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1.
Front Vet Sci ; 10: 1254940, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37808114

RESUMO

Thoroughly analyzing the sperm and exploring the information obtained using artificial intelligence (AI) could be the key to improving fertility estimation. Artificial neural networks have already been applied to calculate zootechnical indices in animals and predict fertility in humans. This method of estimating the results of reproductive biotechnologies, such as in vitro embryo production (IVEP) in cattle, could be valuable for livestock production. This study was developed to model IVEP estimates in Senepol animals based on various sperm attributes, through retrospective data from 290 IVEP routines performed using 38 commercial doses of semen from Senepol bulls. All sperm samples that had undergone the same procedure during sperm selection for in vitro fertilization were evaluated using a computer-assisted sperm analysis (CASA) system to define sperm subpopulations. Sperm morphology was also analyzed in a wet preparation, and the integrity of the plasma and acrosomal membranes, mitochondrial potential, oxidative status, and chromatin resistance were evaluated using flow cytometry. A previous study identified three sperm subpopulations in such samples and the information used in tandem with other sperm quality variables to perform an AI analysis. AI analysis generated models that estimated IVEP based on the season, donor, percentage of viable oocytes, and 18 other sperm predictor variables. The accuracy of the results obtained for the three best AI models for predicting the IVEP was 90.7, 75.3, and 79.6%, respectively. Therefore, applying this AI technique would enable the estimation of high or low embryo production for individual bulls based on the sperm analysis information.

2.
Int J Mol Sci ; 24(18)2023 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-37762363

RESUMO

During embryo development, the endoplasmic reticulum (ER) acts as an important site for protein biosynthesis; however, in vitro culture (IVC) can negatively affect ER homeostasis. Therefore, the aim of our study was to evaluate the effects of the supplementation of tauroursodeoxycholic acid (TUDCA), an ER stress inhibitor, in the IVC of bovine embryos. Two experiments were carried out: Exp. 1: an evaluation of blastocyst rate, hatching kinetics, and gene expression of hatched embryos after being treated with different concentrations of TUDCA (50, 200, or 1000 µM) in the IVC; Exp. 2: an evaluation of the re-expansion, hatching, and gene expression of hatched embryos previously treated with 200 µM of TUDCA at IVC and submitted to vitrification. There was no increase in the blastocyst and hatched blastocyst rates treated with TUDCA in the IVC. However, embryos submitted to vitrification after treatment with 200 µM of TUDCA underwent an increased hatching rate post-warming together with a down-regulation in the expression of ER stress-related genes and the accumulation of lipids. In conclusion, this work showed that the addition of TUDCA during in vitro culture can improve the cryotolerance of the bovine blastocyst through the putative modulation of ER and oxidative stress.


Assuntos
Retículo Endoplasmático , Ácido Tauroquenodesoxicólico , Bovinos , Animais , Ácido Tauroquenodesoxicólico/farmacologia , Suplementos Nutricionais
3.
Theriogenology ; 204: 1-7, 2023 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-37030172

RESUMO

Supplementation of culture media with IGF-1 during in vitro culture of embryos has had controversial results over the years. In the present study, we show that differences previously observed in response to IGF addition might be related to intrinsic heterogeneity of the embryos. In other words, the effects exerted by IGF-1 are dependent on the characteristics of the embryos and their ability to modulate metabolism and overcome stressful conditions, such as the ones found in a non-optimized in vitro culture system. To test this hypothesis, in vitro produced bovine embryos with distinct morphokinetics (fast- and slow-cleavage) were submitted to treatment with IGF-1 and then evaluated for embryo production rates, total cell number, gene expression and lipid profile. Our results show that remarkable differences were found when fast and slow embryos treated with IGF-1 were compared. Fast embryos respond by upregulating genes related to mitochondrial function, stress response, and lipid metabolism, whereas slow embryos presented lower mitochondrial efficiency and lipid accumulation. We conclude that indeed the treatment with IGF-1 selectively affects embryonic metabolism according to early morphokinetics phenotypes, and this information is relevant for decision-making in the design of more appropriate in vitro culture systems.


Assuntos
Desenvolvimento Embrionário , Fator de Crescimento Insulin-Like I , Animais , Bovinos , Fator de Crescimento Insulin-Like I/metabolismo , Desenvolvimento Embrionário/fisiologia , Blastocisto/fisiologia , Embrião de Mamíferos , Lipídeos , Fertilização in vitro/métodos , Fertilização in vitro/veterinária
4.
Front Cell Dev Biol ; 10: 938709, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36187479

RESUMO

Several opportunities for embryo development, stem cell maintenance, cell fate, and differentiation have emerged using induced pluripotent stem cells (iPSCs). However, the difficulty in comparing bovine iPSCs (biPSCs) with embryonic stem cells (ESCs) was a challenge for many years. Here, we reprogrammed fetal fibroblasts by transient expression of the four transcription factors (Oct4, Sox2, Klf4, and c-Myc, collectively termed "OSKM" factors) and cultured in iPSC medium, supplemented with bFGF, bFGF2i, leukemia inhibitory factor (LIF), or LIF2i, and then compared these biPSC lines with bESC to evaluate the pluripotent state. biPSC lines were generated in all experimental groups. Particularly, reprogrammed cells treated with bFGF were more efficient in promoting the acquisition of pluripotency. However, LIF2i treatment did not promote continuous self-renewal. biPSCs (line 2) labeled with GFP were injected into early embryos (day 4.5) to assess the potential to contribute to chimeric blastocysts. The biPSC lines show a pluripotency state and are differentiated into three embryonic layers. Moreover, biPSCs and bESCs labeled with GFP were able to contribute to chimeric blastocysts. Additionally, biPSCs have shown promising potential for contributing to chimeric blastocysts and for future studies.

5.
Sci Rep ; 12(1): 11228, 2022 07 04.
Artigo em Inglês | MEDLINE | ID: mdl-35787650

RESUMO

Giant unilamellar vesicles (GUVs) are composed of lipophilic layers and are sensitive to the action of reactive oxygen species (ROS). The use of GUVs as microcarriers of biological macromolecules is particularly interesting since ROS produced by gametes or embryos during in vitro culture can induce the opening of pores in the membrane of these vesicles and cause the release of their content. This study investigated the behavior of GUVs [composed of 2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl)] in co-culture with in vitro produced bovine embryos, as well as their embryotoxicity and effectiveness as cysteine carriers in culture medium. Embryonic developmental rates were unaffected, demonstrating the absence of toxicity of GUVs co-cultured with the embryos. No increase of intracellular ROS levels was observed in the embryos co-cultured with GUVs, indicating that the higher lipid content of the culture environment resulting from the lipid composition of the GUV membrane itself did not increase oxidative stress. Variations in the diameter and number of GUVs demonstrated their sensitivity to ROS produced by embryos cultured under conditions that generate oxidative stress. Encapsulation of cysteine in GUVs was found to be more effective in controlling the production of ROS in embryonic cells than direct dilution of this antioxidant in the medium. In conclusion, the use of GUVs in in vitro culture was found to be safe since these vesicles did not promote toxic effects nor did they increase intracellular ROS concentrations in the embryos. GUVs were sensitive to oxidative stress, which resulted in structural changes in response to the action of ROS. The possible slow release of cysteine into the culture medium by GUV rupture would therefore favor the gradual supply of cysteine, prolonging its presence in the medium. Thus, the main implication of the use of GUVs as cysteine microcarriers is the greater effectiveness in preventing the intracytoplasmic increase of ROS in in vitro produced bovine embryos.


Assuntos
Antioxidantes , Lipossomas Unilamelares , Animais , Antioxidantes/farmacologia , Bovinos , Cisteína , Espécies Reativas de Oxigênio , Lipossomas Unilamelares/química
6.
Cells ; 10(6)2021 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-34204517

RESUMO

The event of cellular reprogramming into pluripotency is influenced by several factors, such as in vitro culture conditions (e.g., culture medium and oxygen concentration). Herein, bovine iPSCs (biPSCs) were generated in different levels of oxygen tension (5% or 20% of oxygen) and supplementation (bFGF or bFGF + LIF + 2i-bFL2i) to evaluate the efficiency of pluripotency induction and maintenance in vitro. Initial reprogramming was observed in all groups and bFL2i supplementation initially resulted in a superior number of colonies. However, bFL2i supplementation in low oxygen led to a loss of self-renewal and pluripotency maintenance. All clonal lines were positive for alkaline phosphatase; they expressed endogenous pluripotency-related genes SOX2, OCT4 and STELLA. However, expression was decreased throughout the passages without the influence of oxygen tension. GLUT1 and GLUT3 were upregulated by low oxygen. The biPSCs were immunofluorescence-positive stained for OCT4 and SOX2 and they formed embryoid bodies which differentiated in ectoderm and mesoderm (all groups), as well as endoderm (one line from bFL2i in high oxygen). Our study is the first to compare high and low oxygen environments during and after induced reprogramming in cattle. In our conditions, a low oxygen environment did not favor the pluripotency maintenance of biPSCs.


Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco Pluripotentes Induzidas , Oxigênio/farmacologia , Animais , Bovinos , Reprogramação Celular/efeitos dos fármacos
7.
Front Cell Dev Biol ; 9: 623852, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33681203

RESUMO

In several species, oocyte and embryo competence are improved by the addition of endoplasmic reticulum (ER) stress inhibitors to in vitro maturation (IVM) medium and/or in vitro culture (IVC) medium. This study aimed to evaluate the effects of three concentrations of tauroursodeoxycholic acid (TUDCA; 50, 200, and 1,000 µM), a chemical chaperone for relieving ER stress, during IVM of bovine cumulus-oocyte complexes (COCs) for 24 h. Treated oocytes were analyzed for nuclear maturation, reactive oxygen species (ROS) production, mitochondrial activity, and abundance of target transcripts. In addition, the number of pronuclei in oocytes was evaluated after 18-20 h of insemination, and the rates of blastocyst and hatched blastocyst formation were evaluated after 7 and 8/9 days of culture, respectively. We further evaluated the transcript abundance of embryonic quality markers. Our findings showed that supplementation of IVM medium with 200 µM of TUDCA decreased ROS production and increased abundance of transcripts related to antioxidant activity in oocytes (CAT, GPX1, and HMOX1) and embryos (GPX1 and PRDX3). Interestingly, high concentration of TUDCA (1,000 µM) was toxic to oocytes, reducing the nuclear maturation rate, decreasing mitochondrial activity, and increasing the abundance of ER stress (HSPA5) and cellular apoptosis (CASP3 and CD40) related transcripts. The results of this study suggest that treatment with 200 µM of TUDCA is associated with a greater resistance to oxidative stress and indirectly with ER stress relief in bovine oocytes.

8.
Sci Rep ; 10(1): 19557, 2020 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-33177637

RESUMO

We evaluated the effect of the antral follicle count (AFC) on ovarian follicular dynamics, pregnancy rates, progesterone concentrations, and transcriptional patterns of genes in Nelore cattle (Bos taurus indicus) after a timed artificial insemination (TAI) programme. Cows were separated based on the AFC, and those with a high AFC showed a larger (P < 0.0001) ovarian diameter and area than those with a very low AFC. Females with a very low AFC exhibited a larger (P < 0.01) diameter of the dominant follicle at TAI (13.6 ± 0.3 vs. 12.2 ± 0.4 mm) and a tendency (P = 0.06) to have different serum progesterone concentrations (2.9 ± 0.3 vs. 2.1 ± 0.3 ng/mL; on day 18, considering day 0 as the beginning of the synchronization protocol) than those with a high AFC. The pregnancy rate was higher (P ≤ 0.05) in animals with a very low (57.9%) and low (53.1%) AFC than in those with a high AFC (45.2%). The expression of genes related to intercellular communication, meiotic control, epigenetic modulation, cell division, follicular growth, cell maintenance, steroidogenesis and cellular stress response was assessed on day 5. In females with a low AFC, 8 and 21 genes in oocytes and cumulus cells, respectively, were upregulated (P < 0.05), while 3 and 6 genes in oocytes and cumulus cells, respectively, were downregulated. The results described here will help elucidate the differences in ovarian physiology and the reproductive success of Bos indicus females with a low or high AFC.


Assuntos
Folículo Ovariano/fisiologia , Taxa de Gravidez , Progesterona/sangue , Transcriptoma , Animais , Bovinos , Células do Cúmulo/citologia , Feminino , Inseminação Artificial/veterinária , Oócitos/citologia , Folículo Ovariano/citologia , Ovário/citologia , Ovário/fisiologia , Gravidez
9.
Int J Mol Sci ; 21(18)2020 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-32962179

RESUMO

In many cell types, epigenetic changes are partially regulated by the availability of metabolites involved in the activity of chromatin-modifying enzymes. Even so, the association between metabolism and the typical epigenetic reprogramming that occurs during preimplantation embryo development remains poorly understood. In this work, we explore the link between energy metabolism, more specifically the tricarboxylic acid cycle (TCA), and epigenetic regulation in bovine preimplantation embryos. Using a morphokinetics model of embryonic development (fast- and slow-developing embryos), we show that DNA methylation (5mC) and hydroxymethylation (5hmC) are dynamically regulated and altered by the speed of the first cleavages. More specifically, slow-developing embryos fail to perform the typical reprogramming that is necessary to ensure the generation of blastocysts with higher ability to establish specific cell lineages. Transcriptome analysis revealed that such differences were mainly associated with enzymes involved in the TCA cycle rather than specific writers/erasers of DNA methylation marks. This relationship was later confirmed by disturbing the embryonic metabolism through changes in α-ketoglutarate or succinate availability in culture media. This was sufficient to interfere with the DNA methylation dynamics despite the fact that blastocyst rates and total cell number were not quite affected. These results provide the first evidence of a relationship between epigenetic reprogramming and energy metabolism in bovine embryos. Likewise, levels of metabolites in culture media may be crucial for precise epigenetic reprogramming, with possible further consequences in the molecular control and differentiation of cells.


Assuntos
Blastocisto/enzimologia , Blastocisto/metabolismo , Ciclo do Ácido Cítrico , Metilação de DNA , Animais , Blastocisto/citologia , Bovinos , Meios de Cultura/metabolismo , Desenvolvimento Embrionário/genética , Metabolismo Energético , Epigênese Genética , Feminino , Perfilação da Expressão Gênica , Ontologia Genética , Ácidos Cetoglutáricos/metabolismo , Gravidez , Ácido Succínico/metabolismo
10.
Sci Rep ; 10(1): 15672, 2020 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-32973241

RESUMO

Previous studies have discussed the importance of an optimal range of metabolic activity during preimplantation development. To avoid factors than can trigger an undesirable trajectory, it is important to learn how nutrients and metabolites interact to help launching the correct developmental program of the embryo, and how much the in vitro culture system can impair this process. Here, using the bovine model, we describe a factorial experimental design used to investigate the biochemical and molecular signature of embryos in response to different combinations of morphological features-i.e. speed of development-and external stimuli during in vitro culture-i.e. different oxygen tensions and glucose supplementation. Our analyses demonstrate that the embryos present heterogeneous metabolic responses depending on early morphological phenotypes and the composition of their surroundings. However, despite the contribution of each single stimulus for the embryo phenotype, oxygen tension is determinant for such differences. The lower oxygen environment boosts the metabolism of embryos with faster kinetics, in particular those cultured in lower glucose concentrations.


Assuntos
Adaptação Fisiológica , Técnicas de Cultura Embrionária , Embrião de Mamíferos/fisiologia , Meio Ambiente , Adaptação Fisiológica/efeitos dos fármacos , Animais , Bovinos , Relação Dose-Resposta a Droga , Embrião de Mamíferos/efeitos dos fármacos , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Glucose/farmacologia , Oxigênio/metabolismo
11.
J Assist Reprod Genet ; 37(10): 2359-2376, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32654105

RESUMO

Over the past years, the assisted reproductive technologies (ARTs) have been accompanied by constant innovations. For instance, intracytoplasmic sperm injection (ICSI), time-lapse monitoring of the embryonic morphokinetics, and PGS are innovative techniques that increased the success of the ART. In the same trend, the use of artificial intelligence (AI) techniques is being intensively researched whether in the embryo or spermatozoa selection. Despite several studies already published, the use of AI within assisted reproduction clinics is not yet a reality. This is largely due to the different AI techniques that are being proposed to be used in the daily routine of the clinics, which causes some uncertainty in their use. To shed light on this complex scenario, this review briefly describes some of the most frequently used AI algorithms, their functionalities, and their potential use. Several databases were analyzed in search of articles where applied artificial intelligence algorithms were used on reproductive data. Our focus was on the classification of embryonic cells and semen samples. Of a total of 124 articles analyzed, 32 were selected for this review. From the proposed algorithms, most have achieved a satisfactory precision, demonstrating the potential of a wide range of AI techniques. However, the evaluation of these studies suggests the need for more standardized research to validate the proposed models and their algorithms. Routine use of AI in assisted reproduction clinics is just a matter of time. However, the choice of AI technique to be used is supported by a better understanding of the principles subjacent to each technique, that is, its robustness, pros, and cons. We provide some current (although incipient) and potential uses of AI on the clinic routine, discussing how accurate and friendly it could be. Finally, we propose some standards for AI research on the selection of the embryo to be transferred and other future hints. For us, the imminence of its use is evident, providing a revolutionary milestone that will impact the ART.


Assuntos
Inteligência Artificial/tendências , Reprodução/genética , Técnicas de Reprodução Assistida/tendências , Injeções de Esperma Intracitoplásmicas/tendências , Algoritmos , Feminino , Fertilização in vitro/tendências , Humanos , Masculino , Reprodução/fisiologia , Injeções de Esperma Intracitoplásmicas/métodos , Espermatozoides/crescimento & desenvolvimento
12.
JBRA Assist Reprod ; 24(4): 470-479, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32293823

RESUMO

Based on growing demand for assisted reproduction technology, improved predictive models are required to optimize in vitro fertilization/intracytoplasmatic sperm injection strategies, prioritizing single embryo transfer. There are still several obstacles to overcome for the purpose of improving assisted reproductive success, such as intra- and inter-observer subjectivity in embryonic selection, high occurrence of multiple pregnancies, maternal and neonatal complications. Here, we compare studies that used several variables that impact the success of assisted reproduction, such as blastocyst morphology and morphokinetic aspects of embryo development as well as characteristics of the patients submitted to assisted reproduction, in order to predict embryo quality, implantation or live birth. Thereby, we emphasize the proposal of an artificial intelligence-based platform for a more objective method to predict live birth.


Assuntos
Inteligência Artificial , Desenvolvimento Embrionário/fisiologia , Nascido Vivo , Técnicas de Reprodução Assistida , Feminino , Fertilização in vitro , Humanos , Gravidez , Taxa de Gravidez , Prognóstico
13.
PLoS One ; 14(8): e0220731, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31381602

RESUMO

In this study, we evaluated the modulation effect of long-chain Acyl-CoA synthetase during early embryo development. Bovine embryos were cultured in four groups: positive modulation (ACS+) with GW3965 hydrochloride, negative modulation (ACS-) with Triacsin C, association of both modulators (ACS±), and control. Embryo development rates were not altered (P>0.05) by treatments. Embryonic cytoplasmic lipid content increased in ACS+ but reduced in ACS- compared to the control (P < 0.05), whereas the membrane phospholipids profile was not altered by treatments. The total number of blastomeres did not differ (P > 0.05) between groups; however, an increased apoptotic cells percentage was found in ACS- compared to control. Twenty-four hours after warming, ACS+ and control grade I embryos presented the best hatching rates, whereas the ACS+ group equaled the hatching rates between their embryos of grades I, II and III 48 hours after warming. The relative abundance of transcripts for genes associated with lipid metabolism (ACSL3, ACSL6, ACAT1, SCD, and AUH), heatshock (HSP90AA1 and HSF1), oxidative stress (GPX4), and angiogenesis (VEGF), among other important genes for embryo development were affected by at least one of the treatments. The treatments were effective in modulating the level of transcripts for ACSL3 and the cytoplasmic lipid content. The ACS- was not effective in increasing embryonic cryosurvival, whereas ACS+ restored survival rates after vitrification of embryos with low quality, making them equivalent to embryos of excellent quality.


Assuntos
Bovinos/embriologia , Coenzima A Ligases/metabolismo , Metabolismo dos Lipídeos , Animais , Bovinos/genética , Bovinos/metabolismo , Criopreservação/métodos , Técnicas de Cultura Embrionária/métodos , Desenvolvimento Embrionário , Feminino , Fertilização in vitro/métodos , Gotículas Lipídicas/metabolismo , Fosfolipídeos/metabolismo , Transcriptoma , Vitrificação
14.
Mol Reprod Dev ; 86(11): 1639-1651, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31389116

RESUMO

Studies have shown that the use of equine chorionic gonadotropin (eCG), which binds both follicle stimulating hormone (FSH) and luteinizing hormone (LH) receptors, could modify the female reproductive tract. We, thus, aimed to quantify the messenger RNA (mRNA) abundance of genes related to cumulus-oocyte complexes (COCs) and embryo quality in Nelore cows (Bos taurus indicus) submitted to ovarian superstimulation using only FSH (FSH group; n = 10) or replacement of the last two doses of FSH by eCG (FSH/eCG group; n = 10). All animals were slaughtered and the ovarian antral follicles from both groups (10-14 mm in diameter) were aspirated for cumulus, oocyte and in vitro embryo production gene expression analysis. The relative mRNA abundance of 96 genes related to COCs development and embryo quality was measured by RT-qPCR. We found that oocytes are more affected by eCG use and that 35 genes involved in lipid metabolism, oxidative stress, transcriptional control, and cellular development were upregulated in the FSH/eCG group. In blastocysts, lipid metabolism seems to be the main pathway regulated by eCG use. We suggest that these multiple effects could be due to the ability of eCG to bind LHR and FSHR, which could activate multiple signal transduction pathways in the superstimulated ovary, further impacting the transcriptional profile of COCs and blastocysts.


Assuntos
Blastocisto/metabolismo , Gonadotropina Coriônica/farmacologia , Células do Cúmulo/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Oócitos/metabolismo , Superovulação/metabolismo , Transcriptoma/efeitos dos fármacos , Animais , Blastocisto/citologia , Bovinos , Células do Cúmulo/citologia , Feminino , Perfilação da Expressão Gênica , Cavalos , Oócitos/citologia
15.
Reprod Fertil Dev ; 31(3): 451-461, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30301510

RESUMO

Follicular fluid composition and the transcription pattern of granulosa cells were analysed to better comprehend associations between embryo development and morphokinetics. Bovine follicles were punctured and their respective follicular fluid and granulosa cells were collected. Cumulus-oocyte complexes derived from these follicles were matured and fertilised invitro. Embryo morphology and kinetics were evaluated at 40h after insemination, when embryos were classified as fast (FCL, four or more cells), slow (SCL, 2-3 cells) or non-cleaved (NCL). Their development was followed until the blastocyst stage. Glucose, pyruvate, cholesterol and oestradiol were quantified in the follicular fluid and the transcription pattern of 96 target genes was evaluated in granulosa cells by large-scale quantitative reverse transcription polymerase chain reaction. Follicular fluid from the blastocyst group had increased levels of glucose, total cholesterol and pyruvate compared to the non-blastocyst group, whereas higher levels of oestradiol were observed in the follicular fluid of embryos and blastocysts with fast cleavage. The transcriptional pattern revealed altered metabolic pathways between groups, such as lipid metabolism, cellular stress and cell signalling. In conclusion, both follicular fluid and granulosa cells are associated with the possibility of identifying follicles that may generate embryos with high potential to properly develop to the blastocyst stage.


Assuntos
Desenvolvimento Embrionário/fisiologia , Líquido Folicular/metabolismo , Folículo Ovariano/metabolismo , Animais , Bovinos , Colesterol/metabolismo , Estradiol/metabolismo , Feminino , Glucose/metabolismo , Células da Granulosa/metabolismo , Cinética , Ácido Pirúvico/metabolismo
16.
Epigenetics Chromatin ; 11(1): 1, 2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29310712

RESUMO

BACKGROUND: The timing of the first cell divisions may predict the developmental potential of an embryo, including its ability to establish pregnancy. Besides differences related to metabolism, stress, and survival, embryos with different speeds of development present distinct patterns of gene expression, mainly related to energy and lipid metabolism. As gene expression is regulated by epigenetic factors, and that includes DNA methylation patterns, in this study we compared the global DNA methylation profile of embryos with different kinetics of development in order to identify general pathways and regions that are most influenced by this phenotype. For this purpose, bovine embryos were in vitro produced using sexed semen (female), classified as fast (four or more cells) or slow (two cells) at 40 hpi and cultured until blastocyst stage, when they were analyzed. RESULTS: Genome-wide DNA methylation analysis identified 11,584 differently methylated regions (DMRs) (7976 hypermethylated regions in fast and 3608 hypermethylated regions in slow embryos). Fast embryos presented more regions classified as hypermethylated distributed throughout the genome, as in introns, exons, promoters, and repeat elements while in slow embryos, hypermethylated regions were more present in CpG islands. DMRs were clustered by means of biological processes, and the most affected pathways were related to cell survival/differentiation and energy/lipid metabolism. Transcripts profiles from DM genes connected with these pathways were also assessed, and the most part disclosed changes in relative quantitation. CONCLUSION: The kinetics of the first cleavages influences the DNA methylation and expression profiles of genes related to metabolism and differentiation pathways and may affect embryo viability.


Assuntos
Blastocisto/citologia , Metilação de DNA , Desenvolvimento Embrionário , Animais , Blastocisto/química , Bovinos , Ilhas de CpG , Epigênese Genética , Feminino , Cinética
17.
Oxid Med Cell Longev ; 2017: 1502489, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29209446

RESUMO

High oxygen levels during in vitro culture (IVC) can induce oxidative stress through accumulation of reactive oxygen species (ROS), negatively affecting embryo development. This study evaluated the effect of different O2 tensions during IVC on bovine blastocyst development and transcriptional status, considering transcription factors that play an essential role during early embryo development. For this purpose, embryos were produced in vitro by conventional protocols and cultured in two different oxygen tensions, physiological (5%) and atmospheric (20%). Expanded blastocysts were subjected to transcript quantitation analysis by RT-qPCR with Biomark™ HD System (Fluidigm, US), using 67 TaqMan assays specific for Bos taurus. Differences were observed in genes related to oxidation-reduction processes, DNA-dependent transcription factors, and factors related to important functional pathways for embryo development. Blastocyst rate was higher in the 5% O2 group and the number of cells was assessed, with the 5% O2 group having a higher number of cells. ROS concentration was evaluated, with a higher ROS presence in the 20% O2 group. Taken together, these results allow us to conclude that IVC of embryos at atmospheric O2 tension affects the expression of important transcription factors involved in multiple cell biology pathways that can affect embryo development, quality, and viability.


Assuntos
Embrião de Mamíferos/metabolismo , Estresse Oxidativo , Fatores de Transcrição/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Blastocisto/citologia , Blastocisto/metabolismo , Fator de Transcrição CDX2/metabolismo , Bovinos , Embrião de Mamíferos/citologia , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Fertilização in vitro , Microscopia de Fluorescência , Oócitos/citologia , Fatores de Transcrição Otx/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Oxigênio/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transcriptoma/efeitos dos fármacos
18.
Sci Data ; 4: 170192, 2017 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-29257125

RESUMO

There is currently no objective, real-time and non-invasive method for evaluating the quality of mammalian embryos. In this study, we processed images of in vitro produced bovine blastocysts to obtain a deeper comprehension of the embryonic morphological aspects that are related to the standard evaluation of blastocysts. Information was extracted from 482 digital images of blastocysts. The resulting imaging data were individually evaluated by three experienced embryologists who graded their quality. To avoid evaluation bias, each image was related to the modal value of the evaluations. Automated image processing produced 36 quantitative variables for each image. The images, the modal and individual quality grades, and the variables extracted could potentially be used in the development of artificial intelligence techniques (e.g., evolutionary algorithms and artificial neural networks), multivariate modelling and the study of defined structures of the whole blastocyst.


Assuntos
Blastocisto , Animais , Bovinos , Feminino , Processamento de Imagem Assistida por Computador , Gravidez
19.
Anim. Reprod. ; 14(3): 635-644, 017t.2017t.2017. graf, tab
Artigo em Inglês | VETINDEX | ID: vti-16165

RESUMO

The association of OPU-IVEP is an important instrument to drive genetic progress. In vitro embryo production (IVEP) has remarkably expanded in the last decade compared to in vivo embryo production. Because of the high repeatability of oocyte retrieval within oocyte-donors, studies exploring the relationship between the number of oocytes recovered per OPU section with IVEP efficiency, as well as with field fertility (pregnancy results following embryo transfer; P/ET) are extremely important to guide cow-donor selection and optimize field reproduction efficiency and the herds genetic gain. Based on this rationale, our group conducted a retrospective analysis of a large database comprising IVEP records from several cattle breeds, including Bos indicus and Bos taurus for either beef or dairy purposes. A total of 205,140 oocytes recovered from 7,906 OPU procedures of 6,902 donors (5,227 beef and 1,675 dairy) of Brazilian farms were analyzed. Beef breeds analyzed were Nelore (Bos indicus) and Senepol (Bos taurus) and dairy breeds Were Gyr (Bos indicus) and Holstein (Bos taurus). According to our analysis, the IVEP in beef cattle had a great improvement throughout the last years, with a remarkable increase in numbers of pregnancies per OPU compared to late 90s (averaging only 1 pregnancy per OPU in 1998 vs 2,4 in 2014). As for the distribution of oocytes retrieved, both Bos indicus beef (Nelore = 27.2) and dairy (Gyr = 23.8) breeds seem to yield greater average numbers of oocytes per OPU compared to Bos taurus (Senepol = 21.8; Holstein = 19.3).(AU)


Assuntos
Animais , Feminino , Gravidez , Bovinos , Bovinos/embriologia , Taxa de Gravidez , Fase Folicular , Oócitos/crescimento & desenvolvimento , Transferência Embrionária/veterinária
20.
Anim. Reprod. (Online) ; 14(3): 635-644, July.-Sept.2017. graf, tab
Artigo em Inglês | VETINDEX | ID: biblio-1461294

RESUMO

The association of OPU-IVEP is an important instrument to drive genetic progress. In vitro embryo production (IVEP) has remarkably expanded in the last decade compared to in vivo embryo production. Because of the high repeatability of oocyte retrieval within oocyte-donors, studies exploring the relationship between the number of oocytes recovered per OPU section with IVEP efficiency, as well as with field fertility (pregnancy results following embryo transfer; P/ET) are extremely important to guide cow-donor selection and optimize field reproduction efficiency and the herd’s genetic gain. Based on this rationale, our group conducted a retrospective analysis of a large database comprising IVEP records from several cattle breeds, including Bos indicus and Bos taurus for either beef or dairy purposes. A total of 205,140 oocytes recovered from 7,906 OPU procedures of 6,902 donors (5,227 beef and 1,675 dairy) of Brazilian farms were analyzed. Beef breeds analyzed were Nelore (Bos indicus) and Senepol (Bos taurus) and dairy breeds Were Gyr (Bos indicus) and Holstein (Bos taurus). According to our analysis, the IVEP in beef cattle had a great improvement throughout the last years, with a remarkable increase in numbers of pregnancies per OPU compared to late 90’s (averaging only 1 pregnancy per OPU in 1998 vs 2,4 in 2014). As for the distribution of oocytes retrieved, both Bos indicus beef (Nelore = 27.2) and dairy (Gyr = 23.8) breeds seem to yield greater average numbers of oocytes per OPU compared to Bos taurus (Senepol = 21.8; Holstein = 19.3).


Assuntos
Feminino , Animais , Gravidez , Bovinos , Bovinos/embriologia , Fase Folicular , Oócitos/crescimento & desenvolvimento , Taxa de Gravidez , Transferência Embrionária/veterinária
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