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1.
Reprod Fertil Dev ; 362024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38894494

RESUMO

Context Altered signalling of androgens, anti-Müllerian hormone or transforming growth factor beta (TGFß) during foetal development have been implicated in the predisposition to polycystic ovary syndrome (PCOS) in later life, aside from its genetic predisposition. In foetal ovarian fibroblasts, TGFß1 has been shown to regulate androgen signalling and seven genes located in loci associated with PCOS. Since PCOS exhibits a myriad of symptoms, it likely involves many different organs. Aims To identify the relationships between TGFß signalling molecules and PCOS candidate genes in different tissues associated with PCOS. Methods Using RNA sequencing data, we examined the expression patterns of TGFß signalling molecules in the human ovary, testis, heart, liver, kidney, brain tissue, and cerebellum from 4 to 20weeks of gestation and postnatally. We also examined the correlations between gene expression of TGFß signalling molecules and PCOS candidate genes. Key results TGFß signalling molecules were dynamically expressed in most tissues prenatally and/or postnatally. FBN3 , a PCOS candidate gene involved in TGFß signalling, was expressed during foetal development in all tissues. The PCOS candidate genes HMGA2, YAP1 , and RAD50 correlated significantly (P TGFBR1 in six out of the seven tissues examined. Conclusions This study suggests that possible crosstalk occurs between genes in loci associated with PCOS and TGFß signalling molecules in multiple tissues, particularly during foetal development. Implications Thus, alteration in TGFß signalling during foetal development could affect many tissues contributing to the multiple phenotypes of PCOS in later life.


Assuntos
Síndrome do Ovário Policístico , Transdução de Sinais , Fator de Crescimento Transformador beta , Humanos , Feminino , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/genética , Adulto , Ovário/metabolismo , Feto/metabolismo , Masculino , Gravidez , Regulação da Expressão Gênica no Desenvolvimento , Testículo/metabolismo , Testículo/embriologia , Fibrilinas
2.
Development ; 151(12)2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38895963

RESUMO

The sixth SY-Stem Symposium, jointly organized by the Research Institute of Molecular Pathology and the Institute of Molecular Biotechnology took place in Vienna in March 2024. Again, aspiring new group leaders were given a stage to present their work and vision of their labs. To round up the excellent program, the scientific organizers included renowned keynote speakers. Here, we provide a summary of the talks covering topics such as early embryogenesis, nervous system development and disease, regeneration and the latest technologies.


Assuntos
Desenvolvimento Embrionário , Humanos , Animais , Regeneração/fisiologia , Células-Tronco/citologia , Sistema Nervoso/embriologia , Diferenciação Celular
3.
Open Biol ; 14(6): 240065, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38896085

RESUMO

The transition from oocyte to embryo requires translation of maternally provided transcripts that in Drosophila is activated by Pan Gu kinase to release a rapid succession of 13 mitotic cycles. Mitotic entry is promoted by several protein kinases that include Greatwall/Mastl, whose Endosulfine substrates antagonize Protein Phosphatase 2A (PP2A), facilitating mitotic Cyclin-dependent kinase 1/Cyclin B kinase activity. Here we show that hyperactive greatwallScant can not only be suppressed by mutants in its Endos substrate but also by mutants in Pan Gu kinase subunits. Conversely, mutants in me31B or trailer hitch, which encode a complex that represses hundreds of maternal mRNAs, enhance greatwallScant . Me31B and Trailer Hitch proteins, known substrates of Pan Gu kinase, copurify with Endos. This echoes findings that budding yeast Dhh1, orthologue of Me31B, associates with Igo1/2, orthologues of Endos and substrates of the Rim15, orthologue of Greatwall. endos-derived mutant embryos show reduced Me31B and elevated transcripts for the mitotic activators Cyclin B, Polo and Twine/Cdc25. Together, our findings demonstrate a previously unappreciated conservation of the Greatwall-Endosulfine pathway in regulating translational repressors and its interactions with the Pan Gu kinase pathway to regulate translation and/or stability of maternal mRNAs upon egg activation.


Assuntos
Proteínas de Drosophila , Regulação da Expressão Gênica no Desenvolvimento , Oócitos , Proteína Fosfatase 2 , Animais , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Oócitos/metabolismo , Oócitos/citologia , Proteína Fosfatase 2/metabolismo , Proteína Fosfatase 2/genética , Biossíntese de Proteínas , Drosophila melanogaster/genética , Drosophila melanogaster/embriologia , Drosophila melanogaster/metabolismo , Mutação , Feminino , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Embrião não Mamífero/metabolismo , Estabilidade de RNA , RNA Mensageiro Estocado/metabolismo , RNA Mensageiro Estocado/genética , RNA Helicases DEAD-box
5.
Biol Open ; 13(6)2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38887972

RESUMO

Regular spatial patterns are ubiquitous forms of organization in nature. In animals, regular patterns can be found from the cellular scale to the tissue scale, and from early stages of development to adulthood. To understand the formation of these patterns, how they assemble and mature, and how they are affected by perturbations, a precise quantitative description of the patterns is essential. However, accessible tools that offer in-depth analysis without the need for computational skills are lacking for biologists. Here, we present PatternJ, a novel toolset to analyze regular one-dimensional patterns precisely and automatically. This toolset, to be used with the popular imaging processing program ImageJ/Fiji, facilitates the extraction of key geometric features within and between pattern repeats in static images and time-lapse series. We validate PatternJ with simulated data and test it on images of sarcomeres from insect muscles and contracting cardiomyocytes, actin rings in neurons, and somites from zebrafish embryos obtained using confocal fluorescence microscopy, STORM, electron microscopy, and brightfield imaging. We show that the toolset delivers subpixel feature extraction reliably even with images of low signal-to-noise ratio. PatternJ's straightforward use and functionalities make it valuable for various scientific fields requiring quantitative one-dimensional pattern analysis, including the sarcomere biology of muscles or the patterning of mammalian axons, speeding up discoveries with the bonus of high reproducibility.


Assuntos
Axônios , Processamento de Imagem Assistida por Computador , Sarcômeros , Somitos , Peixe-Zebra , Animais , Axônios/fisiologia , Processamento de Imagem Assistida por Computador/métodos , Sarcômeros/ultraestrutura , Somitos/embriologia , Software , Algoritmos
6.
Dev Cell ; 59(12): 1487-1488, 2024 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-38889690

RESUMO

In this issue of Developmental Cell, Bolondi et al. systematically assesses neuro-mesodermal progenitor (NMP) dynamics by combining a mouse stem-cell-based embryo model with molecular recording of single cells, shedding light on the dynamics of neural tube and paraxial mesoderm formation during mammalian development.


Assuntos
Mesoderma , Animais , Camundongos , Mesoderma/citologia , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Tubo Neural/citologia , Tubo Neural/embriologia , Diferenciação Celular/fisiologia , Células-Tronco/citologia , Células-Tronco/metabolismo , Padronização Corporal
7.
Nat Commun ; 15(1): 5210, 2024 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-38890321

RESUMO

Cell-fate decisions during mammalian gastrulation are poorly understood outside of rodent embryos. The embryonic disc of pig embryos mirrors humans, making them a useful proxy for studying gastrulation. Here we present a single-cell transcriptomic atlas of pig gastrulation, revealing cell-fate emergence dynamics, as well as conserved and divergent gene programs governing early porcine, primate, and murine development. We highlight heterochronicity in extraembryonic cell-types, despite the broad conservation of cell-type-specific transcriptional programs. We apply these findings in combination with functional investigations, to outline conserved spatial, molecular, and temporal events during definitive endoderm specification. We find early FOXA2 + /TBXT- embryonic disc cells directly form definitive endoderm, contrasting later-emerging FOXA2/TBXT+ node/notochord progenitors. Unlike mesoderm, none of these progenitors undergo epithelial-to-mesenchymal transition. Endoderm/Node fate hinges on balanced WNT and hypoblast-derived NODAL, which is extinguished upon endodermal differentiation. These findings emphasise the interplay between temporal and topological signalling in fate determination during gastrulation.


Assuntos
Embrião de Mamíferos , Endoderma , Gastrulação , Regulação da Expressão Gênica no Desenvolvimento , Análise de Célula Única , Animais , Endoderma/citologia , Endoderma/metabolismo , Endoderma/embriologia , Suínos , Camundongos , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Diferenciação Celular , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/metabolismo , Transcriptoma , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Linhagem da Célula , Proteínas com Domínio T/metabolismo , Proteínas com Domínio T/genética , Transição Epitelial-Mesenquimal/genética
8.
Elife ; 132024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38869942

RESUMO

Movement is a key feature of animal systems, yet its embryonic origins are not fully understood. Here, we investigate the genetic basis underlying the embryonic onset of movement in Drosophila focusing on the role played by small non-coding RNAs (microRNAs, miRNAs). To this end, we first develop a quantitative behavioural pipeline capable of tracking embryonic movement in large populations of fly embryos, and using this system, discover that the Drosophila miRNA miR-2b-1 plays a role in the emergence of movement. Through the combination of spectral analysis of embryonic motor patterns, cell sorting and RNA in situs, genetic reconstitution tests, and neural optical imaging we define that miR-2b-1 influences the emergence of embryonic movement by exerting actions in the developing nervous system. Furthermore, through the combination of bioinformatics coupled to genetic manipulation of miRNA expression and phenocopy tests we identify a previously uncharacterised (but evolutionarily conserved) chloride channel encoding gene - which we term Movement Modulator (Motor) - as a genetic target that mechanistically links miR-2b-1 to the onset of movement. Cell-specific genetic reconstitution of miR-2b-1 expression in a null miRNA mutant background, followed by behavioural assays and target gene analyses, suggest that miR-2b-1 affects the emergence of movement through effects in sensory elements of the embryonic circuitry, rather than in the motor domain. Our work thus reports the first miRNA system capable of regulating embryonic movement, suggesting that other miRNAs are likely to play a role in this key developmental process in Drosophila as well as in other species.


Assuntos
MicroRNAs , Animais , MicroRNAs/metabolismo , MicroRNAs/genética , Drosophila melanogaster/genética , Drosophila melanogaster/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Movimento , Embrião não Mamífero/metabolismo , Drosophila/genética , Drosophila/embriologia , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo
9.
Biol Open ; 13(6)2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38874999

RESUMO

The neural crest (NC) is an embryonic multipotent and transitory population of cells that appears during late gastrulation/early neurulation in the developing embryos of vertebrate organisms. Often called "the fourth germ layer", the NC is characterised by incredible mobility, which allows the NC cells to migrate throughout the whole embryo, giving rise to an astonishing number of different derivatives in the adult organism, such as craniofacial skeleton, adrenal gland, enteric nervous system and melanocytes. Because of these properties, neurocristopathies (NCPs), which is the term used to classify genetic diseases associated with NC developmental defects, are often syndromic and, taken all together, are the most common type of genetic disease. The NEUcrest consortium is an EU funded innovative training network (ITN) that aims to study the NC and NCPs. In March 2024, the early stage researchers (ESRs) in the NEUcrest consortium organised an in-person conference for well-established and early career researchers to discuss new advances in the NC and NCPs field, starting from the induction of the NC, and then moving on to migration and differentiation processes they undergo. The conference focused heavily on NCPs associated with each of these steps. The conference also included events, such as a round table to discuss the future of the NC research, plus a talk by a person living with an NCP. This 3-day conference aimed to bring together the past, present and future of this field to try and unravel the mysteries of this unique cell population.


Assuntos
Crista Neural , Crista Neural/citologia , Crista Neural/embriologia , Humanos , Animais , Diferenciação Celular , Movimento Celular
11.
Proc Natl Acad Sci U S A ; 121(25): e2318838121, 2024 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-38870057

RESUMO

Hertwig's rule states that cells divide along their longest axis, usually driven by forces acting on the mitotic spindle. Here, we show that in contrast to this rule, microtubule-based pulling forces in early Caenorhabditis elegans embryos align the spindle with the short axis of the cell. We combine theory with experiments to reveal that in order to correct this misalignment, inward forces generated by the constricting cytokinetic ring rotate the entire cell until the spindle is aligned with the cell's long axis. Experiments with slightly compressed mouse zygotes indicate that this cytokinetic ring-driven mechanism of ensuring Hertwig's rule is general for cells capable of rotating inside a confining shell, a scenario that applies to early cell divisions of many systems.


Assuntos
Caenorhabditis elegans , Fuso Acromático , Animais , Caenorhabditis elegans/embriologia , Camundongos , Fuso Acromático/metabolismo , Microtúbulos/metabolismo , Citocinese/fisiologia , Rotação , Zigoto/metabolismo , Zigoto/citologia , Zigoto/crescimento & desenvolvimento , Embrião não Mamífero/citologia , Desenvolvimento Embrionário/fisiologia , Modelos Biológicos
12.
J Vis Exp ; (207)2024 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-38856229

RESUMO

The mammalian heart is a complex organ formed during development via highly diverse populations of progenitor cells. The origin, timing of recruitment, and fate of these progenitors are vital for the proper development of this organ. The molecular mechanisms that govern the morphogenesis of the heart are essential for understanding the pathogenesis of congenital heart diseases and embryonic cardiac regeneration. Classical approaches to investigate these mechanisms employed the generation of transgenic mice to assess the function of specific genes during cardiac development. However, mouse transgenesis is a complex, time-consuming process that often cannot be performed to assess the role of specific genes during heart development. To address this, we have developed a protocol for efficient electroporation and culture of mouse embryonic hearts, enabling transient transgenesis to rapidly assess the effect of gain- or loss-of-function of genes involved in cardiac development. Using this methodology, we successfully overexpressed Meis1 in the embryonic heart, with a preference for epicardial cell transfection, demonstrating the capabilities of the technique.


Assuntos
Eletroporação , Técnicas de Transferência de Genes , Coração , Animais , Camundongos , Coração/embriologia , Eletroporação/métodos , Camundongos Transgênicos , Feminino , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Gravidez
13.
Appl Opt ; 63(13): 3712-3724, 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38856558

RESUMO

This study aimed to evaluate the effects of herbicide 2, 4-D-dichlorophenoxy acetic acid on golden apple snail eggs and embryos. Additionally, the study assessed the applicability of optical coherence tomography (OCT), a non-invasive depth cross-sectional microscopic imaging technique, as a novel method, to the best of our knowledge, for studying morphological changes in golden apple snail eggs and embryos, in comparison to the conventional approach of using white light microscopy. The study revealed that the herbicide 2,4-D-dichlorophenoxy acetic acid affected the hatchery rate and morphological changes of the eggs and embryos. The lethal concentration (LC50), representing the concentration of a substance that is expected to cause death in half of the population being studied, of the golden apple eggs and embryos increased with longer exposure time and higher concentrations. The estimated median effective concentration (EC50), which denotes the concentration producing the desired effect in 50% of the exposed golden apple embryos, exhibited a similar trend of change as the LC50. When compared to the microscopic study, it was observed that OCT could be employed to investigate morphological changes of golden apple snail eggs and embryos, enabling evaluation of alterations in both 2D and 3D structures.


Assuntos
Ácido 2,4-Diclorofenoxiacético , Embrião não Mamífero , Herbicidas , Tomografia de Coerência Óptica , Animais , Ácido 2,4-Diclorofenoxiacético/farmacologia , Ácido 2,4-Diclorofenoxiacético/toxicidade , Tomografia de Coerência Óptica/métodos , Herbicidas/farmacologia , Herbicidas/toxicidade , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/embriologia , Caramujos/embriologia , Caramujos/efeitos dos fármacos , Óvulo/efeitos dos fármacos
14.
BMC Genomics ; 25(1): 570, 2024 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-38844864

RESUMO

Compound eyes formation in decapod crustaceans occurs after the nauplius stage. However, the key genes and regulatory mechanisms of compound eye development during crustacean embryonic development have not yet been clarified. In this study, RNA-seq was used to investigate the gene expression profiles of Neocaridina denticulata sinensis from nauplius to zoea stage. Based on RNA-seq data analysis, the phototransduction and insect hormone biosynthesis pathways were enriched, and molting-related neuropeptides were highly expressed. There was strong cell proliferation in the embryo prior to compound eye development. The formation of the visual system and the hormonal regulation of hatching were the dominant biological events during compound eye development. The functional analysis of DEGs across all four developmental stages showed that cuticle formation, muscle growth and the establishment of immune system occurred from nauplius to zoea stage. Key genes related to eye development were discovered, including those involved in the determination and differentiation of the eye field, eye-color formation, and visual signal transduction. In conclusion, the results increase the understanding of the molecular mechanism of eye formation in crustacean embryonic stage.


Assuntos
Olho Composto de Artrópodes , Perfilação da Expressão Gênica , Animais , Olho Composto de Artrópodes/metabolismo , Olho Composto de Artrópodes/crescimento & desenvolvimento , Transcriptoma , Regulação da Expressão Gênica no Desenvolvimento , Decápodes/genética , Decápodes/crescimento & desenvolvimento , Olho/metabolismo , Olho/embriologia , Olho/crescimento & desenvolvimento
15.
J Pineal Res ; 76(5): e12984, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38874070

RESUMO

The antidepressant venlafaxine, a selective serotonin and norepinephrine reuptake inhibitor, is commonly prescribed to treat major depressive disorder and is found at high concentrations in the aquatic environment. Concerns have been raised related to the health of aquatic organisms in response to this nontargeted pharmaceutical exposure. For instance, we previously demonstrated that exposure to venlafaxine perturbs neurodevelopment, leading to behavioural alterations in zebrafish (Danio rerio). We also observed disruption in serotonin expression in the pineal and raphe, regions critical in regulating circadian rhythms, leading us to hypothesize that zygotic exposure to venlafaxine disrupts the circadian locomotor rhythm in larval zebrafish. To test this, we microinjected zebrafish embryos with venlafaxine (1 or 10 ng) and recorded the locomotor activity in 5-day-old larvae over a 24-h period. Venlafaxine deposition reduced larval locomotor activity during the light phase, but not during the dark phase of the diurnal cycle. The melatonin levels were higher in the dark compared to during the light photoperiod and this was not affected by embryonic venlafaxine deposition. Venlafaxine exposure also did not affect the transcript abundance of clock genes, including clock1a, bmal2, cry1a and per2, which showed a clear day/night rhythmicity. A notable finding was that exposure to luzindole, a melatonin receptor antagonist, decreased the locomotor activity in the control group in light, whereas the activity was higher in larvae raised from the venlafaxine-deposited embryos. Overall, zygotic exposure to venlafaxine disrupts the locomotor activity of larval zebrafish fish during the day, demonstrating the capacity of antidepressants to disrupt the circadian rhythms in behaviour. Our results suggest that disruption in melatonin signalling may be playing a role in the venlafaxine impact on circadian behaviour, but further investigation is required to elucidate the possible mechanisms in larval zebrafish.


Assuntos
Ritmo Circadiano , Larva , Locomoção , Cloridrato de Venlafaxina , Peixe-Zebra , Animais , Peixe-Zebra/embriologia , Cloridrato de Venlafaxina/farmacologia , Cloridrato de Venlafaxina/toxicidade , Larva/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Zigoto/efeitos dos fármacos , Zigoto/metabolismo , Atividade Motora/efeitos dos fármacos , Melatonina/farmacologia
16.
BMC Genomics ; 25(1): 553, 2024 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-38831310

RESUMO

Development of the human pancreas requires the precise temporal control of gene expression via epigenetic mechanisms and the binding of key transcription factors. We quantified genome-wide patterns of DNA methylation in human fetal pancreatic samples from donors aged 6 to 21 post-conception weeks. We found dramatic changes in DNA methylation across pancreas development, with > 21% of sites characterized as developmental differentially methylated positions (dDMPs) including many annotated to genes associated with monogenic diabetes. An analysis of DNA methylation in postnatal pancreas tissue showed that the dramatic temporal changes in DNA methylation occurring in the developing pancreas are largely limited to the prenatal period. Significant differences in DNA methylation were observed between males and females at a number of autosomal sites, with a small proportion of sites showing sex-specific DNA methylation trajectories across pancreas development. Pancreas dDMPs were not distributed equally across the genome and were depleted in regulatory domains characterized by open chromatin and the binding of known pancreatic development transcription factors. Finally, we compared our pancreas dDMPs to previous findings from the human brain, identifying evidence for tissue-specific developmental changes in DNA methylation. This study represents the first systematic exploration of DNA methylation patterns during human fetal pancreas development and confirms the prenatal period as a time of major epigenomic plasticity.


Assuntos
Metilação de DNA , Pâncreas , Humanos , Pâncreas/metabolismo , Pâncreas/embriologia , Feminino , Masculino , Regulação da Expressão Gênica no Desenvolvimento , Ilhas de CpG , Epigênese Genética , Genoma Humano , Feto/metabolismo
17.
Brief Bioinform ; 25(4)2024 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-38851297

RESUMO

The development of the human central nervous system initiates in the early embryonic period until long after delivery. It has been shown that several neurological and neuropsychiatric diseases originate from prenatal incidents. Mathematical models offer a direct way to understand neurodevelopmental processes better. Mathematical modelling of neurodevelopment during the embryonic period is challenging in terms of how to 'Approach', how to initiate modelling and how to propose the appropriate equations that fit the underlying dynamics of neurodevelopment during the embryonic period while including the variety of elements that are built-in naturally during the process of neurodevelopment. It is imperative to answer where and how to start modelling; in other words, what is the appropriate 'Approach'? Therefore, one objective of this study was to tackle the mathematical issue broadly from different aspects and approaches. The approaches were divided into three embryonic categories: cell division, neural tube growth and neural plate growth. We concluded that the neural plate growth approach provides a suitable platform for simulation of brain formation/neurodevelopment compared to cell division and neural tube growth. We devised a novel equation and designed algorithms that include geometrical and topological algorithms that could fit most of the necessary elements of the neurodevelopmental process during the embryonic period. Hence, the proposed equations and defined mathematical structure would be a platform to generate an artificial neural network that autonomously grows and develops.


Assuntos
Tubo Neural , Humanos , Tubo Neural/embriologia , Neurogênese , Neurônios/citologia , Algoritmos , Modelos Neurológicos , Animais , Redes Neurais de Computação , Divisão Celular , Desenvolvimento Embrionário , Placa Neural/citologia , Placa Neural/embriologia
18.
Reprod Domest Anim ; 59(6): e14627, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38837827

RESUMO

The efficiency of bovine in vitro embryo production can be significantly improved by splitting embryos at different stages. However, the blastocyst quality of in vitro-produced demi-embryos remains unexplored. The objective of this research was to compare embryo developmental rates and quality of bovine demi-embryos produced by two different strategies: (a) embryo bisection (BSEC) and (b) 2-cell blastomere separation (BSEP). To determine demi-embryos quality, we evaluated total blastocyst cell number and proportion of SOX2+ cells. Additionally, the expression of SOX2, NANOG, OCT4, CDX2, IFNT, BAX and BCL genes and let-7a and miRNA-30c Micro RNAs was analysed. BSEP resulted in improved blastocyst development, higher ICM cells and a significantly higher expression of IFNΤ than demi-embryos produced by BSEC. Let-7a, which is associated with low pregnancy establishment was detected in BSEC, while miRNA-30c expression was observed in all treatments. In conclusion, BSEP of 2-cell embryos is more efficient to improve in vitro bovine embryo development and to produce good quality demi-embryos based on ICM cell number and the expression pattern of the genes explored compared to BSEC.


Assuntos
Blastocisto , Blastômeros , Técnicas de Cultura Embrionária , Desenvolvimento Embrionário , Animais , Bovinos/embriologia , Feminino , Técnicas de Cultura Embrionária/veterinária , Blastômeros/citologia , Fertilização in vitro/veterinária , MicroRNAs/genética , MicroRNAs/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Gravidez
19.
Reprod Domest Anim ; 59(6): e14621, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38828534

RESUMO

Estimating the parturition date in dogs is challenging due to their reproductive peculiarities that. Ultrasonographic examination serves as a tool for studying embryo/foetal biometry and estimating the time of parturition by measuring foetal and extra-foetal structures. However, due to reproductive differences among various dog breeds, such estimates may have a non-significant pattern, representing inaccuracies in the estimated date of birth. This study aimed to monitor pregnant Toy Poodle bitches and establish relationships between ultrasonographically measured foetal and extra-foetal dimensions and the remaining time until parturition. Eighteen pregnant Toy Poodle bitches were subjected to weekly ultrasonographic evaluations and measurements of the inner chorionic cavity diameter, craniocaudal length (CCL), biparietal diameter (BPD), diameter of the deep portion of diencephalo-telencephalic vesicle (DPTV), abdominal diameter, thorax diameter (TXD), placental thickness and the renal diameter (REND). These parameters were retrospectively correlated with the date of parturition and linear regressions were established between gestational measurements and days before parturition (DBP). All analyses were conducted using the Statistical Package for Social Sciences (IBM® SPSS®) program at a 5% significance level. The foetal measurements that showed a high correlation (r) and reliability (R2) with DBP were BPD [(DBP = [15.538 × BPD] - 39.756), r = .97 and R2 = .93], TXD [(DBP = [8.933 × TXD] - 32.487), r = .94 and R2 = .89], DPTV [(DBP = [34.580 × DPTV] - 39.403), r = .93 and R2 = .86] and REND [(DBP = [13.735 × REND] - 28.937), r = .91 and R2 = .82]. This statistically validates the application of these specific formulas to estimate the parturition date in Toy Poodle bitches.


Assuntos
Parto , Ultrassonografia Pré-Natal , Animais , Feminino , Gravidez , Cães/embriologia , Ultrassonografia Pré-Natal/veterinária , Biometria , Feto/anatomia & histologia , Feto/diagnóstico por imagem , Estudos Retrospectivos , Placenta/diagnóstico por imagem , Placenta/anatomia & histologia , Embrião de Mamíferos/fisiologia , Idade Gestacional
20.
J Vis Exp ; (207)2024 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-38829132

RESUMO

Microglia are highly dynamic cells and their migration and colonization of the brain parenchyma is a crucial step for proper brain development and function. Externally developing zebrafish embryos possess optical transparency, which along with well-characterized transgenic reporter lines that fluorescently label microglia, make zebrafish an ideal vertebrate model for such studies. In this paper, we take advantage of the unique features of the zebrafish model to visualize the dynamics of microglia cells in vivo and under physiological conditions. We use confocal microscopy to record a timelapse of microglia cells in the optic tectum of the zebrafish embryo and then, extract tracking data using the IMARIS 10.0 software to obtain the cells' migration path, mean speed, and distribution in the optic tectum at different developmental stages. This protocol can be a useful tool to elucidate the physiological significance of microglia behavior in various contexts, contributing to a deeper characterization of these highly motile cells.


Assuntos
Microglia , Microscopia Confocal , Peixe-Zebra , Animais , Peixe-Zebra/embriologia , Microglia/citologia , Microscopia Confocal/métodos , Movimento Celular/fisiologia , Colículos Superiores/citologia , Colículos Superiores/fisiologia , Embrião não Mamífero/citologia
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