: 97-99, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29908142

RESUMO

We aimed to evaluate the effect of three extracellular cryoprotectants on the morphology of vitrified feline preantral follicles. Feline ovarian fragments (0.5 × 2 × 2 mm) collected from five domestic adult cats subjected to ovariohysterectomy for routine castration were vitrified with ethylene glycol (EG) 40% combined or not with sucrose (0.1 or 0.5 M), trehalose (0.1 or 0.5 M), or raffinose (0.1 M). After vitrification using the solid-surface method and warming of the tissues, cryoprotectants were washed out of the ovarian tissues, which were fixed for histological analysis. The percentages of normal follicles were similar to the control (fresh) (62.9 ± 4.1%) only for tissues exposed and cryopreserved with EG + trehalose at concentrations of 0.1 (35.8 ± 8.3%) and 0.5 M (33.4 ± 5.4%). All the other sugars decreased the percentages of morphologically normal follicles as compared to the control group and the trehalose groups. Based on the results of the present study, we recommend the use of trehalose as the extracellular cryoprotectant for the vitrification of feline ovarian tissue.


Assuntos
Criopreservação/métodos , Crioprotetores/farmacologia , Preservação de Órgãos/métodos , Folículo Ovariano/efeitos dos fármacos , Vitrificação , Animais , Gatos , Etilenoglicol/farmacologia , Feminino , Rafinose/farmacologia , Sacarose/farmacologia , Trealose/farmacologia
8.
Biopreserv Biobank ; 16(4): 258-269, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29957024

RESUMO

AIM: The present study evaluates the effect of different concentrations of antioxidants (catalase - CAT and alpha lipoic acid - ALA) on the follicular activation and morphology, DNA damage, ROS production, and mitochondrial activity in vitrified sheep ovarian tissue. METHODS: This experiment was divided into two steps. First, ovarian fragments were distributed into the following treatments: fresh tissue or control (CTR), incubation (INC), vitrification without antioxidant (VWA), with CAT (10, 20, or 40 IU mL-1) or ALA (25, 50, or 100 µM mL-1). After vitrification/warming, the fragments were additionally incubated for 24 hours and evaluated for morphology and follicular activation, as well as reactive oxygen species (ROS) levels in the culture medium. For the second step, other ovarian fragments were submitted to CTR, VWA, CAT40, and ALA100. After vitrification/warming, the fragments were incubated for 24 hours and evaluated by cell density of ovarian stroma, DNA damage, and mitochondrial and intracellular ROS levels. RESULTS: The percentage of morphologically normal follicles in vitrified ovarian tissue in the presence of ALA in all concentrations did not differ (p > 0.05) from fresh tissue or CTRs. The percentage of activated follicles was higher in ALA100 µM mL-1 than those observed for the treatments INC, CAT (40 IU mL-1), or ALA (25 or 50 µM mL-1). The use of CAT affected (p < 0.05) the density of stromal cells (40 IU mL-1), ROS levels (10 and 20 IU mL-1), as well as DNA damage revealed by ©H2AX (40 IU mL-1). CONCLUSIONS: Although 100 µM/mL of ALA did not alter intracellular ROS, this concentration reduced the levels of ROS in the culture medium, preserved both the follicular morphology, as well as the mitochondrial activity, promoted follicle activation, and protected the follicles from DNA damage.


Assuntos
Catalase/farmacologia , Criopreservação/métodos , Ovário/citologia , Ovário/metabolismo , Ácido Tióctico/farmacologia , Vitrificação , Animais , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Feminino , Folículo Ovariano/citologia , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovário/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Ovinos
9.
Pesqui. vet. bras ; 38(1): 175-182, Jan. 2018. tab, ilus
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-895548

RESUMO

The aim of this study was to characterize the preantral ovarian follicular population in agoutis (D. leporina) by estimating the number of follicles at each developmental category, and also describe the morphometry and the specific features of the follicle and the oocyte by using light and transmission electron microscopy. The length of each ovary was measured using a caliper rule, longitudinally sectioned into two halves and both were immediately fixed to perform the estimation of follicular population and ultrastructural analysis. The mean (±S.E.M.) population of follicular per pair of ovary was estimated at 4419.8±532.26 and 5397.52±574.91 for right and left ovaries, respectively, but no differences were observed between them. The diameters for follicles, oocyte and nuclei were: 18.62±3.40µm, 12.28±2.37µm and 6.10±0.93µm for primordial, 23.75±5.70µm, 14.22±3.00µm and 6.70±1.24µm for primary and 88.55±17.61µm, 52.85±17.56µm and 22.33±17.61µm for secondary follicles, respectively. The most of the follicles found belonged to the primordial category (86.63%), followed by primary (13.01%) and secondary (0.35%) one. Additionally, polyovular follicles were observed in all the animals and they represented 7.51% of the total follicles counted. The ultrastructural analysis showed that the oocyte presented a central and regular nuclei, displaying a homogenous mass. Among the organelles, the mitochondria were the most abundant and the oocyte Golgi apparatus was rarely observed. In conclusion, this work shows for the first time the characterization of the population of preantral follicles in the ovary of Dasyprocta leporina. Those information will be useful for further development and adaptation of biotechniques such as germplasm cryopreservation and in vitro gametes manipulation.(AU)


O objetivo deste trabalho foi caracterizar a população folicular ovariana pré-antral em cutias (D. leporina) estimando o número de folículos em cada categoria de desenvolvimento, e também descrever a morfometria e as características específicas do folículo e oócito usando microscopia de luz e eletrônica de transmissão. O comprimento de cada ovário foi medido utilizando um paquímetro, seccionados longitudinalmente em duas metades e ambos foram imediatamente fixados para realizar a estimativa da população folicular e análise ultraestrutural. A média (±S.E.M.) da população folicular por par de ovário foi estimada em 4419,8±532,26 e 5397,52±574,91 nos ovários direito e esquerdo, respectivamente, mas não foram observadas diferenças entre eles. Os diâmetros dos folículos, oócito e núcleos, respectivamente, foram: 18,62±3,40µm, 12,28±2.37µm e 6,10±0,93µm para primordial, 23,75±5,70µm, 14,22±3,00µm e 6,70±1,24µm para primário e 88,55±17,61µm, 52,85±17,56µm e 22,33±17,61µm de folículos secundários. A maioria dos folículos encontrados pertencia à categoria primordial (86,63%), seguido pelo primário (13,01%) e um secundário (0,35%). Adicionalmente, os folículos poliovulares foram observados em todos os animais e representavam 7,51% do total de folículos contados. A análise ultra-estrutural mostrou que o oócito apresentou núcleos centrais e regulares, exibindo uma massa homogênea. Dentre as organelas, as mitocôndrias foram as mais abundantes e o aparelho de Golgi do oócito foi raramente observado. Em conclusão, este trabalho mostra pela primeira vez a caracterização da população de folículos pré-antrais do ovário da Dasyprocta leporina. Essas informações serão úteis para o desenvolvimento e adaptação de biotécnicas, como a criopreservação de germoplasma e manipulação de gametas in vitro.(AU)


Assuntos
Animais , Dasyproctidae/anatomia & histologia , Oócitos , Folículo Ovariano/anatomia & histologia , Microscopia Eletrônica de Transmissão/veterinária
10.
Microsc Res Tech ; 80(4): 406-418, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27921341

RESUMO

The aims of this study were to investigate the effects of medium replacement system (experiment I) and of FSH presentations (homeopathic - FSH 6cH and allopathic FSH - rFSH; experiment II) on the in vitro development, hormone production and gene expression of isolated ovine preantral follicles cultured for 6 days. In experiment I, secondary follicles were cultured in the α-MEM+ supplemented with FSH 6cH (0.05 fg/ml) or recombinant bovine FSH (100 ng/ml) without/with daily medium addition. The homeopathic FSH treatments with/without medium addition improved (p < .05) follicular development compared to rFSH100 treatment without addition. FSH 6cH with addition showed the highest (p < .05) estradiol production. To verify whether the effects of homeopathic FSH were not due to its vehicle, experiment II was performed. The α-MEM+ was supplemented or not with alcohol (0.2% grain ethanol, v/v), FSH 6cH or rFSH100 with daily medium addition. Surprisingly, we found that all treatments improved follicular development compared to the α-MEM+ (p < .05). Moreover, homeopathic FSH was similar to the other treatments including its vehicle. In conclusion, its vehicle (ethanol) causes the effect of homeopathic FSH on in vitro development of isolated ovine preantral follicles.


Assuntos
Proliferação de Células/efeitos dos fármacos , Etanol/farmacologia , Hormônio Foliculoestimulante/farmacologia , Hormônios/biossíntese , Técnicas de Cultura de Órgãos/métodos , Folículo Ovariano/crescimento & desenvolvimento , Animais , Apoptose/genética , Caspase 3/análise , Conexina 43/análise , Conexinas/análise , Fragmentação do DNA , Estradiol/biossíntese , Etanol/química , Feminino , Homeopatia , Hormônios/farmacologia , Folículo Ovariano/efeitos dos fármacos , Progesterona/biossíntese , Proteínas Recombinantes/farmacologia , Ovinos
11.
Reprod Fertil Dev ; 27(3): 440-8, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25481978

RESUMO

Effective methods for gamete preservation should have low impact on DNA integrity. The present study investigated the effects of vitrification of goat ovarian tissues on the occurrence of DNA fragmentation and DNA double-stand breaks using the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) assay and detection of phosphorylated histone H2AX (γH2AX), respectively. Goat ovaries were collected at a local abattoir and 12 tissue fragments were prepared from each ovarian pair. Tissue fragments were used as fresh control samples or were cultured in vitro, vitrified or vitrified and cultured. Vitrification was performed using the Ovarian Tissue Cryosystem. Fragments from all groups (control and treatments) were processed for histology, transmission electron microscopy, TUNEL assay and immunofluorescence. Compared with fresh control samples, a lower percentage of morphologically normal follicles was detected in the vitrification followed by culture treatment group (P<0.05). Normal follicular ultrastructure was observed in all groups. Immunofluorescence revealed the presence of γH2AX foci in few oocytes and ovarian stromal cells. TUNEL-positive follicles were found in samples without significant differences among groups (P>0.05). In conclusion, the vitrification protocol used in the present study did not increase DNA damage in preantral follicles enclosed in goat ovarian tissues.


Assuntos
Criopreservação/métodos , Crioprotetores/farmacologia , Dano ao DNA/efeitos dos fármacos , Ovário/efeitos dos fármacos , Preservação de Tecido/métodos , Vitrificação , Animais , Feminino , Cabras , Folículo Ovariano/efeitos dos fármacos
12.
Cell Tissue Res ; 348(1): 225-38, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22382392

RESUMO

This study evaluated the levels of bone morphogenetic protein receptors BMPRIB and BMPRII mRNA in goat follicles and the effects of bone morphogenetic protein-15 (BMP-15) on the in vitro development of cultured preantral follicles. Real-time polymerase chain reaction (PCR) was used to analyze the levels of BMPRIB and BMPRII mRNA in caprine preantral follicles and in small and large antral follicles. Preantral follicles (≥150 µm) were also isolated from goat ovaries and cultured for 18 days in α-MEM(+) supplemented with or without BMP-15 (10, 50, or 100 ng/ml). At the end of culture, some follicles were fixed for ultrastructural evaluation. Real-time PCR showed a reduction in BMPRII mRNA levels from the primary to secondary follicles. Higher levels of BMPRIB mRNA were observed in granulosa/theca cells from large antral follicles compared with small antral follicles. Moreover, BMPRII mRNA was expressed to a greater extent in cumulus-oocyte complexes from large antral follicles than in their respective granulosa/theca cells. In culture, 50 ng/ml BMP-15 positively influenced antral cavity formation and follicle growth after 18 days and also maintained follicular integrity. Thus, BMPRIB and BMPRII mRNAs are present in all follicular categories. BMP-15 (50 ng/ml) stimulates growth, antrum formation and the ultrastructural integrity of isolated caprine preantral follicles after 18 days of culture.


Assuntos
Proteína Morfogenética Óssea 15/farmacologia , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Folículo Ovariano/crescimento & desenvolvimento , Folículo Ovariano/metabolismo , Animais , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/metabolismo , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/metabolismo , Proliferação de Células/efeitos dos fármacos , Feminino , Cabras/genética , Humanos , Meiose/efeitos dos fármacos , Microscopia de Fluorescência , Oócitos/citologia , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Folículo Ovariano/citologia , Folículo Ovariano/ultraestrutura , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Técnicas de Cultura de Tecidos , Sobrevivência de Tecidos/efeitos dos fármacos
13.
Biopreserv Biobank ; 10(4): 338-42, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24849881

RESUMO

The objectives of this study were to determine: 1) the optimal concentration (1.0 or 1.5 M) and duration of exposure (5, 10, or 20 min) of ovarian tissue to 1,2-propanediol (PROH) on morphology and viability of caprine preantral follicles; and 2) the effect of supplementing cryopreservation medium supplementation with Trolox(®) (0.1, 0.5, or 1.0 mM) or catalase (5, 10, or 20 IU/mL) on follicular morphology, viability, and lipid peroxidation. Cryopreservation decreased (p<0.05) percentages of normal follicles relative to the control (84%). Although supplementation of the cryopreservation medium (1.0 M PROH) with catalase (10 or 20 IU/mL) or Trolox(®) (0.1 mM) resulted in follicular morphology and viability similar to that in the controls (P>0.05), lipid peroxidation was reduced only when 20 IU/mL catalase was added to the cryopreservation medium.


Assuntos
Catalase/metabolismo , Criopreservação/métodos , Crioprotetores/efeitos adversos , Congelamento , Folículo Ovariano/enzimologia , Propilenoglicol/efeitos adversos , Animais , Feminino , Cabras , Peroxidação de Lipídeos/efeitos dos fármacos , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovário/efeitos dos fármacos , Ovário/enzimologia , Ovário/metabolismo
14.
Pesqui. vet. bras ; 30(4): 305-310, abr. 2010. ilus, graf, tab
Artigo em Inglês | LILACS | ID: lil-548881

RESUMO

This study was conducted in order to verify the effect of different concentrations of BMP-7 in the in vitro survival and development of caprine preantral follicles. Fragments of caprine ovarian cortical tissue were cultured for 1 or 7 days in Minimum Essential Medium (MEM+) supplemented with different concentrations of BMP-7 (1, 10, 50 or 100ng/ml). Non-cultured fragments or those cultured for 1 or 7 days were processed for classical histology and transmission electron microscopy (TEM). Parameters such as follicular survival, activation and growth were evaluated. The results showed that, after 1 or 7 days of culture, the percentage of morphologically normal follicles was significantly reduced in all treatments when compared with fresh control, except at 1ng/ml of BMP-7 for 1 day. In addition, the concentration of 10ng/ml of BMP-7 significantly increases follicular diameter from day 1 to 7 of culture. There was no influence of the other concentrations of BMP-7 regarding to the follicular and oocyte diameter. Ultrastructure studies confirmed follicular integrity after 7 days of culture in 1ng/ml BMP-7. In conclusion, small concentrations of BMP-7 can improve the survival and growth of caprine preantral follicles during in vitro culture.


O presente trabalho foi conduzido de modo a se verificar o efeito de diferentes concentrações da BMP-7 no desenvolvimento in vitro de folículos pré-antrais caprinos. Fragmentos de tecido cortical ovariano caprino foram cultivados por 1 ou 7 dias em Minimum Essential Medium (MEM+) suplementado com diferentes concentrações de BMP-7 (1, 10, 50 ou 100ng/ml). Os fragmentos não cultivados ou aqueles cultivados por 1 ou 7 dias foram processados para histologia clássica e microscopia eletrônica de transmissão (TEM), sendo avaliados parâmetros morfológicos indicativos de viabilidade, ativação e crescimento. Os resultados mostraram que o percentual de folículos morfologicamente normais diminuiu significativamente em todos os tratamentos quando comparados ao controle, exceto na concentração de 1ng/ml por 1 dia de cultivo. Já no D7 todos os tratamentos reduziram significativamente os percentuais de folículos morfologicamente normais. Utilizando 10ng/ml de BMP-7 foi observado um aumento significativo no diâmetro folicular quando comparados os diferentes períodos de cultivo. Não houve influência das demais concentrações de BMP-7 quando avaliados além do diâmetro folicular o diâmetro oocitário. A análise por TEM confirmou a integridade ultra-estrutural nos folículos após 7 dias de cultivo com 1ng/ml de BMP-7 . Em conclusão, o BMP-7 em baixas concentrações pode melhorar a sobrevivência e o crescimento durante o cultivo in vitro de folículos pré-antrais caprinos.


Assuntos
Animais , Ovário/anatomia & histologia , Proteínas Morfogenéticas Ósseas/administração & dosagem , Cabras
15.
Biopreserv Biobank ; 8(4): 219-21, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24846107

RESUMO

Exposure time and addition of sucrose to the vitrification medium as well as the solid-surface vitrification (SSV) on the morphology of bovine preantral follicles were evaluated. Ovarian tissue was exposed for 1, 5, or 10 min to 4.0 M ethylene glycol with or without the addition of 0.5 M sucrose. Subsequently, the tissue was washed out from cryoprotectants or vitrified by the SSV method. Independently of the presence of sucrose, exposure to vitrification solution for 10 min did reduce the percentages of normal follicles when compared with control. However, the highest rates of normal follicles were attained when tissue was previously exposed to the vitrification solution, with sucrose added or not, for 10 min. Although the SSV is a promising procedure to be applied in ovarian tissue, an optimal vitrification solution for bovine ovarian tissue needs to be developed.

16.
Zygote ; 18(1): 89-92, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19586559

RESUMO

Ovarian cortical fragments from five adult ewes were in vitro cultured for 1, 3 or 5 days in the presence of minimum essential medium either supplemented or not by follicle-stimulating hormone (FSH) (100 ng/ml) or indole-3-acetic acid (IAA) (10, 20, 40 or 100 ng/ml), alone or in combination. After in vitro culture, ovarian fragments were submitted to follicular isolation and viability test was performed using trypan blue. Addition of IAA (10 ng/ml) to a free-FSH medium resulted in the highest percentages of viable follicles, but was progressively deleterious in higher concentrations (20, 40 and 100 ng/ml) if in absence of FSH. Follicular development was observed only when FSH was added to an IAA-free medium. In conclusion, IAA at a concentration of 10 ng/ml increases follicular survival in vitro. However, at high concentrations (20, 40 or 100 ng/ml), this auxin may be deleterious to preantral follicles, the addition of FSH to the medium being necessary.


Assuntos
Hormônio Foliculoestimulante/farmacologia , Ácidos Indolacéticos/farmacologia , Folículo Ovariano/efeitos dos fármacos , Animais , Sobrevivência Celular , Feminino , Folículo Ovariano/citologia , Ovinos , Técnicas de Cultura de Tecidos
17.
Rev. bras. cardiol. invasiva ; 16(3): 301-306, jul.-set. 2008. tab
Artigo em Português | LILACS | ID: lil-503476

RESUMO

Introdução: As complicações vasculares (CV) são uma das principais causas de morbidade e mortalidade em pacientes submetidos a intervenções coronárias percutâneas (ICPs), porém estudos anteriores não refletem a prática atual. Objetivos: Avaliar a incidência de CV e seus preditores, em uma população de pacientes tratada com ICP contemporaneamente. Método: Estudo observacional de corte transversal, com implantes de stents coronários, de janeiro de 2000 a dezembro de 2007. As características clínicas e angiográficas e a evolução intra-hospitalar foram avaliadas e registradas em banco de dados informatizados. Foram excluídos aqueles com óbito hospitalar ou cirurgia cardíaca de urgência. CV foram definidas como sangramento maior, cirurgia vascular ou hematoma > 10 cm. Os dados foram analisados com SPSS 11,0, e as características dos pacientes com e sem CV foram comparadas com teste t de Student e teste do qui-quadrado. Os preditores independentes de CV foram identificados por análise de regressão logística múltipla. Resultados: Total de 4.595 pacientes com 5.485 stents implantados, com média de idade de 60,64 + - 10,65 anos e 32 por cento de mulheres. As ICPs foram realizadas pela via femoral...


Assuntos
Humanos , Feminino , Idoso , Angioplastia Coronária com Balão/efeitos adversos , Cateterismo Cardíaco/efeitos adversos , Doenças Vasculares/etiologia , Aspirina/administração & dosagem , Heparina/administração & dosagem , Fatores de Risco , Ticlopidina/administração & dosagem
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