Science.gov

Sample records for embryos

  1. Embryos, microscopes, and society.

    PubMed

    Maienschein, Jane

    2016-06-01

    Embryos have different meanings for different people and in different contexts. Seen under the microscope, the biological embryo starts out as one cell and then becomes a bunch of cells. Gradually these divide and differentiate to make up the embryo, which in humans becomes a fetus at eight weeks, and then eventually a baby. At least, that happens in those cases that carry through normally and successfully. Yet a popular public perception imagines the embryo as already a little person in the very earliest stages of development, as if it were predictably to become an adult. In actuality, cells can combine, pull apart, and recombine in a variety of ways and still produce embryos, whereas most embryos never develop into adults at all. Biological embryos and popular imaginations of embryos diverge. This paper looks at some of the historical reasons for and social implications of that divergence.

  2. Potentiality and human embryos.

    PubMed

    Lizza, John P

    2007-09-01

    Consideration of the potentiality of human embryos to develop characteristics of personhood, such as intellect and will, has figured prominently in arguments against abortion and the use of human embryos for research. In particular, such consideration was the basis for the call of the US President's Council on Bioethics for a moratorium on stem cell research on human embryos. In this paper, I critique the concept of potentiality invoked by the Council and offer an alternative account. In contrast to the Council's view that an embryo's potentiality is determined by definition and is not affected by external conditions that may prevent certain possibilities from ever being realized, I propose an empirically grounded account of potentiality that involves an assessment of the physical and decisional conditions that may restrict an embryo's possibilities. In my view, some human embryos lack the potentiality to become a person that other human embryos have. Assuming for the sake of argument that the potential to become a person gives a being special moral status, it follows that some human embryos lack this status. This argument is then used to support Gene Outka's suggestion that it is morally permissible to experiment on 'spare' frozen embryos that are destined to be destroyed.

  3. Measuring embryo metabolism to predict embryo quality.

    PubMed

    Thompson, Jeremy G; Brown, Hannah M; Sutton-McDowall, Melanie L

    2016-01-01

    Measuring the metabolism of early embryos has the potential to be used as a prospective marker for post-transfer development, either alone or in conjunction with other embryo quality assessment tools. This is necessary to maximise the opportunity of couples to have a healthy child from assisted reproduction technology (ART) and for livestock breeders to efficiently improve the genetics of their animals. Nevertheless, although many promising candidate substrates (e.g. glucose uptake) and methods (e.g. metabolomics using different spectroscopic techniques) have been promoted as viability markers, none has yet been widely used clinically or in livestock production. Herein we review the major techniques that have been reported; these are divided into indirect techniques, where measurements are made from the embryo's immediate microenvironment, or direct techniques that measure intracellular metabolic activity. Both have strengths and weaknesses, the latter ruling out some from contention for use in human ART, but not necessarily for use in livestock embryo assessment. We also introduce a new method, namely multi- (or hyper-) spectral analysis, which measures naturally occurring autofluorescence. Several metabolically important molecules have fluorescent properties, which we are pursuing in conjunction with improved image analysis as a viable embryo quality assessment methodology.

  4. Ethics for embryos

    PubMed Central

    Parker, C

    2007-01-01

    This paper responds to DW Brock's technically strong case for the use of human embryonic stem cells in medical research. His main issue in this context is the question of whether it is moral to destroy viable human embryos. He offers a number of reasons to support his view that it is moral to destroy them, but his use of conceptual arguments is not adequate to secure his position. The purpose and scope of this paper is wholly concerned with his arguments rather than with the conclusion that it is justifiable to destroy human embryos. The author proceeds through his variety of arguments and offers reasons for rejecting them. The author concludes that Brock has not shown that it is moral to destroy viable human embryos. PMID:17906062

  5. The First Human Cloned Embryo.

    ERIC Educational Resources Information Center

    Cibelli, Jose B.; Lanza, Robert P.; West, Michael D.; Ezzell, Carol

    2002-01-01

    Describes a process known as parthenogenesis which produces cloned, early-stage embryos and human embryos generated only from eggs. Speculates that this technology puts therapeutic cloning within reach. (DDR)

  6. Researchers Create Artificial Mouse 'Embryo'

    MedlinePlus

    ... news/fullstory_163881.html Researchers Create Artificial Mouse 'Embryo' Experiment used two types of gene-modified stem ... they've created a kind of artificial mouse embryo using stem cells, which can be coaxed to ...

  7. The Virtual Embryo Project

    EPA Science Inventory

    The v-Embryo™ is a far reaching new research program at the US EPA to develop a working computer model of a mammalian embryo that can be used to better understand the prenatal risks posed by environmental chemicals and to eventually predict a chemical’s potential developmental to...

  8. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality

    PubMed Central

    Buemo, Carla Paola; Gambini, Andrés; Moro, Lucia Natalia; Hiriart, María Inés; Fernández-Martín, Rafael; Collas, Philippe; Salamone, Daniel Felipe

    2016-01-01

    In this study, we analyzed the effects of the cloned embryo aggregation on in vitro embryo development and embryo quality by measuring blastocyst diameter and cell number, DNA fragmentation levels and the expression of genes associated with pluripotency, apoptosis, trophoblast and DNA methylation in the porcine. Zona-free reconstructed cloned embryos were cultured in the well of the well system, placing one (1x non aggregated group) or three (3x group) embryos per microwell. Our results showed that aggregation of three embryos increased blastocyst formation rate and blastocyst diameter of cloned pig embryos. DNA fragmentation levels in 3x aggregated cloned blastocysts were significantly decreased compared to 1x blastocysts. Levels of Oct4, Klf4, Igf2, Bax and Dnmt 1 transcripts were significantly higher in aggregated embryos, whereas Nanog levels were not affected. Transcripts of Cdx2 and Bcl-xl were essentially non-detectable. Our study suggests that embryo aggregation in the porcine may be beneficial for cloned embryo development and embryo quality, through a reduction in apoptotic levels and an improvement in cell reprogramming. PMID:26894831

  9. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality.

    PubMed

    Buemo, Carla Paola; Gambini, Andrés; Moro, Lucia Natalia; Hiriart, María Inés; Fernández-Martín, Rafael; Collas, Philippe; Salamone, Daniel Felipe

    2016-01-01

    In this study, we analyzed the effects of the cloned embryo aggregation on in vitro embryo development and embryo quality by measuring blastocyst diameter and cell number, DNA fragmentation levels and the expression of genes associated with pluripotency, apoptosis, trophoblast and DNA methylation in the porcine. Zona-free reconstructed cloned embryos were cultured in the well of the well system, placing one (1x non aggregated group) or three (3x group) embryos per microwell. Our results showed that aggregation of three embryos increased blastocyst formation rate and blastocyst diameter of cloned pig embryos. DNA fragmentation levels in 3x aggregated cloned blastocysts were significantly decreased compared to 1x blastocysts. Levels of Oct4, Klf4, Igf2, Bax and Dnmt 1 transcripts were significantly higher in aggregated embryos, whereas Nanog levels were not affected. Transcripts of Cdx2 and Bcl-xl were essentially non-detectable. Our study suggests that embryo aggregation in the porcine may be beneficial for cloned embryo development and embryo quality, through a reduction in apoptotic levels and an improvement in cell reprogramming.

  10. Ensoulment and IVF embryos.

    PubMed Central

    Shea, M C

    1987-01-01

    This paper examines the metaphysical question of 'ensoulment' in relation to the theory, put forward in an earlier paper, that human life begins when the newly formed body organs and systems of the embryo begin to function as an organised whole, at which stage there is evidence of a change of nature. Although Roman Catholic theology teaches that a human being is a union of physical body and spiritual soul, it is incorrect to interpret this in a dualistic sense. The meaning of 'soul' is considered and the conclusion reached that although both in the religious context and apart from it abortion is difficult to justify at any stage after conception, it does not follow that the use of 'spare' In Vitro Fertilisation (IVF) embryos should be rejected. If 'ensoulment' does not occur until the new organism functions as a whole then a decision not to make use of IVF embryos for medical purposes would be a heavy responsibility and not a 'safe' way out. PMID:3612702

  11. Gender determination of avian embryo

    DOEpatents

    Daum, Keith A.; Atkinson, David A.

    2002-01-01

    Disclosed is a method for gender determination of avian embryos. During the embryo incubation process, the outer hard shells of eggs are drilled and samples of allantoic fluid are removed. The allantoic fluids are directly introduced into an ion mobility spectrometer (IMS) for analysis. The resulting spectra contain the relevant marker peaks in the positive or negative mode which correlate with unique mobilities which are sex-specific. This way, the gender of the embryo can be determined.

  12. Electrothermal branding for embryo labeling.

    PubMed

    Wang, L; Beebe, D J; Williams, A R; Easley, K D

    1997-11-01

    A novel embryo labeling technique based on electrothermal branding is developed. Two types of micro branding irons are fabricated and tested. One utilizes 25 microns tungsten wire as the heating element. The other utilizes surface micromachining techniques to fabricate polysilicon branding irons. The thermal behavior of the branding irons and the heat distributions in the embryos are analytically modeled. Micron-scale labels on unfertilized bovine embryos are achieved.

  13. Ethics and embryos.

    PubMed

    Poplawski, N; Gillett, G

    1991-06-01

    In this paper we argue that the human form should be seen to exist, in a longitudinal way, throughout the continuum of human growth and development. This entails that the moral value of that form, which we link analytically to the adult, interacting, social and rational being, attaches to all phases of human life to some extent. Having established this we discuss the consequences it has for the moral status of the human embryo. We then apply this argument, and the resulting moral status, to the area of reproductive technology. In doing this we show that there are certain regulations and controls which ought to apply to the use of these infertility treatments.

  14. Human research cloning, embryos, and embryo-like artifacts.

    PubMed

    Hyun, Insoo; Jung, Kyu Won

    2006-01-01

    Research suggests that cloning is incapable of producing a viable embryo when it is used on primate eggs. In fact, the entity created may not qualify as an embryo at all. If the results stand, cloning avoids the moral objections typically lodged against it, and cloning is itself an "alternative source" of stem cells.

  15. Resolving disputes over frozen embryos.

    PubMed

    Robertson, J A

    1989-01-01

    The relation between respect for family and reproductive choice and use of IVF technology is in dispute in recent legal cases on the disposition of frozen embryos. Couples in IVF programs should be encouraged to stipulate in advance binding instructions regarding the disposition of such embryos.

  16. Genes, embryos, and future people.

    PubMed

    Glannon, Walter

    1998-07-01

    Testing embryonic cells for genetic abnormalities gives us the capacity to predict whether and to what extent people will exist with disease and disability. Moreover, the freezing of embryos for long periods of time enables us to alter the length of a normal human lifespan. After highlighting the shortcomings of somatic-cell gene therapy and germ-line genetic alteration, I argue that the testing and selective termination of genetically defective embryos is the only medically and morally defensible way to prevent the existence of people with severe disability, pain and suffering that make their lives not worth living for them on the whole. In addition, I consider the possible harmful effects on children born from frozen embryos after the deaths of their biological parents, or when their parents are at an advanced age. I also explore whether embryos have moral status and whether the prospects for disease-preventing genetic alteration can justify long-term cryopreservation of embryos.

  17. Cryobiological preservation of Drosophila embryos

    SciTech Connect

    Mazur, P.; Schreuders, P.D.; Cole, K.W.; Hall, J.W. ); Mahowald, A.P. )

    1992-12-18

    The inability to cryobiologically preserve the fruit fly Drosophila melanogaster has required that fly stocks be maintained by frequent transfer of adults. This method is costly in terms of time and can lead to loss of stocks. Traditional slow freezing methods do not succeed because the embryos are highly sensitive to chilling. With the procedures described here, 68 percent of precisely staged 15-hour Oregon R (wild-type) embryos hatch after vitrification at -205[degree]C, and 40 percent of the resulting larvae develop into normal adult flies. These embryos are among the most complex organisms successfully preserved by cryobiology.

  18. Physical influences on embryo development.

    PubMed

    Deeming, D C; Rowlett, K; Simkiss, K

    1987-01-01

    There is a critical period between 3 and 7 days of incubation when the absence of turning in eggs of the domestic fowl leads to increased mortality and decreased embryo growth. This critical period coincides with the time of subembryonic fluid formation, and it is suggested that the absence of turning leads to the presence of unstirred layer effects in fluid secretion. This fluid deficiency persists throughout the subsequent development of the embryo. Experiments on shell-less culture systems support this interpretation in preference to other explanations of embryo death in unturned eggs, which usually refer to chorion adhesion to shell membranes.

  19. Ion currents in embryo development.

    PubMed

    Tosti, Elisabetta; Boni, Raffaele; Gallo, Alessandra

    2016-03-01

    Ion channels are proteins expressed in the plasma membrane of electrogenic cells. In the zygote and blastomeres of the developing embryo, electrical modifications result from ion currents that flow through these channels. This phenomenon implies that ion current activity exerts a specific developmental function, and plays a crucial role in signal transduction and the control of embryogenesis, from the early cleavage stages and during growth and development of the embryo. This review describes the involvement of ion currents in early embryo development, from marine invertebrates to human, focusing on the occurrence, modulation, and dynamic role of ion fluxes taking place on the zygote and blastomere plasma membrane, and at the intercellular communication between embryo cell stages.

  20. The Climatology of Hailstone Embryos.

    NASA Astrophysics Data System (ADS)

    Knight, Nancy C.

    1981-07-01

    Data on hailstone embryo types, using a broad classification as graupel or frozen drops, are presented from several geographical areas representing distinctly different storm `climatologies.' The relative frequency of the two embryo types varies greatly from area to area, in a Way that correlates rather well with average cloud-base temperature. The warmer based clouds produce hail with more frozen drop embryos. The correlation may be explainable either in terms of the dominant precipitation growth process-liquid coalescence or the ice process-or in terms of recycling of embryos, or both. In light of these results, the transferability of any hail suppression technology from one area to another should not be considered to be automatic.

  1. DAPI Staining of Drosophila Embryos.

    PubMed

    Rothwell, Wendy F; Sullivan, William

    2007-10-01

    INTRODUCTIONDrosophila embryos can be stained with specific fluorescent probes or antibodies through either direct or indirect immunofluorescence. In particular, several effective probes exist for visualizing DNA. 4',6-diamidino-2-phenylindole (DAPI) is a commonly used DNA-binding dye. Because it is specific for double-stranded DNA, no prior RNase treatment is required. While the embryo staining method described here uses DAPI, other fluorescent DNA probes can be processed similarly.

  2. Phaseolus immature embryo rescue technology.

    PubMed

    Geerts, Pascal; Toussaint, André; Mergeai, Guy; Baudoin, Jean-Pierre

    2011-01-01

    Predominant among the production constraints of the common bean Phaseolus vulgaris are infestation of Ascochyta blight, Bean Golden Mosaic virus (BGMV), and Bean Fly. Interbreeding with Phaseolus -coccineus L. and/or Phaseolus polyanthus Greenm has been shown to provide P. vulgaris with greater resistance to these diseases. For interspecific crosses to be successful, it is important to use P. coccineus and P. polyanthus as female parents; this prevents rapid reversal to the recurrent parent P. vulgaris. Although incompatibility barriers are post-zygotic, early hybrid embryo abortion limits the success of F1 crosses. While rescue techniques for globular and early heart-shaped embryos have improved in recent years, -success in hybridization remains very low. In this study, we describe six steps that allowed us to rescue 2-day-old P. vulgaris embryos using a pod culture technique. Our methods consisted of (i) pod culture, (ii) extraction and culture of immature embryos, (iii) dehydration of embryos, (iv) germination of embryos, (v) rooting of developed shoots, and (vi) hardening of plantlets.

  3. Lessons from Embryos: Haeckel's Embryo Drawings, Evolution, and Secondary Biology Textbooks

    ERIC Educational Resources Information Center

    Wellner, Karen L.

    2014-01-01

    In 1997, developmental biologist Michael Richardson compared his research team's embryo photographs to Ernst Haeckel's 1874 embryo drawings and called Haeckel's work "noncredible". "Science" soon published "Haeckel's Embryos: Fraud Rediscovered," and Richardson's comments further reinvigorated criticism of Haeckel by…

  4. Avian embryos in hypoxic environments.

    PubMed

    León-Velarde, F; Monge-C, C

    2004-08-12

    Avian embryos at high altitude do not benefit of the maternal protection against hypoxia as in mammals. Nevertheless, avian embryos are known to hatch successfully at altitudes between 4,000 and 6,500 m. This review considers some of the processes that bring about the outstanding modifications in the pressure differences between the environment and mitochondria of avian embryos in hypoxic environments. Among species, some maintain normal levels of oxygen consumption ( VO2) have a high oxygen carrying capacity, lower the air cell-arterial pressure difference ( PAO2 - PaO2 ) with a constant pH. Other species decrease VO2, increase only slightly the oxygen carrying capacity, have a higher PAO2 - PaO2 difference than sea-level embryos and lower the PCO2 and pH. High altitude embryos, and those exposed to hypoxia have an accelerated decline of erythrocyte ATP levels during development and an earlier stimulation of 2,3-BPG synthesis. A higher Bohr effect may ensure high tissue PO2 in the presence of the high-affinity hemoglobin. Independently of the strategy used, they serve together to promote suitable rates of development and successful hatching of high altitude birds in hypoxic environments.

  5. Observations of the Embryos of Graupel.

    NASA Astrophysics Data System (ADS)

    Takahashi, Tsuneya; Fukuta, Norihiko

    1988-11-01

    Embryos in natural graupel particles were studied with a setereomicroscope by carefully disassembling samples gathered at two locations globally apart.For the majority of graupel particle clearly identifiable embryos did not exist, suggesting that embryos of graupel occurred as a result of rime breakup. In some cases, ice particles, frozen drops, plates, dendritic crystals, and broken crystals were found as embryos; the minimum sizes of embryos of graupel were 0.3 mm (dendritic crystal) and 0.1 mm (others). Isometric crystals, which show a high probability of growing into graupel due to their fast fall velocities, were also confirmed as being embryos of graupel.

  6. Feminists on the inalienability of human embryos.

    PubMed

    McLeod, Carolyn; Baylis, Francoise

    2006-01-01

    The feminist literature against the commodification of embryos in human embryo research includes an argument to the effect that embryos are "intimately connected" to persons, or morally inalienable from them. We explore why embryos might be inalienable to persons and why feminists might find this view appealing. But, ultimately, as feminists, we reject this view because it is inconsistent with full respect for women's reproductive autonomy and with a feminist conception of persons as relational, embodied beings. Overall, feminists should avoid claims about embryos' being inalienable to persons in arguments for or against the commodification of human embryos.

  7. In amnio MRI of mouse embryos.

    PubMed

    Roberts, Thomas A; Norris, Francesca C; Carnaghan, Helen; Savery, Dawn; Wells, Jack A; Siow, Bernard; Scambler, Peter J; Pierro, Agostino; De Coppi, Paolo; Eaton, Simon; Lythgoe, Mark F

    2014-01-01

    Mouse embryo imaging is conventionally carried out on ex vivo embryos excised from the amniotic sac, omitting vital structures and abnormalities external to the body. Here, we present an in amnio MR imaging methodology in which the mouse embryo is retained in the amniotic sac and demonstrate how important embryonic structures can be visualised in 3D with high spatial resolution (100 µm/px). To illustrate the utility of in amnio imaging, we subsequently apply the technique to examine abnormal mouse embryos with abdominal wall defects. Mouse embryos at E17.5 were imaged and compared, including three normal phenotype embryos, an abnormal embryo with a clear exomphalos defect, and one with a suspected gastroschisis phenotype. Embryos were excised from the mother ensuring the amnion remained intact and stereo microscopy was performed. Embryos were next embedded in agarose for 3D, high resolution MRI on a 9.4T scanner. Identification of the abnormal embryo phenotypes was not possible using stereo microscopy or conventional ex vivo MRI. Using in amnio MRI, we determined that the abnormal embryos had an exomphalos phenotype with varying severities. In amnio MRI is ideally suited to investigate the complex relationship between embryo and amnion, together with screening for other abnormalities located outside of the mouse embryo, providing a valuable complement to histology and existing imaging methods available to the phenotyping community.

  8. Embryo adoption: Some further considerations

    PubMed Central

    Patterson, Colin

    2015-01-01

    Recent discussions of embryo adoption have sought to make sense of the teaching of the Congregation for the Doctrine of the Faith (CDF) document Dignitas personae which appeared to provide a negative judgment on such a practice. This article aims to provide a personalist account of the process of fertilization and implantation that might serve as the basis for the negative judgment of the CDF document. In doing so, it relies upon the idea that a person, including an embryo, is not to be considered in isolation, but always in relation to God and to others. This approach extends the substantialist conceptualizations commonly employed in discussions of this issue. More generally, the article seeks to highlight the value of a personalist re-framing for an understanding of the moral questions surrounding the beginning of life. Lay summary: This article seeks to make sense of what appears to be a clear-cut rejection, set out in the Congregation for the Doctrine of the Faith (CDF) document Dignitas personae, of the proposal for women to “adopt” surplus frozen embryos. It draws upon more recently developed modes of philosophical/theological reasoning to argue that, in human procreation, both fertilization and implantation represent constitutive dimensions of divine creative activity and so must be protected from manipulative technological intervention. Since embryo adoption requires this kind of technology, it makes sense for the Church document not to approve it. PMID:25698841

  9. Untwisting the Caenorhabditis elegans embryo

    PubMed Central

    Christensen, Ryan Patrick; Bokinsky, Alexandra; Santella, Anthony; Wu, Yicong; Marquina-Solis, Javier; Guo, Min; Kovacevic, Ismar; Kumar, Abhishek; Winter, Peter W; Tashakkori, Nicole; McCreedy, Evan; Liu, Huafeng; McAuliffe, Matthew; Mohler, William; Colón-Ramos, Daniel A; Bao, Zhirong; Shroff, Hari

    2015-01-01

    The nematode Caenorhabditis elegans possesses a simple embryonic nervous system with few enough neurons that the growth of each cell could be followed to provide a systems-level view of development. However, studies of single cell development have largely been conducted in fixed or pre-twitching live embryos, because of technical difficulties associated with embryo movement in late embryogenesis. We present open-source untwisting and annotation software (http://mipav.cit.nih.gov/plugin_jws/mipav_worm_plugin.php) that allows the investigation of neurodevelopmental events in late embryogenesis and apply it to track the 3D positions of seam cell nuclei, neurons, and neurites in multiple elongating embryos. We also provide a tutorial describing how to use the software (Supplementary file 1) and a detailed description of the untwisting algorithm (Appendix). The detailed positional information we obtained enabled us to develop a composite model showing movement of these cells and neurites in an 'average' worm embryo. The untwisting and cell tracking capabilities of our method provide a foundation on which to catalog C. elegans neurodevelopment, allowing interrogation of developmental events in previously inaccessible periods of embryogenesis. DOI: http://dx.doi.org/10.7554/eLife.10070.001 PMID:26633880

  10. 9 CFR 98.9 - Embryos refused entry.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Embryos refused entry. 98.9 Section 98... EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses and Asses § 98.9 Embryos refused entry. Any embryo refused entry into...

  11. 9 CFR 98.9 - Embryos refused entry.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Embryos refused entry. 98.9 Section 98... EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses and Asses § 98.9 Embryos refused entry. Any embryo refused entry into...

  12. 9 CFR 98.9 - Embryos refused entry.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Embryos refused entry. 98.9 Section 98... EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses and Asses § 98.9 Embryos refused entry. Any embryo refused entry into...

  13. 9 CFR 98.9 - Embryos refused entry.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Embryos refused entry. 98.9 Section 98... EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses and Asses § 98.9 Embryos refused entry. Any embryo refused entry into...

  14. 9 CFR 98.9 - Embryos refused entry.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Embryos refused entry. 98.9 Section 98... EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses and Asses § 98.9 Embryos refused entry. Any embryo refused entry into...

  15. Embryo technologies in the horse.

    PubMed

    Squires, E L; Carnevale, E M; McCue, P M; Bruemmer, J E

    2003-01-01

    Recent studies demonstrated that zwitterionic buffers could be used for satisfactory storage of equine embryos at 5 degrees C. The success of freezing embryos is dependent upon size and stage of development. Morulae and blastocysts <300 microm can be slowly cooled or vitrified with acceptable pregnancy rates after transfer. The majority of equine embryos are collected from single ovulating mares, as there is no commercially available product for superovulation in equine. However, pituitary extract, rich in FSH, can be used to increase embryo recovery three- to four-fold. Similar to human medicine, assisted reproductive techniques have been developed for the older, subfertile mare. Transfer of in vivo-matured oocytes from young, healthy mares into a recipient's oviduct results in a 70-80% pregnancy rate compared with a 30-40% pregnancy rate when the oocytes are from older, subfertile mares. This procedure can also be used to evaluate in vitro maturation systems. In vitro production of embryos is still quite difficult in the horse. However, intracytoplasmic sperm injection (ICSI) has been used to produce several foals. Cleavage rates of 60% and blastocyst rates of 30% have been reported after ICSI of in vitro-matured oocytes. Gamete intrafallopian tube transfer (GIFT) is a possible treatment for subfertile stallions. Transfer of in vivo-matured oocytes with 200,000 sperm into the oviduct of normal mares resulted in a pregnancy rate of 55-82%. Oocyte freezing is a technique that has proven difficult in most species. However, equine oocytes vitrified in a solution of ethylene glycol, DMSO, and Ficoll and loaded onto a cryoloop resulted in three pregnancies of 26 transfers and two live foals produced. Production of a cloned horse appears to be likely, as several cloned pregnancies have recently been produced.

  16. Comparison of three embryo culture methods for derivation of human embryonic stem cells from discarded embryos.

    PubMed

    Liu, Ying; Li, Yang; Hwang, Andrew; Wang, Shu-yu; Jia, Chan-wei; Yu, Lan; Li, Jian

    2011-06-01

    Human embryonic stem cells (hESC) are self-renewing and pluripotent cells that hold great promise. Our objective was to compare the effect of three different embryo culture methods for derivation of human embryonic stem cells from discarded embryos. A prospective and randomized trial was conducted using 381 discarded human embryos at days 2-3 postfertilization in Beijing Obstetrics and Gynecology Hospital IVF center. After removal of the zona pellucida, discarded human embryos were cultured by three different methods as multiple embryo aggregates, single embryo, and blastomeres. Outgrowth of embryos and hESC derivation were observed. The outgrowth rate of embryos cultured as multiple embryo aggregates was higher than that of those cultured as single embryos or blastomeres (p < 0.05). Three propagating hESC lines were derived from poor quality day 2-3 postfertilization nonblastocyst embryos cultured as multiple embryo aggregates. Derived hESC lines expressed hESC-specific markers of pluripotency and had normal diploid karyotype. The cells were able to form derivatives of all three germ layers in vivo as teratomas. Our results demonstrate that culturing these discarded embryos as multiple embryo aggregates was more profitable for outgrowth and derivation of ESC line than culturing these as single embryo or blastomeres.

  17. Adoption first? The disposition of human embryos.

    PubMed

    Murphy, Timothy F

    2014-06-01

    Anja Karnein has suggested that because of the importance of respect for persons, law and policy should require some human embryos created in vitro to be available for adoption for a period of time. If no one comes forward to adopt the embryos during that time, they may be destroyed (in the case of embryos left over from fertility medicine) or used in research (in the case of embryos created for that purpose or left over from fertility medicine). This adoption option would increase the number of embryos available for couples looking for help in having children, but that effect is less important--Karnein argues--than the observance of respect for human persons. As possible persons, she holds that embryos ought to be treated, as if they will become children, if only for a while. If enacted as a matter of law and policy, an 'adoption option' would wrongly interfere with the dispositional rights women and men ought to have over embryos they create in the course of trying to have children. Karnein's proposal would also deprive researchers of certainty that the embryos they create for research would actually be available that way, leading to increased burdens of time and money and maybe even to more embryos than would otherwise be produced. Karnein's analysis does not show, moreover, that any duty of rescue applies to embryos. No woman is required to adopt any embryo, which significantly undercuts the justification for an obligatory adoption period.

  18. Induction of autophagy improves embryo viability in cloned mouse embryos

    PubMed Central

    Shen, XingHui; Zhang, Na; Wang, ZhenDong; Bai, GuangYu; Zheng, Zhong; Gu, YanLi; Wu, YanShuang; Liu, Hui; Zhou, DongJie; Lei, Lei

    2015-01-01

    Autophagy is an essential cellular mechanism that degrades cytoplasmic proteins and organelles to recycle their components. Moreover, autophagy is essential for preimplantation development in mammals. Here we show that autophagy is also important for reprogramming in somatic cell nuclear transfer (SCNT). Our data indicate that unlike fertilized oocytes, autophagy is not triggered in SCNT embryos during 6 hours of activation. Mechanistically, the inhibited autophagic induction during SCNT activation is due to the cytochalasin B (CB) caused depolymerization of actin filaments. In this study, we induced autophagy during SCNT activation by rapamycin and pp242, which could restore the expected level of autophagy and significantly enhance the development of SCNT embryos to the blastocyst stage when compared with the control (68.5% and 68.7% vs. 41.5%, P < 0.05). Furthermore, the treatment of rapamycin and pp242 accelerates active DNA demethylation indicated by the conversion of 5 mC to 5 hmC, and treatment of rapamycin improves degradation of maternal mRNA as well. Thus, our findings reveal that autophagy is important for development of SCNT embryos and inhibited autophagic induction during SCNT activation might be one of the serious causes of low efficiency of SCNT. PMID:26643778

  19. Zeatin reductase in Phaseolus embryos

    SciTech Connect

    Martin, R.C.; Mok, David, W.S.; Mok, M.C. )

    1989-04-01

    Zeatin was converted to O-xylosylzeatin in embryos of Phaseolus vulgaris . O-xylosyldihydrozeatin was also identified as a zeatin metabolite. Incubation of embryo extracts with {sup 14}C-zeatin and {sup 14}C-O-xylosylzeatin revealed that reduction preceeds the O-xylosylation of zeatin. An enzyme responsible for reducing the N{sup 6}-side chain was isolated and partially purified using ammonium sulfate fractionation and affinity, gel filtration and anion exchange chromatography. The NADPH dependent reductase was zeatin specific and did not recognize cis-zeatin, ribosylzeatin, i{sup 6}Ade or i{sup 6}Ado. Two forms of the reductase could be separated by either gel filtration or anion exchange HPLC. The HMW isozyme (Mr. 55,000) eluted from the anion exchange column later than the LMW isozyme (Mr. 25,000). Interspecific differences in zeatin reductase activity were also detected.

  20. Electroporation into Cultured Mammalian Embryos

    NASA Astrophysics Data System (ADS)

    Nomura, Tadashi; Takahashi, Masanori; Osumi, Noriko

    Over the last century, mammalian embryos have been used extensively as a common animal model to investigate fundamental questions in the field of developmental biology. More recently, the establishment of transgenic and gene-targeting systems in laboratory mice has enabled researchers to unveil the genetic mechanisms under lying complex developmental processes (Mak, 2007). However, our understanding of cell—cell interactions and their molecular basis in the early stages of mammalian embryogenesis is still very fragmentary. One of the major problems is the difficulty of precise manipulation and limited accessibility to mammalian embryos via uterus wall. Unfortunately, existing tissue and organotypic culture systems per se do not fully recapitulate three-dimensional, dynamic processes of organogenesis observed in vivo. Although transgenic animal technology and virus-mediated gene delivery are useful to manipulate gene expression, these techniques take much time and financial costs, which limit their use.

  1. Human embryo culture media comparisons.

    PubMed

    Pool, Thomas B; Schoolfield, John; Han, David

    2012-01-01

    Every program of assisted reproduction strives to maximize pregnancy outcomes from in vitro fertilization and selecting an embryo culture medium, or medium pair, consistent with high success rates is key to this process. The common approach is to replace an existing medium with a new one of interest in the overall culture system and then perform enough cycles of IVF to see if a difference is noted both in laboratory measures of embryo quality and in pregnancy. This approach may allow a laboratory to select one medium over another but the outcomes are only relevant to that program, given that there are well over 200 other variables that may influence the results in an IVF cycle. A study design that will allow for a more global application of IVF results, ones due to culture medium composition as the single variable, is suggested. To perform a study of this design, the center must have a patient caseload appropriate to meet study entrance criteria, success rates high enough to reveal a difference if one exists and a strong program of quality assurance and control in both the laboratory and clinic. Sibling oocytes are randomized to two study arms and embryos are evaluated on day 3 for quality grades. Inter and intra-observer variability are evaluated by kappa statistics and statistical power and study size estimates are performed to bring discriminatory capability to the study. Finally, the complications associated with extending such a study to include blastocyst production on day 5 or 6 are enumerated.

  2. Genetics of early embryo survival

    SciTech Connect

    Niswander, L.; Edstroem, J.E.; Rinchik, E.M.; Magnuson, T.

    1989-01-01

    The albino-deletion complex represents one of the few regions of the mouse genome where a set of specific developmental defects are associated with a series of overlapping chromosomal deletions. A total of 37 deletions exist, all of which remove the region of mouse chromosome 7 that surrounds and includes the albino coat-color locus. The complementation patterns among these deletion chromosomes indicate that at least 9 distinct functional units exist in this region. Two of these units contain genes whose expression is required around the time the basic body plan is being established in the early postimplantation embryo. Embryos homozygous for the deletions that remove one of these units develop to day 8.5. Extensive development of the extraembryonic structures occurs, and the three primary germ layers form but there is no organization of mesoderm into somites or induction of the neural axis. Embryos homozygous for deletions that remove the second region, as well as the first, do not undergo gastrulation, and there is no development of the extraembryonic structures. 10 refs., 2 figs., 1 tab.

  3. Embryo donation in Iran: an ethical review.

    PubMed

    Afshar, Leila; Bagheri, Alireza

    2013-12-01

    Iran is the only Muslim country that has legislation on embryo donation, adopted in 2003. With an estimated 10-15% of couples in the country that are infertile, there are not any legal or religious barriers that prohibit an infertile couple from taking advantage of Assisted Reproductive Technologies (ARTs). Although all forms of ARTs available in Iran have been legitimized by religious authorities, there is a lack of legislation in all ARTs except embryo donation. By highlighting ethical issues in embryo donation, the paper presents a critical review of the Act of Embryo Donation in Iran. The paper argues that the Act does not provide enough safeguards for the future child and assurance for the safety of the donated embryos. It also does not restrict embryo donation to surplus embryos from infertile couples and is silent about the number of embryos that could be donated by each couple as well as the number of recipients for donated embryos by a couple. The Act is also silent about the issues of genetic linkage (nasab) and heritage which are challenging issues, especially in a conservative Islamic society. As a result, the future child may not inherit from their birth parents, as it is not required by the Act, or from the genetically related parents under the anonymity policy. Finally there is no standard national protocol or guidelines to evaluate the safety of the donated embryos. The paper concludes that despite its benefits, the Act lacks clarity, and it is subject to misunderstanding and confusion.

  4. Enhance beef cattle improvement by embryo biotechnologies.

    PubMed

    Wu, B; Zan, L

    2012-10-01

    Embryo biotechnology has become one of the prominent high businesses worldwide. This technology has evolved through three major changes, that is, traditional embryo transfer (in vivo embryo production by donor superovulation), in vitro embryo production by ovum pick up with in vitro fertilization and notably current cloning technique by somatic cell nuclear transfer and transgenic animal production. Embryo biotechnology has widely been used in dairy and beef cattle industry and commercial bovine embryo transfer has become a large international business. Currently, many developed biotechnologies during the period from early oocyte stage to pre-implantation embryos can be used to create new animal breeds and accelerate genetic progression. Based on recent advances in embryo biotechnologies and authors current studies, this review will focus on a description of the application of this technology to beef cattle improvement and discuss how to use this technology to accelerate beef cattle breeding and production. The main topics of this presentation include the following: (i) how to increase calf production numbers from gametes including sperm and oocyte; (ii) multiple ovulation and embryo transfer breeding schemes; (iii) in vitro fertilization and intracytoplasm sperm injection in bovine; (iv) pronuclear development and transgenic animals; (v) sex selection from sperm and embryos; (vi) cloning and androgenesis; (vii) blastocyst development and embryonic stem cells; (viii) preservation of beef cattle genetic resources; and (ix) conclusions.

  5. Generation and developmental characteristics of porcine tetraploid embryos and tetraploid/diploid chimeric embryos.

    PubMed

    He, Wenteng; Kong, Qingran; Shi, Yongqian; Xie, Bingteng; Jiao, Mingxia; Huang, Tianqing; Guo, Shimeng; Hu, Kui; Liu, Zhonghua

    2013-10-01

    The aim of this study was to optimize electrofusion conditions for generating porcine tetraploid (4n) embryos and produce tetraploid/diploid (4n/2n) chimeric embryos. Different electric field intensities were tested and 2 direct current (DC) pulses of 0.9 kV/cm for 30 μs was selected as the optimum condition for electrofusion of 2-cell embryos to produce 4n embryos. The fusion rate of 2-cell embryos and the development rate to blastocyst of presumably 4n embryos, reached 85.4% and 28.5%, respectively. 68.18% of the fused embryos were found to be 4n as demonstrated by fluorescent in situ hybridization (FISH). Although the number of blastomeres in 4n blastocysts was significantly lower than in 2n blastocysts (P<0.05), there was no significant difference in developmental rates of blastocysts between 2n and 4n embryos (P>0.05), suggesting that the blastocyst forming capacity in 4n embryos is similar to those in 2n embryos. Moreover, 4n/2n chimeric embryos were obtained by aggregation of 4n and 2n embryos. We found that the developmental rate and cell number of blastocysts of 4-cell (4n)/4-cell (2n) chimeric embryos were significantly higher than those of 2-cell (4n)/4-cell (2n), 4-cell (4n)/8-cell (2n), 4-cell (4n)/2-cell (2n) chimeric embryos (P<0.05). Consistent with mouse chimeras, the majority of 4n cells contribute to the trophectoderm (TE), while the 2n cells are mainly present in the inner cell mass (ICM) of porcine 4n/2n chimeric embryos. Our study established a feasible and efficient approach to produce porcine 4n embryos and 4n/2n chimeric embryos.

  6. Radioactive labeling of proteins in cultured postimplantation mouse embryos. I. Influence of the embryo preparation method

    SciTech Connect

    Nowak, J.; Klose, J. )

    1989-07-01

    Conditions for optimum incorporation of radioactive amino acids into proteins of cultured postimplantation mouse embryos were investigated under the aspect of using these proteins for two-dimensional electrophoretic separations followed by fluorography. The aim was to obtain highly radioactive proteins under conditions as physiological as possible. Embryos at Days 10, 11, and 12 of gestation were prepared in different ways and incubated for 4 h in Tyrode's solution containing ({sup 3}H)amino acids (mixture) at a concentration of 27 microCi/ml medium. The preparations were: (a) yolk sac opened, placenta and blood circulation intact; (b) yolk sac and amnion opened, placenta and blood circulation intact (Day 10 embryos only); (c) placenta, yolk sac, and amnion removed (embryo naked); (d) naked embryos cut randomly into pieces (Day 10 embryos only). After incubation whole embryos or certain parts (tail, liver, rest body) were investigated by determining the radioactivity taken up by the protein. The results are given in dpm per mg protein per embryo. Radioactivity of proteins was about 3 times higher in naked embryos than in embryos left in their yolk sacs. This was true for all three stages investigated. However, the degree of radioactivity in the various parts of naked embryos differed by a factor of 15, whereas radioactivity was evenly distributed in embryos incubated in their yolk sacs. Therefore, embryos prepared according to the first method (see above) fulfilled the conditions required at the best.

  7. Palaeontology: pterosaur embryo from the Early Cretaceous.

    PubMed

    Wang, Xiaolin; Zhou, Zhonghe

    2004-06-10

    Dinosaur embryos have been discovered all over the world, but so far no pterosaur embryos have been reported. Here we describe a Chinese fossil from the Early Cretaceous period containing an embryo that is unambiguously a pterosaur. The embryonic skeleton, which is exquisitely preserved in its egg, is associated with eggshell fragments, wing membranes and skin imprints. This discovery confirms that pterosaurs were egg-layers and sheds new light on our understanding of pterosaur development.

  8. 9 CFR 98.20 - Embryos refused entry.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Embryos refused entry. 98.20 Section... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.20 Embryos refused entry. If any embryos are determined to be...

  9. 9 CFR 98.16 - The embryo collection unit.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false The embryo collection unit. 98.16... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.16 The embryo collection unit. Ruminant and swine embryos may...

  10. 9 CFR 98.20 - Embryos refused entry.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Embryos refused entry. 98.20 Section... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.20 Embryos refused entry. If any embryos are determined to be...

  11. 9 CFR 98.16 - The embryo collection unit.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false The embryo collection unit. 98.16... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.16 The embryo collection unit. Ruminant and swine embryos may...

  12. 9 CFR 98.20 - Embryos refused entry.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Embryos refused entry. 98.20 Section... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.20 Embryos refused entry. If any embryos are determined to be...

  13. 9 CFR 98.16 - The embryo collection unit.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false The embryo collection unit. 98.16... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.16 The embryo collection unit. Ruminant and swine embryos may...

  14. 9 CFR 98.20 - Embryos refused entry.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Embryos refused entry. 98.20 Section... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.20 Embryos refused entry. If any embryos are determined to be...

  15. 9 CFR 98.16 - The embryo collection unit.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false The embryo collection unit. 98.16... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.16 The embryo collection unit. Ruminant and swine embryos may...

  16. 9 CFR 98.20 - Embryos refused entry.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Embryos refused entry. 98.20 Section... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.20 Embryos refused entry. If any embryos are determined to be...

  17. 9 CFR 98.16 - The embryo collection unit.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false The embryo collection unit. 98.16... CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.16 The embryo collection unit. Ruminant and swine embryos may...

  18. Transmission OF Campylobacter coli in chicken embryos

    PubMed Central

    Rossi, Daise Aparecida; Fonseca, Belchiolina Beatriz; de Melo, Roberta Torres; Felipe, Gutembergue da Silva; da Silva, Paulo Lourenço; Mendonça, Eliane Pereira; Filgueiras, Ana Luzia Lauria; Beletti, Marcelo Emilio

    2012-01-01

    Campylobacter coli is an important species involved in human cases of enteritis, and chickens are carriers of the pathogen mainly in developing country. The current study aimed to evaluate the transmission of C. coli and its pathogenic effects in chicken embryos. Breeder hens were inoculated intra-esophageally with C. coli isolated from chickens, and their eggs and embryos were analyzed for the presence of bacteria using real-time PCR and plate culture. The viability of embryos was verified. In parallel, SPF eggs were inoculated with C. coli in the air sac; after incubation, the embryos were submitted to the same analysis as the embryos from breeder hens. In embryos and fertile eggs from breeder hens, the bacterium was only identified by molecular methods; in the SPF eggs, however, the bacterium was detected by both techniques. The results showed no relationship between embryo mortality and positivity for C. coli in the embryos from breeder hens. However, the presence of bacteria is a cause of precocious mortality for SPF embryos. This study revealed that although the vertical transmission is a possible event, the bacteria can not grow in embryonic field samples. PMID:24031861

  19. Research on human embryos--a justification.

    PubMed

    Brown, J

    1986-12-01

    The philosophical debate surrounding the moral status of the embryo has reached the public arena. The author of this paper examines some of the common arguments against embryo experimentation, including an influential article by Professor Ian Kennedy. He concludes that these arguments do not succeed in demonstrating that the intentional creation of embryos for research purposes is wrong, unless they also succeed in demonstrating that contemporary liberal abortion laws are also wrong. The author also criticises the conclusions of the Warnock Report, and suggests that the reasons for permitting embryo research must be given a wider public audience.

  20. Cryopreservation of immature embryos of Theobroma cacao.

    PubMed

    Pence, V C

    1991-06-01

    Immature, white zygotic embryos of Theobroma cacao L. (cacao) retained the ability to produce callus and to undergo somatic embryogenesis after slow hydrated freezing and desiccated fast freezing in liquid nitrogen. The highest rate of somatic embryogenesis occurred in embryos which were precultured on a medium containing 3% sucrose, frozen slowly with cryoprotectants before exposure to liquid nitrogen, and recovered on a medium containing 3 mg/liter NAA. Embryos precultured on media containing sucrose increasing to 21% had a higher rate of survival but were less embryogenic after freezing. These results suggest that immature embryos might be used for long-term germplasm storage of T. cacao germplasm.

  1. In vitro culture and embryo metabolism of cattle and sheep embryos - a decade of achievement.

    PubMed

    Thompson, J G

    2000-07-02

    At the beginning of the 1990s, co-culture of cattle and sheep embryos was the most favoured method to support embryo development, but the use of this system has hampered progress in raising the efficiency of embryo production. Furthermore, little was known of the requirements of embryos and the biochemistry of early embryo development. As the decade progressed, energy metabolism studies improved our understanding of the energy substrate requirements for embryo development. Furthermore, an appreciation of the reproductive tract environment increased. This resulted in more "defined" systems, which have evolved further in the development of "sequential" media systems, where components change in accordance to the needs of the embryo. Nevertheless, wholly defined systems, such as the replacement of albumin with PVA, are less able to support similar levels of development as protein-containing medium, and the resulting embryos are metabolically compromised. This highlights the nutritive role of albumin. One area in which much work has been conducted, but yet no unifying theory has emerged, is that of the interactive roles of growth factors (including autocrine/paracrine), cytokines and extra-cellular matrix molecules in the development of a viable embryo. A new concept is that of regulation of energy metabolism. Compounds such as ethylenediamine tetraacetic acid (EDTA), NaN(3) and 2,4-dinitrophenol have been shown to increase embryo development and quality of resulting embryos. This demonstrates that the process of ATP production is a key regulator of in vitro embryo development.

  2. Navigating the site for embryo implantation: biomechanical and molecular regulation of intrauterine embryo distribution.

    PubMed

    Chen, Qi; Zhang, Ying; Elad, David; Jaffa, Ariel J; Cao, Yujing; Ye, Xiaoqin; Duan, Enkui

    2013-10-01

    The distribution of intrauterine embryo implantation site(s) in most mammalian species shows remarkably constant patterns: in monotocous species such as humans, an embryo tends to implant in the uterine fundus; in polytocous species such as rodents, embryos implant evenly along the uterine horns. These long-time evolved patterns bear great biological significance because disruption of these patterns can have adverse effects on pregnancies. However, lack of suitable models and in vivo monitoring techniques has impeded the progress in understanding the mechanisms of intrauterine embryo distribution. These obstacles are being overcome by genetically engineered mouse models and newly developed high-resolution ultrasound. It has been revealed that intrauterine embryo distribution involves multiple events including uterine sensing of an embryo, fine-tuned uterine peristaltic movements, time-controlled uterine fluid reabsorption and uterine luminal closure, as well as embryo orientation. Diverse molecular factors, such as steroid hormone signaling, lipid signaling, adrenergic signaling, developmental genes, ion/water channels, and potentially embryonic signaling are actively involved in intrauterine embryo distribution. This review covers the biomechanical and molecular aspects of intrauterine embryo distribution (embryo spacing at the longitudinal axis and embryo orientation at the vertical axis), as well as its pathophysiological roles in human reproductive medicine. Future progress requires multi-disciplinary research efforts that will integrate in vivo animal models, clinical cases, physiologically relevant in vitro models, and biomechanical/computational modeling. Understanding the mechanisms for intrauterine embryo distribution could potentially lead to development of therapeutics for treating related conditions in reproductive medicine.

  3. Effects of Fluoxetine on Human Embryo Development

    PubMed Central

    Kaihola, Helena; Yaldir, Fatma G.; Hreinsson, Julius; Hörnaeus, Katarina; Bergquist, Jonas; Olivier, Jocelien D. A.; Åkerud, Helena; Sundström-Poromaa, Inger

    2016-01-01

    The use of antidepressant treatment during pregnancy is increasing, and selective serotonin reuptake inhibitors (SSRIs) are the most widely prescribed antidepressants in pregnant women. Serotonin plays a role in embryogenesis, and serotonin transporters are expressed in two-cell mouse embryos. Thus, the aim of the present study was to evaluate whether fluoxetine, one of the most prescribed SSRI antidepressant world-wide, exposure influences the timing of different embryo developmental stages, and furthermore, to analyze what protein, and protein networks, are affected by fluoxetine in the early embryo development. Human embryos (n = 48) were randomly assigned to treatment with 0.25 or 0.5 μM fluoxetine in culture medium. Embryo development was evaluated by time-lapse monitoring. The fluoxetine-induced human embryo proteome was analyzed by shotgun mass spectrometry. Protein secretion from fluoxetine-exposed human embryos was analyzed by use of high-multiplex immunoassay. The lower dose of fluoxetine had no influence on embryo development. A trend toward reduced time between thawing and start of cavitation was noted in embryos treated with 0.5 μM fluoxetine (p = 0.065). Protein analysis by shotgun mass spectrometry detected 45 proteins that were uniquely expressed in fluoxetine-treated embryos. These proteins are involved in cell growth, survival, proliferation, and inflammatory response. Culturing with 0.5 μM, but not 0.25 μM fluoxetine, caused a significant increase in urokinase-type plasminogen activator (uPA) in the culture medium. In conclusion, fluoxetine has marginal effects on the timing of developmental stages in embryos, but induces expression and secretion of several proteins in a manner that depends on dose. For these reasons, and in line with current guidelines, the lowest possible dose of SSRI should be used in pregnant women who need to continue treatment. PMID:27378857

  4. Methionine requirements for the preimplantation bovine embryo.

    PubMed

    BONILLA, Luciano; LUCHINI, Daniel; DEVILLARD, Estelle; HANSEN, Peter J

    2010-10-01

    The early embryo's nutritional environment plays an important role in establishing its developmental potential. However, little is known about the specific nutrient requirements of the embryo. The objective of the present study was to determine requirements of the in vitro produced bovine embryo for the essential amino acid methionine. In addition to serving as a precursor for polypeptides, methionine plays roles in regulation of translation, DNA methylation, and antioxidant balance. In the first experiment, embryos were cultured in potassium simplex optimized medium - bovine embryo modification 2 containing 0, 35, 50, 100, 200 or 400 µmol/l L-methionine for 8 days. There was no effect of methionine concentration on cleavage rate. The percent of oocytes that developed to blastocyst was lower for embryos without methionine at Day 7 and 8 than other groups but was similar for embryos cultured with 35-400 µmol/l. Neither total cell number, allocation of cells to trophectoderm or inner cell mass, or frequency of apoptosis was affected by methionine concentration. In the second experiment, embryos were cultured with 0, 7, 14, 21, 28 or 35 µmol/l methionine. There was no effect of methionine concentration on cleavage rate. The percent of oocytes that developed to blastocyst was lower for embryos without methionine at Day 7 and 8 but was not different between embryos cultured with 7-35 µmol/l methionine. However, the proportion of blastocysts that were expanded, hatching or hatched on Day 7 was reduced at lower concentrations of methionine (7 and 14). DNA methylation of blastocyst nuclei was unaffected by methionine concentration but intracellular glutathione content was higher for embryos cultured without methionine. In conclusion, the methionine requirement for preimplantation development is between 14 and 21 µmol/l. These concentrations are lower or similar to those found in the reproductive tract and suggest that methionine deficiency is not a common cause of

  5. Embryo development in dairy cattle.

    PubMed

    Lonergan, Pat; Fair, Trudee; Forde, Niamh; Rizos, Dimitrios

    2016-07-01

    During the past 50 years, the fertility of high-producing lactating dairy cows has decreased, associated with intensive selection for increased milk production. The physiological and metabolic changes associated with high milk production, including decreased (glucose, insulin, IGF-I) or increased (nonesterified fatty acids, ketone bodies) concentrations of circulating metabolites during nutrient partitioning associated with negative energy balance as well as uterine and nonuterine diseases have been linked with poor reproductive efficiency. Fertilization is typically above 80% and does not seem to be the principal factor responsible for the low fertility in dairy cows. However, early embryonic development is compromised in high-producing dairy cows, as observed by most embryonic losses occurring during the first 2 weeks after fertilization and may be linked to compromised oocyte quality due to a poor follicular microenvironment, suboptimal reproductive tract environment for the embryo, and/or inadequate maternal-embryonic communication. These and other factors related to embryo development will be discussed.

  6. The Virtual Embryo Project (v-Embryo™)

    EPA Science Inventory

    The v-Embryo is a far reaching new research program at the US EPA to develop a working computer model of a mammalian embryo that can be used to better understand the prenatal risks posed by environmental chemicals and to eventually predict a chemical's potential developmental tox...

  7. Murine cytomegalovirus infection of cultured mouse embryos.

    PubMed Central

    Tsutsui, Y.; Naruse, I.

    1987-01-01

    Isolated mouse whole embryos of 7.5 days' gestation were infected with murine cytomegalovirus (MCMV) and cultured in pure rat serum. Although the MCMV infection had little effect on the survival and development of the embryos during 3 days of cultivation, immunohistochemical analysis of their serial sections using monoclonal antibody showed MCMV-infected cells in various portions of the embryos. This monoclonal antibody, when tested with the use of infected cultured mouse fibroblasts, reacted with nuclear antigen within 2 hours after infection and also reacted with nuclear inclusions in the late phase of infection. The viral antigen-positive cells detected by the monoclonal antibody were present in almost all of the ectoplacental cone and the yolk sac and in about 82% of the embryos. In the embryos, antigen-positive cells were frequently observed in the epithelium of the digestive tracts, endothelial cells of the blood vessels, and the mesodermal cells. In some of the embryos, viral antigen-positive cells were clearly observed in a small percentage of the blood cells. These findings indicate that blood cells, in addition to cell migration during embryogenesis, may play an important role in transmission of infectious virus into the embryos. Mouse whole embryo culture infected with MCMV can provide a model for the study of cellular tropism related to congenital infection by cytomegalovirus. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:3034066

  8. Supplement of autologous ooplasm into porcine somatic cell nuclear transfer embryos does not alter embryo development.

    PubMed

    Lee, W-J; Lee, J-H; Jeon, R-H; Jang, S-J; Lee, S-C; Park, J-S; Lee, S-L; King, W-A; Rho, G-J

    2017-02-13

    Somatic cell nuclear transfer (SCNT) is considered as the technique in which a somatic cell is introduced into an enucleated oocyte to make a cloned animal. However, it is unavoidable to lose a small amount of the ooplasm during enucleation step during SCNT procedure. The present study was aimed to uncover whether the supplement of autologous ooplasm could ameliorate the oocyte competence so as to improve low efficiency of embryo development in porcine SCNT. Autologous ooplasm-transferred (AOT) embryos were generated by the supplementation with autologous ooplasm into SCNT embryos. They were comparatively evaluated with respect to embryo developmental potential, the number of apoptotic body formation and gene expression including embryonic lineage differentiation, apoptosis, epigenetics and mitochondrial activity in comparison with parthenogenetic, in vitro-fertilized (IVF) and SCNT embryos. Although AOT embryos showed perfect fusion of autologous donor ooplasm with recipient SCNT embryos, the supplement of autologous ooplasm could not ameliorate embryo developmental potential in regard to the rate of blastocyst formation, total cell number and the number of apoptotic body. Furthermore, overall gene expression of AOT embryos was presented with no significant alterations in comparison with that of SCNT embryos. Taken together, the results of AOT demonstrated inability to make relevant values improved from the level of SCNT embryos to their IVF counterparts.

  9. Intrauterine embryo transfer with canine embryos cryopreserved by the slow freezing and the Cryotop method

    PubMed Central

    HORI, Tatsuya; USHIJIMA, Hitoshi; KIMURA, Taku; KOBAYASHI, Masanori; KAWAKAMI, Eiichi; TSUTSUI, Toshihiko

    2016-01-01

    Canine embryos (8-cell to blastocyst stages) frozen-thawed using the slow-freezing method with glycerol (four recipients) or dimethyl sulfoxide (three recipients) as a cryoprotectant and vitrified-warmed using the Cryotop method (five recipients) were surgically transferred into the unilateral uterine horn of recipient bitches. As a result, the morphology of embryos frozen-thawed using the slow-freezing method was judged to be normal, but no conception occurred in any of the recipient bitches. Two of the five bitches that received transferred embryos (morula to early blastocyst stages) vitrified-warmed using the Cryotop method became pregnant and produced normal pups (1/9 embryos, 11.1% and 1/6 embryos, 17.0%). It was concluded that the Cryotop method was more appropriate for canine embryo cryopreservation than the slow-freezing method, which is used for the cryopreservation of embryos of other mammalian species. PMID:27041356

  10. Intrauterine embryo transfer with canine embryos cryopreserved by the slow freezing and the Cryotop method.

    PubMed

    Hori, Tatsuya; Ushijima, Hitoshi; Kimura, Taku; Kobayashi, Masanori; Kawakami, Eiichi; Tsutsui, Toshihiko

    2016-08-01

    Canine embryos (8-cell to blastocyst stages) frozen-thawed using the slow-freezing method with glycerol (four recipients) or dimethyl sulfoxide (three recipients) as a cryoprotectant and vitrified-warmed using the Cryotop method (five recipients) were surgically transferred into the unilateral uterine horn of recipient bitches. As a result, the morphology of embryos frozen-thawed using the slow-freezing method was judged to be normal, but no conception occurred in any of the recipient bitches. Two of the five bitches that received transferred embryos (morula to early blastocyst stages) vitrified-warmed using the Cryotop method became pregnant and produced normal pups (1/9 embryos, 11.1% and 1/6 embryos, 17.0%). It was concluded that the Cryotop method was more appropriate for canine embryo cryopreservation than the slow-freezing method, which is used for the cryopreservation of embryos of other mammalian species.

  11. Physiological and molecular determinants of embryo implantation

    PubMed Central

    Zhang, Shuang; Lin, Haiyan; Kong, Shuangbo; Wang, Shumin; Wang, Hongmei; Wang, Haibin; Armant, D. Randall

    2014-01-01

    Embryo implantation involves the intimate interaction between an implantation-competent blastocyst and a receptive uterus, which occurs in a limited time period known as the window of implantation. Emerging evidence shows that defects originating during embryo implantation induce ripple effects with adverse consequences on later gestation events, highlighting the significance of this event for pregnancy success. Although a multitude of cellular events and molecular pathways involved in embryo-uterine crosstalk during implantation have been identified through gene expression studies and genetically engineered mouse models, a comprehensive understanding of the nature of embryo implantation is still missing. This review focuses on recent progress with particular attention to physiological and molecular determinants of blastocyst activation, uterine receptivity, blastocyst attachment and uterine decidualization. A better understanding of underlying mechanisms governing embryo implantation should generate new strategies to rectify implantation failure and improve pregnancy rates in women. PMID:23290997

  12. Embryo transfer in domestic South American camelids.

    PubMed

    Sumar, Julio B

    2013-01-10

    Intraspecific and interspecific embryo transfer in domestic South American camelids is developing into a well-established technique. Reports reveal many benefits of using reproductive biotechnologies to allow rapid propagation of alpacas and llamas of high genetic merit (e.g., high fiber quality, preserve color variation). The objective of this review is to provide up-to-date information about embryo transfer in domestic South American camelids. Specific information is provided on criteria for male selection, donor and recipient synchronization, the practice of single- vs. super-ovulation protocols, embryo recovery and transfer techniques, advances in cryopreservation of embryos, results of intra- and inter-specific transfer, and the future of the embryo transfer in domestic South American camelids.

  13. Undernutrition affects embryo quality of superovulated ewes.

    PubMed

    Abecia, J A; Forcada, F; Palacín, I; Sánchez-Prieto, L; Sosa, C; Fernández-Foren, A; Meikle, A

    2015-02-01

    To determine the effect of undernutrition on embryo production and quality in superovulated sheep, 45 ewes were allocated into two groups to be fed diets that provided 1.5 (control, C; n = 20) or 0.5 (low nutrition, L; n = 25) times daily requirements for maintenance, from oestrous synchronization with intravaginal sponges to embryo collection. Embryos were collected 7 days after the onset of oestrus (day 0). Low nutrition resulted in lower live weight and body condition at embryo collection (P < 0.05). Diet (P < 0.01) and day of sampling (P < 0.001) significantly affected plasma non-esterified fatty acid (NEFA) and insulin concentrations. Plasma leptin concentrations decreased on day 7 only in L ewes. A significant effect of dietary treatment (P < 0.05) and day (P < 0.0001) was observed on plasma insulin-like growth factor (IGF)-I concentrations. The number of recovered oocytes and embryos did not differ between the groups (L: 15.4 ± 0.4; C: 12.4 ± 0.4). Recovery rate was lower (P < 0.05) in the L (60%) than in the C group (73%). The total number of embryos and number of viable-transferable embryos (5.0 ± 0.3 and 3.4 ± 0.3 embryos, respectively) of the L group were lower (P < 0.1) when compared with controls (8.4 ± 0.4 and 6.2 ± 0.4 embryos, respectively). Undernutrition during the period of superovulation and early embryonic development reduced total and viable number of embryos. These effects might be mediated by disruption of endocrine homeostasis, oviduct environment and/or oocyte quality.

  14. Refrigeration of rainbow trout gametes and embryos.

    PubMed

    Babiak, Igor; Dabrowski, Konrad

    2003-12-01

    Prolonged access to early embryos composed of undifferentiated, totipotent blastomeres is desirable in situations when multiple collections of gametes are not possible. The objective of the present study is to examine whether the refrigeration of rainbow trout Oncorhynchus mykiss gametes and early embryos would be a suitable, reliable, and efficient tool for prolonging the availability of early developmental stages up to the advanced blastula stage. The study was conducted continuously during fall, winter, and spring spawning seasons. In all, more than 500 experimental variants were performed involving individual samples from 26 females and 33 males derived from three strains. These strains represented three possible circumstances. In optimal one, gametes from good quality donors were obtained soon after ovulation. In the two non-optimal sources, either donors were of poor genetic quality or gametes were collected from a distant location and transported as unfertilized gametes. A highly significant effect of variability of individual sample quality on efficiency of gamete and embryo refrigeration was revealed. The source of gametes significantly affected viability of refrigerated oocytes and embryos, but not spermatozoa. On average, oocytes from optimal source retained full fertilization viability for seven days of chilled storage, significantly longer than from non-optimal sources. Spermatozoa, regardless of storage method, retained full fertilization ability for the first week of storage. Refrigeration of embryos at 1.4+/-0.4 degrees C significantly slowed the development. Two- week-old embryos were still in blastula stage. Average survival rate of embryos refrigerated for 10 days and then transferred to regular incubation temperatures of 9-14 degrees C was 92% in optimal and 51 and 71% in non-optimal source variants. No effect of gamete and embryo refrigeration on the occurrence of developmental abnormalities was observed. Cumulative refrigeration of oocytes and

  15. Debate in embryo donation: embryo donation or both-gamete donation?

    PubMed

    Samani, Reza Omani; Moalem, Mohammad Reza Rezania; Merghati, Seyed Taha; Alizadeh, Leila

    2009-01-01

    So far, more than 2 million babies have been born worldwide through assisted reproduction technologies. For many couples, there is no treatment except by involving a third party. Recently, embryo donation law has been approved by Iran's parliament and now it is legal in Iran. But there is a misunderstanding regarding the source of embryos: they can be obtained from surplus frozen embryos of infertile couples or embryos can be made from donated spermatozoa and eggs from fertile married couples. Here in this paper we discuss ethical, religious and legal aspects of these two procedures and present the advantages and disadvantages of them. Meanwhile, the new term 'both-gamete donation' was defined for the procedure that is practised here instead of 'embryo donation'. In conclusion we can say: (i) Iranian law means only embryo donation and covers only surplus embryos from other infertile couples and not both-gamete donation; (ii) as gamete donation is practised in Iran upon decrees of clergy leaders, we have no law or legislation against both-gamete donation; (iii) there are many ethical, legal and religious questions about both-gamete donation to be answered; (iv) ethical and religious questions are fewer concerning embryo donation compared with both-gamete donation; and (v) embryo sharing is a good way for donation of fresh embryos.

  16. Characterization of embryo-specific genes

    SciTech Connect

    Sung, Z.R.

    1988-01-01

    The objective of the proposed research is to characterize the structure and function of a set of genes whose expression is regulated in embryo development, and that are not expressed in mature tissues -- the embryogenic genes. In order to isolate these genes, we immunized a rabbit with total extracts of somatic embryos of carrot, and enriched the anti-embryo antiserum for antibodies reacting with extracts of carrot somatic embryos. Using this enriched antiserum, we screened a lambda gt11 cDNA library constructed from embryo poly A{sup +} RNA, and isolated 10 cDNA clones that detect embryogenic mRNAs. Monospecific antibodies have been purified for proteins corresponding to each cDNA sequence. Four cDNA clones were further characterized in terms of the expression of their corresponding mRNA and protein in somatic embryos of carrot. In some cases, comparable gene sequences or products have been detected in somatic and zygotic embryos of other plant species. The characteristics of these 4 cDNA clones -- clone Nos. 8, 59, and 66 -- are described in this report. 3 figs.

  17. Glassfrog embryos hatch early after parental desertion

    PubMed Central

    Delia, Jesse R. J.; Ramírez-Bautista, Aurelio; Summers, Kyle

    2014-01-01

    Both parental care and hatching plasticity can improve embryo survival. Research has found that parents can alter hatching time owing to a direct effect of care on embryogenesis or via forms of care that cue the hatching process. Because parental care alters conditions critical for offspring development, hatching plasticity could allow embryos to exploit variation in parental behaviour. However, this interaction of parental care and hatching plasticity remains largely unexplored. We tested the hypothesis that embryos hatch early to cope with paternal abandonment in the glassfrog Hyalinobatrachium fleischmanni (Centrolenidae). We conducted male-removal experiments in a wild population, and examined embryos' response to conditions with and without fathers. Embryos hatched early when abandoned, but extended development in the egg stage when fathers continued care. Paternal care had no effect on developmental rate. Rather, hatching plasticity was due to embryos actively hatching at different developmental stages, probably in response to deteriorating conditions without fathers. Our experimental results are supported by a significant correlation between the natural timing of abandonment and hatching in an unmanipulated population. This study demonstrates that embryos can respond to conditions resulting from parental abandonment, and provides insights into how variation in care can affect selection on egg-stage adaptations. PMID:24789892

  18. Toxicity of chlorine to zebrafish embryos.

    PubMed

    Kent, Michael L; Buchner, Cari; Barton, Carrie; Tanguay, Robert L

    2014-01-16

    Surface disinfection of fertilized fish eggs is widely used in aquaculture to reduce extraovum pathogens that may be released from brood fish during spawning, and this is routinely used in zebrafish Danio rerio research laboratories. Most laboratories use approximately 25 to 50 ppm unbuffered chlorine solution for 5 to 10 min. Treatment of embryos with chlorine has significant germicidal effects for many Gram-negative bacteria, viruses, and trophozoite stages of protozoa, but is less effective against cyst or spore stages of protozoa and certain Mycobacterium spp. Therefore, we evaluated the toxicity of unbuffered and buffered chlorine solutions to embryos exposed at 6 or 24 h post-fertilization (hpf) to determine whether higher concentrations can be used for treating zebrafish embryos. Most of our experiments entailed using an outbred line (5D), with both mortality and malformations as endpoints. We found that 6 hpf embryos consistently were more resistant than 24 hpf embryos to the toxic effects of chlorine. Chlorine is more toxic and germicidal at lower pH, and chlorine causes elevated pH. Consistent with this, we found that unbuffered chlorine solutions (pH ca. 8-9) were less toxic at corresponding concentrations than solutions buffered to pH 7. Based on our findings here, we recommend treating 6 hpf embryos for 10 min and 24 hpf embryos for 5 min with unbuffered chlorine solution at 100 ppm.

  19. Glassfrog embryos hatch early after parental desertion.

    PubMed

    Delia, Jesse R J; Ramírez-Bautista, Aurelio; Summers, Kyle

    2014-06-22

    Both parental care and hatching plasticity can improve embryo survival. Research has found that parents can alter hatching time owing to a direct effect of care on embryogenesis or via forms of care that cue the hatching process. Because parental care alters conditions critical for offspring development, hatching plasticity could allow embryos to exploit variation in parental behaviour. However, this interaction of parental care and hatching plasticity remains largely unexplored. We tested the hypothesis that embryos hatch early to cope with paternal abandonment in the glassfrog Hyalinobatrachium fleischmanni (Centrolenidae). We conducted male-removal experiments in a wild population, and examined embryos' response to conditions with and without fathers. Embryos hatched early when abandoned, but extended development in the egg stage when fathers continued care. Paternal care had no effect on developmental rate. Rather, hatching plasticity was due to embryos actively hatching at different developmental stages, probably in response to deteriorating conditions without fathers. Our experimental results are supported by a significant correlation between the natural timing of abandonment and hatching in an unmanipulated population. This study demonstrates that embryos can respond to conditions resulting from parental abandonment, and provides insights into how variation in care can affect selection on egg-stage adaptations.

  20. Effect of zidovudine on preimplantation murine embryos.

    PubMed Central

    Toltzis, P; Mourton, T; Magnuson, T

    1993-01-01

    It previously has been demonstrated that zidovudine (AZT) is lethal to early murine embryos. The effect of the drug on pre- and postimplantation embryos was examined to delineate the timing of this toxicity and to investigate its possible mechanisms. Embryos exposed in the whole mouse during preblastocyst development were unable to proceed beyond the blastocyst stage. Similarly, when two-cell embryos harvested from unexposed females were exposed to low-concentration (1 microM) AZT in vitro over 24 h, development beyond the blastocyst stage was inhibited. In contrast, drug exposure during in vitro blastocyst and postblastocyst development resulted in little or no morphologic toxicity. Further investigation revealed that preblastocyst AZT exposure resulted in the development of blastocysts with significantly lower cell numbers than control embryos. While embryonic exposure to AZT at the blastocyst and postblastocyst stages also resulted in retarded cell division, the effects were milder than those recorded after preblastocyst exposure. These data demonstrate that the critical period of AZT toxicity toward murine embryos is between ovulation and implantation and indicate that AZT directly suppresses cell division in the preimplantation embryo. PMID:8215271

  1. Skeletogenesis in sea urchin interordinal hybrid embryos.

    PubMed

    Brandhorst, B P; Davenport, R

    2001-07-01

    Reciprocal interordinal crosses were made between the sea urchins Strongylocentrotus purpuratus and Lytechinus pictus. Previous research indicated that the expression of many L. pictus genes is reduced in the hybrid embryos. The S. purpuratus gene encoding the spicule matrix protein SM50 and the L. pictus gene encoding its orthologue LSM34 were both expressed at normal levels per gene copy in hybrid embryos, and in about 32 skeletogenic primary mesenchyme cells (PMCs) in hybrid and natural gastrulae. In many embryos of all crosses, 16 PMCs initially ingressed, while 32-64 PMCs were present in gastrulae. The skeletal spicules of most hybrid plutei were predominantly like those of S. purpuratus, consistent with the predominance of expression of S. purpuratus genes in hybrid embryos. The spicules of some hybrid plutei showed features characteristic of L. pictus, such as recurrent rods, branched body rod tips, or convergent ventral transverse rods; a few hybrid spicules were predominantly like those of L. pictus. Based on our observations and the literature, we propose the following. Cues from the ectodermal epithelium position the PMCs as they elaborate the initial triradiate spicules. Their orientation and outgrowth appears to be responsible for the convergence of the tips of body rods in most S. purpuratus and hybrid embryos, unlike in most L. pictus embryos. Variations among hybrid and natural embryos in skeletal branching pattern reflect differences in interpretation by PMCs of patterning cues produced by the ectodermal epithelium that probably have similar spatial distributions in the two species.

  2. Deep cytoplasmic rearrangements in ventralized Xenopus embryos

    NASA Technical Reports Server (NTRS)

    Brown, E. E.; Denegre, J. M.; Danilchik, M. V.

    1993-01-01

    Following fertilization in Xenopus, dramatic rearrangements of the egg cytoplasm relocalize maternally synthesized egg components. During the first cell cycle the vegetal yolk mass rotates relative to the egg surface, toward the sperm entry point (SEP) (J. P. Vincent, G. F. Oster, and J. C. Gerhart, 1986, Dev. Biol. 113, 484-500), while concomitant deep cytoplasmic rearrangements occur in the animal hemisphere (M. V. Danilchik and J. M. Denegre, 1991, Development 111, 845-856). In this paper we examine the role of vegetal yolk mass rotation in producing the animal cytoplasmic rearrangements. We inhibited rotation by uv-irradiating embryos during the first cell cycle, a treatment that yields an extremely ventralized phenotype. Both uv-irradiated embryos and unirradiated control embryos show cytoplasmic rearrangements in the animal hemisphere during the first cell cycle. Cytoplasmic rearrangements on the SEP side of the embryo associated with the path of the sperm pronucleus, plus a swirl on the anti-SEP (dorsal) side, are seen, whether or not yolk mass rotation has occurred. This result suggests a role for the expanding sperm aster in directing animal hemisphere cytoplasmic movements. In unirradiated control embryos the anti-SEP (dorsal) swirl is larger than that in uv-irradiated embryos and often extends into the vegetal hemisphere, consistent with the animal cytoplasm having been pulled dorsally and vegetally by the sliding vegetal yolk mass. Thus the yolk mass rotation may normally enhance the dorsalward cytoplasmic movement, begun by the sperm aster, enough to induce normal axis formation. We extended our observations of unirradiated control and uv-irradiated embryos through early cleavages. The vegetal extent of the anti-SEP (dorsal) swirl pattern seen in control embryos persists through the early cleavage period, such that labeled animal cytoplasm extends deep into dorsal third-tier blastomeres at the 32-cell stage. Significantly, in uv-irradiated embryos

  3. Piglets born after intrauterine laparoscopic embryo transfer.

    PubMed

    Wieczorek, J; Koseniuk, J; Mandryk, I; Poniedziałek-Kempny, K

    2015-01-01

    The aim of the study was the preliminary development of laparoscopic transfer of embryos to the uterus in the pig, which can become the alternative for more invasive surgical methods. We proposed the original method of embryo transfer. Donors (n = 40) and recipients (n = 15) of embryos were sows of age of 6-8 months. The estrus cycle of both recipients and donors was routinely synchronized. The experimental animals were divided into two groups. In the first group (10 donors and 3 recipients) embryos were transplanted according to the method described earlier and in the second group (30 donors and 12 recipients) embryos were transplanted according to our own proposed method. As the control group, we used 16 sows after insemination (AI). In animals from both experimental groups pregnancy was diagnosed between 28-31 day after transplantation and in the control group between 28-31 day after insemination. All animals were observed during pregnancy and weaning period in pig farm. Embryos at the development stage of 2-4 cell were obtained surgically and cultured in vitro for 4 days. Obtained blastocysts were transferred to donors. The original set of catheters for blastocysts transfer to pig uterus was constructed. Three trocars were placed in abdominal cavity for inserting endoscope and 2 grasps for uterus stabilization. After uterus stabilization, the slide was inserted into abdomen which was used for putting the needle to puncture uterus. Through this needle catheter with embryos was inserted into the uterus cavity. Embryos were placed by injection into lumen of the one uterine horn. From 12 recipients pregnancy was diagnosed in 6 recipients. From 6 litters, 57 piglets were born. We weaned 41 piglets (71.9%). In our study we obtained 50% efficacy, with the mean number of 9.5 alive piglets in litter and 6.8 weaned piglets. The efficacy of developed method of laparoscopic transfer of porcine embryos allows it to be used in routine practice.

  4. Somatic embryos from callus of Sequoia sempervirens.

    PubMed

    Bourgkard, F; Favre, J M

    1988-10-01

    Compact calli with a potential for somatic embryogenesis were obtained from complete or split mature zygotic embryos or from cotyledons and hypocotyls of in vitro grown seedlings of Sequoia sempervirens. Somatic embryos which showed a typical bipolar structure, were formed together with adventitious buds. When placed on filter paper supports they developed into complete plantlets. Of the various combinations tested, culture medium adapted from Murashige and Skoog mineral solution complemented with 6-benzylaminopurine (2 μM), kinetin (2 μM) and 2,4-dichlorophenoxyacetic acid (2.5 μM) was established as the optimal for somatic embryo production.

  5. Moral qualms, future persons, and embryo research.

    PubMed

    Shaw, David Martin

    2008-05-01

    Many people have moral qualms about embryo research, feeling that embryos must deserve some kind of protection, if not so much as is afforded to persons. This paper will show that these qualms serve to camouflage motives that are really prudential, at the cost of also obscuring the real ethical issues at play in the debate concerning embryo research and therapeutic cloning. This in turn leads to fallacious use of the Actions/Omissions Distinction and ultimately neglects the duties that we have towards future persons.

  6. Biofluid Flow Simulations of Embryo Transfer

    NASA Astrophysics Data System (ADS)

    Shi, W. P.; Ding, D. L.

    2011-09-01

    The investigation of the fluid flow for embryo transfer (ET) procedure may find the way to increase the success rate of the assisted reproductive technologies. In this paper, the transferred liquid flow in the uterine cavity during ET procedure is simulated by a two dimensional multiphase flow model, and the discrete phase model is adopted to trace the embryo motion. Through the investigation on the transferred liquid outline and the track of each embryo in ET cases with different parameters, we summarize the effect of transferred liquid viscosity and distance between catheter tip and uterine fundus. According to the numerical results, we recommend the optimizing standard to perform the ET procedure.

  7. Association between Number of Formed Embryos, Embryo Morphology and Clinical Pregnancy Rate after Intracytoplasmic Sperm Injection.

    PubMed

    Luz, Caroline Mantovani da; Giorgi, Vanessa Silvestre Innocenti; Coelho Neto, Marcela Alencar; Martins, Wellington de Paula; Ferriani, Rui Alberto; Navarro, Paula Andrea

    2016-09-01

    Introduction Infertility has a high prevalence in the general population, affecting ∼ 5 to 15% of couples in reproductive age. The assisted reproduction techniques (ART) include in vitro manipulation of gametes and embryos and are an important treatment indicated to these couples. It is well accepted that the implantation rate is positively influenced by the morphology of transferred embryos. However, we question if, apart from the assessment of embryo morphology, the number of produced embryos per cycle is also related to pregnancy rates in the first fresh transfer cycle. Purpose To evaluate the clinical pregnancy rate according to the number of formed embryos and the transfer of top quality embryos (TQEs). Methods In a retrospective cohort study, between January 2011 and December 2012, we evaluated women who underwent intracytoplasmic sperm injection (ICSI), aged < 40 years, and with at least 1 formed embryo fresh transferred in cleavage stage. These women were stratified into 3 groups according to the number of formed embryos (1 embryo, 2-3 and ≥ 4 embryos). Each group was divided into 2 subgroups according to the presence or not of at least 1 transferred TQE (1 with TQE; 1 without TQE; 2-3 with TQE, 2-3 without TQE; ≥ 4 with TQE; ≥ 4 without TQE). The clinical pregnancy rates were compared in each subgroup based on the presence or absence of at least one transferred TQE. Results During the study period, 636 women had at least one embryo to be transferred in the first fresh cycle (17.8% had 1 formed embryo [32.7% with TQE versus 67.3% without TQE], 42.1% of women had 2-3 formed embryos [55.6% with TQE versus 44.4% without TQE], and 40.1% of patients had ≥ 4 formed embryos [73.7% with TQE versus 26.3% without TQE]). The clinical pregnancy rate was significantly higher in the subgroup with ≥ 4 formed embryos with at least 1 transfered TQE (45.2%) compared with the subgroup without TQE (28.4%). Conclusions Having at

  8. The ART of studying early embryo development: progress and challenges in ruminant embryo culture.

    PubMed

    Lonergan, Pat; Fair, Trudee

    2014-01-01

    The study of preimplantation mammalian embryo development is challenging due to difficulties in accessing in vivo-derived embryos in large numbers at the early stages and the inability to culture embryos in vitro much beyond the blastocyst stage. Nonetheless, embryos exhibit an amazing plasticity and tolerance when it comes to adapting to the environment in which they are cultured. They are capable of developing in media ranging in composition from simple balanced salt solutions to complex systems involving serum and somatic cells. At least a proportion of the blastocysts that develop in culture are developmentally competent as evidenced by the fact that live offspring have resulted following transfer. However, several studies using animal models have shown that such embryos are sensitive to environmental conditions that can affect future pre- and post-natal growth and developmental potential. This review summarises some key aspects of early embryo development and the approaches taken to study this important window in early life.

  9. Multiple-embryo transfer for studying very early maternal-embryo interactions in cattle.

    PubMed

    Gómez, E; Muñoz, M

    2015-08-01

    In the present paper, we highlight the need to study very early maternal-embryo interactions and discuss how these interactions can be addressed. Bovine species normally carry one or, less frequently, two embryos to term; there are very rare cases of triplets or higher-order multiple pregnancies in which all the offspring are born alive. Multiple-embryo transfer (MET) in cattle allows for the detection of endometrial responses in scenarios where single-embryo transfer would not. Although MET is non-physiological, the present study shows that at the very early embryonic stages, a uterus carrying zona-enclosed embryos does not exhibit non-physiological reactions. On the contrary, MET should be considered the sum of multiple individual effects triggered by developing embryos. We provide arguments to support our hypothesis that describe a rationale for current work with MET, and we discuss alternative hypotheses. Using cattle as a model, we describe how technical approaches to analyzing zona-enclosed early embryo-maternal interactions (i.e., transcriptomics, proteomics, and endometrial cell culture) can help identify molecular changes that may be difficult to observe when only a single embryo is present. We conclude that MET can be used for studying very early maternal-embryo interactions in vivo in monotocous species. Free Spanish abstract: A Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/150/2/R35/suppl/DC1.

  10. The embryo's one-sided genes.

    PubMed

    King; Brown

    1995-12-01

    The origin of left-right asymmetry during vertebrate embryogenesis has long been a puzzle; now, for the first time, genes have been identified that are expressed with left-right asymmetric patterns in early embryos.

  11. Steroidal alkaloid toxicity to fish embryos.

    PubMed

    Crawford, L; Kocan, R M

    1993-02-01

    Embryos of two species of fish were evaluated for their suitability as model systems for steroidal alkaloid toxicity, the Japanese rice fish, medaka (Oryzius latipes) and the rainbow trout (Oncorhynchus mykiss). Additionally, the equine neurotoxic sesquiterpene lactone repin, was also tested. A PROBIT program was used to evaluate the EC1, EC50 and EC99 as well as the associated confidence limits. The steroidal alkaloids tested were the Solanum potato glycoalkaloids alpha-chaconine, alpha-solanine, the aglyclones solanidine and solasodine and the Veratrum alkaloid, jervine. Embryo mortality, likely due to structural or functional abnormalities in the early development stages of the embryo, were the only response observed in both species. The rainbow trout exhibited a toxic response to chaconine, solasidine, repin and solanine but the medaka embryos were only affected by the compounds, chaconine and solanine. Rainbow trout may indeed serve as a good lower vertebrate model for studying the toxicity of steroidal alkaloids.

  12. Valuing embryos as both commodities and singularities.

    PubMed

    Legge, Michael; Fitzgerald, Ruth

    2016-03-11

    An argument put forward against gamete and embryo donation, sale and research, is that to do so would treat the gametes or embryos as objects with no intrinsic value as human. Instead, gametes and embryos created and used for donation, sale or research, can be considered more like a commodity created and traded for economic exchange--something that is valuable only for the amount of money or other goods and services that others are willing to exchange. While Kant asserts that humans have dignity rather than object worth, the provision of human gametes and embryos are progressively becoming utilities for resolving childlessness and for certain research investigations. In this paper we discuss the commodity market and the relationship to human reproduction material.

  13. Media composition: macromolecules and embryo growth.

    PubMed

    Meintjes, Marius

    2012-01-01

    Most embryo culture media are still supplemented with proteins rather than with nonprotein macromolecules or recombinant protein products. HSA is probably the most common supplement followed by globulin-enriched preparations. Serum supplementation and Co-Culture of embryos belong to the past. Defined nonprotein or recombinant protein supplements are becoming a viable alternative during gamete and embryo manipulation procedures. Biological protein supplements are still preferred for any extended period of embryo culture. Understanding the goals and purpose of supplemented macromolecules in embryo culture media during each step of the laboratory IVF process should assist us in choosing the safest and most consistent macromolecule for each step, but also selecting a product that has the capability of delivering the best clinical outcome. Each batch of biological protein supplement is unique, even if supplied by the same manufacturer. Each lot of protein supplement typically contains many lot-specific, potentially harmful, and unintended hormone and protein contaminants. Macromolecular embryo culture medium supplements should be identified as one of the highest risk factors in an IVF laboratory that may contribute towards clinical compromise. All efforts should be made to use a proven batch of supplement for as long as the expiration date will allow. The beneficial effect of more complex protein supplements is evident after the activation of the embryonic genome and probably due to the presence of growth factors. Lower live-birth rates due to suboptimum protein supplementation may be a direct result of the preferential loss of female embryos. When deciding on a culture system, thought should be given specifically to the interaction between the culture medium and the macromolecular supplement. Ready-to-use pre-supplemented culture media may be advisable over a more complex product if a comprehensive macromolecular quality management program is not feasible. However, the

  14. RNA Interference in Chicken Embryos

    NASA Astrophysics Data System (ADS)

    van Hateren, Nick J.; Jones, Rachel S.; Wilson, Stuart A.

    The chicken has played an important role in biological discoveries since the 17th century (Stern, 2005). Many investigations into vertebrate development have utilized the chicken due to the accessibility of the chick embryo and its ease of manipulation (Brown et al., 2003). However, the lack of genetic resources has often handicapped these studies and so the chick is frequently overlooked as a model organism for the analysis of vertebrate gene function in favor of mice or zebrafish. In the past six years this situation has altered dramatically with the generation of over half a million expressed sequence tags and >20,000 fully sequenced chicken cDNAs (Boardman et al. 2002; Caldwell et al., 2005; Hubbard et al., 2005) together with a 6X coverage genome sequence (Hillier et al., 2004). These resources have created a comprehensive catalogue of chicken genes with readily accessible cDNA and EST resources available via ARK-GENOMICS (www.ark-genomics.org) for the functional analysis of vertebrate gene function.

  15. Cryopreservation of Arachis pintoi (leguminosae) somatic embryos.

    PubMed

    Rey, H Y; Faloci, M; Medina, R; Dolce, N; Engelmann, F; Mroginski, L

    2013-01-01

    In this study, we successfully cryopreserved cotyledonary somatic embryos of diploid and triploid Arachis pintoi cytotypes using the encapsulation-dehydration technique. The highest survival rates were obtained when somatic embryos were encapsulated in calcium alginate beads and precultured in agitated (80 rpm) liquid establishment medium (EM) with daily increasing sucrose concentration (0.50, 0.75, and 1.0 M). The encapsulated somatic embryos were then dehydrated with silica gel for 5 h to 20% moisture content (fresh weight basis) and cooled either rapidly (direct immersion in liquid nitrogen, LN) or slowly (1 degree C per min from 25 degree C to -30 degree C followed by immersion in LN). Beads were kept in LN for a minimum of 1 h and then were rapidly rewarmed in a 30 degree C water-bath for 2 min. Finally, encapsulated somatic embryos were post-cultured in agitated (80 rpm) liquid EM with daily decreasing sucrose concentration (0.75 and 0.5 M) and transferred to solidified EM. Using this protocol, we obtained 26% and 30% plant regeneration from cryopreserved somatic embryos of diploid and triploid cytotypes. No morphological abnormalities were observed in any of the plants regenerated from cryopreserved embryos and their genetic stability was confirmed with 10 isozyme systems and nine RAPD profiles.

  16. In vitro bovine embryo production in a synthetic medium: embryo development, cryosurvival, and establishment of pregnancy.

    PubMed

    Moreno, D; Neira, A; Dubreil, L; Liegeois, L; Destrumelle, S; Michaud, S; Thorin, C; Briand-Amirat, L; Bencharif, D; Tainturier, D

    2015-10-15

    The aim of this study was to develop an in vitro embryo culture medium without either fetal calf serum or BSA, using various growth factors and cytokines (GFs-CYKs; IGF-I, IGF-II, bFGF, LIF, GM-CSF, TGF-β1, and PDGF-BB), and other molecules with surfactant and embryotrophic properties, such as recombinant albumin (RA) and hyaluronan (HA). The first part of the study was dedicated to define the best combination of GFs-CYKs + RA + HA for optimal embryonic development. Next, we compared development rates and embryo quality (inner cell mass [ICM]-to-total cell number [TCN] ratio), and postthaw survival and hatching rates using this synthetic medium (T1) and a control medium: synthetic oviduct fluid + BSA + ITS (insulin, transferrin, and selenium). The blastocyst rates were significantly higher with T1 than those with the control at 7 and 8 days after fertilization. There was no significant difference in TCN or the ICM/TCN ratio between the two treatments. Survival and hatching rates 48 hours after thawing were similar for both treatments. Finally, nine embryo transfers were conducted using fresh and previously frozen Day-7 blastocysts to evaluate the in vivo viability of embryos produced in this synthetic medium; four gestations were obtained from six fresh embryos and one gestation from three frozen embryos. In conclusion, the fetal calf serum and BSA-free medium, supplemented with GFs-CYKs + RA + HA, improved embryo development and gave comparable ICM/TCN ratios and postthaw survival rates to the control with BSA. Fresh and frozen embryos produced in this medium are viable for embryo transfer. This fully synthetic method of embryo culture is a useful means of reducing the risk of disease transmission via embryo transfer.

  17. Embryo quality and transcervical technique are not the limiting factors in donkey embryo transfer outcome.

    PubMed

    Panzani, D; Rota, A; Crisci, A; Kindahl, H; Govoni, N; Camillo, F

    2012-02-01

    Embryo transfer (ET) in the donkey resulted in a very low recipient pregnancy rates. The aim of these studies was to investigate if nonsurgical transfer techniques or donkey embryo quality affect donkey recipient pregnancy failure. In Study 1, the impact of transfer technique was investigated by evaluating if cervical catheterization is associated with prostaglandin release and suppression of luteal function and if donkey recipients would become pregnant after nonsurgical transfer of horse embryos. Four jennies, from 5 to 8 d after ovulation, were submitted to a sham transcervical ET and to evaluation of PGFM and progesterone plasma concentrations. Five 8 d horse embryos were nonsurgically transferred into synchronized donkey recipients (HD). Cervical stimulation caused a transient PGF(2α) release in two of four jennies in the absence of a significant decrease in progesterone plasma concentration. All transferred horse embryos resulted in pregnancies in the jenny recipients. In Study 2, donkey embryo viability was investigated by 1.2 meters, 6-diamidino-2-phenylindole (DAPI) staining of 10 embryos and by the transfer of 6 and 12 donkey embryos in synchronized mare (DH) and donkey (DD) recipients, respectively, of known fertility. The estimated proportion of dead cells in DAPI stained embryos was 0.9% (range 0-3.9%) and below what is considered normal (20%) for horse embryos. Three of six and six of 12 of the DH and DD ETs, respectively resulted in pregnancies at 14 and 25 d (50%), a higher pregnancy rate than previously reported after DD ET. The overall results of this study suggest that the transcervical technique for ET and donkey embryo viability are not the reasons for the low pregnancy rates that have previously been described in donkey recipients, and that nonsurgical ET in donkeys can result in acceptable results.

  18. Miniaturized embryo array for automated trapping, immobilization and microperfusion of zebrafish embryos.

    PubMed

    Akagi, Jin; Khoshmanesh, Khashayar; Evans, Barbara; Hall, Chris J; Crosier, Kathryn E; Cooper, Jonathan M; Crosier, Philip S; Wlodkowic, Donald

    2012-01-01

    Zebrafish (Danio rerio) has recently emerged as a powerful experimental model in drug discovery and environmental toxicology. Drug discovery screens performed on zebrafish embryos mirror with a high level of accuracy the tests usually performed on mammalian animal models, and fish embryo toxicity assay (FET) is one of the most promising alternative approaches to acute ecotoxicity testing with adult fish. Notwithstanding this, automated in-situ analysis of zebrafish embryos is still deeply in its infancy. This is mostly due to the inherent limitations of conventional techniques and the fact that metazoan organisms are not easily susceptible to laboratory automation. In this work, we describe the development of an innovative miniaturized chip-based device for the in-situ analysis of zebrafish embryos. We present evidence that automatic, hydrodynamic positioning, trapping and long-term immobilization of single embryos inside the microfluidic chips can be combined with time-lapse imaging to provide real-time developmental analysis. Our platform, fabricated using biocompatible polymer molding technology, enables rapid trapping of embryos in low shear stress zones, uniform drug microperfusion and high-resolution imaging without the need of manual embryo handling at various developmental stages. The device provides a highly controllable fluidic microenvironment and post-analysis eleuthero-embryo stage recovery. Throughout the incubation, the position of individual embryos is registered. Importantly, we also for first time show that microfluidic embryo array technology can be effectively used for the analysis of anti-angiogenic compounds using transgenic zebrafish line (fli1a:EGFP). The work provides a new rationale for rapid and automated manipulation and analysis of developing zebrafish embryos at a large scale.

  19. Miniaturized Embryo Array for Automated Trapping, Immobilization and Microperfusion of Zebrafish Embryos

    PubMed Central

    Akagi, Jin; Khoshmanesh, Khashayar; Evans, Barbara; Hall, Chris J.; Crosier, Kathryn E.; Cooper, Jonathan M.; Crosier, Philip S.; Wlodkowic, Donald

    2012-01-01

    Zebrafish (Danio rerio) has recently emerged as a powerful experimental model in drug discovery and environmental toxicology. Drug discovery screens performed on zebrafish embryos mirror with a high level of accuracy the tests usually performed on mammalian animal models, and fish embryo toxicity assay (FET) is one of the most promising alternative approaches to acute ecotoxicity testing with adult fish. Notwithstanding this, automated in-situ analysis of zebrafish embryos is still deeply in its infancy. This is mostly due to the inherent limitations of conventional techniques and the fact that metazoan organisms are not easily susceptible to laboratory automation. In this work, we describe the development of an innovative miniaturized chip-based device for the in-situ analysis of zebrafish embryos. We present evidence that automatic, hydrodynamic positioning, trapping and long-term immobilization of single embryos inside the microfluidic chips can be combined with time-lapse imaging to provide real-time developmental analysis. Our platform, fabricated using biocompatible polymer molding technology, enables rapid trapping of embryos in low shear stress zones, uniform drug microperfusion and high-resolution imaging without the need of manual embryo handling at various developmental stages. The device provides a highly controllable fluidic microenvironment and post-analysis eleuthero-embryo stage recovery. Throughout the incubation, the position of individual embryos is registered. Importantly, we also for first time show that microfluidic embryo array technology can be effectively used for the analysis of anti-angiogenic compounds using transgenic zebrafish line (fli1a:EGFP). The work provides a new rationale for rapid and automated manipulation and analysis of developing zebrafish embryos at a large scale. PMID:22606275

  20. Meanings of the embryo in Japan: narratives of IVF experience and embryo ownership.

    PubMed

    Kato, Masae; Sleeboom-Faulkner, Margaret

    2011-03-01

    This article explores the sociocultural meanings of the embryo implied in the narratives of 58 women who have undergone in vitro fertilisation in Japan over a period from 2006 to 2008. We argue that a lack of sufficient analysis of the sociocultural meanings of the embryo result in a situation where the use of reproductive technologies in Japan advances without reflecting upon the voices of women and couples that use them. Additionally, we argue that the often-heard view that pre-implantation genetic diagnosis causes less pain to women and couples than selective abortion in which foetuses are discarded, should be reviewed in the light of the new empirical evidence offered in this article. Furthermore, this article shows that the view often expounded by Japanese scientists that in Japan the cultural meanings attached to the embryo are insignificant, is incorrect. Consequently, the argument that Japan has no need for an active national debate on the status of embryos should be questioned. Though agreeing with some feminist views on the embryo debate, this article is also critical of feminist views that discuss embryo donation in terms of the loss of ownership of the embryo and the alienation of the embryo due to commodification.

  1. Vitrification of zebrafish embryo blastomeres in microvolumes.

    PubMed

    Cardona-Costa, J; García-Ximénez, F

    2007-01-01

    Cryopreservation of fish embryos may play an important role in biodiversity preservation and in aquaculture, but it is very difficult. In addition, the cryopreservation of fish embryo blastomeres makes conservation strategies feasible when they are used in germ-line chimaerism, including interspecific chimaerism. Fish embryo blastomere cryopreservation has been achieved by equilibrium procedures, but to our knowledge, no data on vitrification procedures are available. In the present work, zebrafish embryo blastomeres were successfully vitrified in microvolumes: a number of 0.25 microl drops, sufficient to contain all the blastomeres of an embryo at blastula stage (from 1000-cell stage to oblong stage), were placed over a 2.5 cm loop of nylon filament. In this procedure, where intracellular cryoprotectant permeation is not required, blastomeres were exposed to cryoprotectants for a maximum of 25 sec prior vitrification. The assayed cryoprotectants (ethylene glycol, propylene glycol, dimethyl sulphoxide, glycerol and methanol) are all frequently used in fish embryo and blastomere cryopreservation. Methanol was finally rejected because of the excessive concentration required for the vitrification (15M). All other cryoprotectants were prepared (individually) at 5 M in Hanks' buffered salt solution (sigma) plus 20% FBS (vitrification solutions: vs). After direct thawing in Hanks' buffered salt solution plus 20% FBS, acceptable survival rates were obtained with ethylene glycol: 82.8%, propylene glycol: 87.7%, dimethyl sulphoxide: 93.4%, and glycerol: 73.9% (p < 0.05). Dimethyl sulphoxide showed the highest blastomere survival rate and allowed the rescue of as much as 20% of the total blastomeres from each zebrafish blastula embryo.

  2. Embryos, individuals, and persons: an argument against embryo creation and research.

    PubMed

    Tollefsen, C

    2001-01-01

    One strategy for arguing that it should be legally permissible to create human embryos, or to use spare human embryos, for scientific research purposes involves the claim that such embryos cannot be persons because they are not human individuals while twinning may yet take place. Being a human individual is considered to be by most people a necessary condition for being a human person. I argue first that such an argument against the personhood of embryos must be rationally conclusive if their destruction in public places such as laboratories is to be countenanced. I base this argument on a popular understanding of the role that the notion of privacy plays in abortion laws. I then argue that such arguments against personhood are not rationally conclusive. The claim that the early embryos is not a human individual is not nearly as obvious as some assert.

  3. Metabolite profiling of somatic embryos of Cyclamen persicum in comparison to zygotic embryos, endosperm, and testa

    PubMed Central

    Winkelmann, Traud; Ratjens, Svenja; Bartsch, Melanie; Rode, Christina; Niehaus, Karsten; Bednarz, Hanna

    2015-01-01

    Somatic embryogenesis has been shown to be an efficient in vitro plant regeneration system for many crops such as the important ornamental plant Cyclamen persicum, for which this regeneration pathway of somatic embryogenesis is of interest for the vegetative propagation of parental lines as well as elite plants. However, somatic embryogenesis is not commercially used in many crops due to several unsolved problems, such as malformations, asynchronous development, deficiencies in maturation and germination of somatic embryos. In contrast, zygotic embryos in seeds develop and germinate without abnormalities in most cases. Instead of time-consuming and labor-intensive experiments involving tests of different in vitro culture conditions and plant growth regulator supplements, we follow a more directed approach. Zygotic embryos served as a reference and were compared to somatic embryos in metabolomic analyses allowing the future optimization of the in vitro system. The aims of this study were to detect differences in the metabolite profiles of torpedo stage somatic and zygotic embryos of C. persicum. Moreover, major metabolites in endosperm and testa were identified and quantified. Two sets of extracts of two to four biological replicates each were analyzed. In total 52 metabolites were identified and quantified in the different tissues. One of the most significant differences between somatic and zygotic embryos was that the proline concentration in the zygotic embryos was about 40 times higher than that found in somatic embryos. Epicatechin, a scavenger for reactive oxygen species, was found in highest abundance in the testa. Sucrose, the most abundant metabolite was detected in significantly higher concentrations in zygotic embryos. Also, a yet unknown trisaccharide, was significantly enriched in zygotic embryos. PMID:26300898

  4. Biosensors for detecting stress in developing embryos

    NASA Astrophysics Data System (ADS)

    Purdey, Malcolm S.; Saini, Avishkar; McLennan, Hanna J.; Pullen, Benjamin J.; Schartner, Erik P.; Sutton-McDowall, Melanie L.; Thompson, Jeremy G.; Monro, Tanya M.; Nicholls, Stephen J.; Abell, Andrew D.

    2016-12-01

    Reactive Oxygen Species (ROS) cause DNA damage and defective function in sperm and also affects the developmental competence of embryos. It is therefore critical to monitor ROS in sperm, oocytes and developing embryos. In particular, hydrogen peroxide (H2O2) is a ROS important to normal cell function and signalling as well as its role in oxidative stress. Here we report the development of a fluorescent sensor for H2O2 using carboxyperoxyfluor-1 (CPF1) in solution and attached to a glass slide or multi-mode optical fibre. CPF1 increases in fluorescence upon reaction with H2O2 to non-invasively detect H2O2 near developing embryos. These probes are constructed by immobilising CPF1 to the optical fibre tip a polyacrylamide layer. Also reported is a new dual optical fibre sensor for detecting both H2O2 and pH that is functional at biologically concentrations of H2O2 and can sense pH to 0.1 units. This research shows promise for the use of optical fibre sensors for monitoring the health of developing embryos. Furthermore, these sensors are applicable for use beyond embryos such as detecting stress in endothelial cells involved in cardiovascular dysfunction.

  5. ROCK inhibition prevents early mouse embryo development.

    PubMed

    Duan, Xing; Chen, Kun-Lin; Zhang, Yu; Cui, Xiang-Shun; Kim, Nam-Hyung; Sun, Shao-Chen

    2014-08-01

    ROCK is a Rho-GTPase effector that is important for actin assembly and is involved in various cellular functions, including cell contraction, migration, motility, and tumor cell invasion. In this study, we investigated ROCK expression and function during early mouse embryo development. Inhibiting ROCK by Y-27632 treatment at the zygote stage resulted in first cleavage failure, and most embryos failed to develop to the 8-cell stage. When adding Y-27632 at the 8-cell stage, embryos failed to undergo compaction and could not develop into blastocysts. In addition, fluorescence staining intensity analysis indicated that actin expression at blastomere membranes was significantly reduced. After ROCK inhibition, two or more nuclei were observed in a cell, which indicated possible cytokinesis failure. Moreover, after ROCK inhibition with Y-27632, the phosphorylation levels of LIMK1/2, a downstream molecule of ROCK, were decreased at blastomere membranes. Thus, our results showed conserved roles for ROCK in this mammalian embryo model and indicated that a ROCK-LIMK1/2-actin pathway might regulate cleavage and blastocyst formation during early mouse embryo development.

  6. [The embryo, the human and the humanized].

    PubMed

    Roa, A

    1992-03-01

    Since the moment of fecundation the human embryo is endowed with the properties of unity and uniqueness and its existence is therefore inviolable. Disputing arguments against this thesis are analyzed. Recent views of some biologists negate the human character to the embryo since the essence of a human being would be its cultural nature and ability to communicate. However, the embryo contains all the genetic information that will allow him to develop the ability to communicate. Any attempt to separate the 3 moments of time, past present and future is a definitive violation of ethics. A basic foundation of ethics is that present and future are implicit in the past and vice-versa. Finally, the idea that the unwanted child is not a cultural being should be discarded.

  7. Characterization of embryo-specific genes

    SciTech Connect

    Not Available

    1989-01-01

    The objective of the proposed research is to characterize the structure and function of a set of genes whose expression is regulated in embryo development, and that is not expressed in mature tissues -- the embryonic genes. In the last two years, using cDNA clones, we have isolated 22 cDNA clones, and characterized the expression pattern of their corresponding RNA. At least 4 cDNA clones detect RNAs of embryonic genes. These cDNA clones detect RNAs expressed in somatic as well as zygotic embryos of carrot. Using the cDNA clones, we screened the genomic library of carrot embryo DNA, and isolated genomic clones for three genes. The structure and function of two genes DC 8 and DC 59 have been characterized and are reported in this paper.

  8. Embryo aggregation does not improve the development of interspecies somatic cell nuclear transfer embryos in the horse.

    PubMed

    Gambini, Andrés; De Stéfano, Adrián; Jarazo, Javier; Buemo, Carla; Karlanian, Florencia; Salamone, Daniel Felipe

    2016-09-01

    The low efficiency of interspecies somatic cell nuclear transfer (iSCNT) makes it necessary to investigate new strategies to improve embryonic developmental competence. Embryo aggregation has been successfully applied to improve cloning efficiency in mammals, but it remains unclear whether it could also be beneficial for iSCNT. In this study, we first compared the effect of embryo aggregation over in vitro development and blastocyst quality of porcine, bovine, and feline zona-free (ZF) parthenogenetic (PA) embryos to test the effects of embryo aggregation on species that were later used as enucleated oocytes donors in our iSCNT study. We then assessed whether embryo aggregation could improve the in vitro development of ZF equine iSCNT embryos after reconstruction with porcine, bovine, and feline ooplasm. Bovine- and porcine-aggregated PA blastocysts had significantly larger diameters compared with nonaggregated embryos. On the other hand, feline- and bovine-aggregated PA embryos had higher blastocyst cell number. Embryo aggregation of equine-equine SCNT was found to be beneficial for embryo development as we have previously reported, but the aggregation of three ZF reconstructed embryos did not improve embryo developmental rates on iSCNT. In vitro embryo development of nonaggregated iSCNT was predominantly arrested around the stage when transcriptional activation of the embryonic genome is reported to start on the embryo of the donor species. Nevertheless, independent of embryo aggregation, equine blastocyst-like structures could be obtained in our study using domestic feline-enucleated oocytes. Taken together, these results reported that embryo aggregation enhance in vitro PA embryo development and embryo quality but effects vary depending on the species. Embryo aggregation also improves, as expected, the in vitro embryo development of equine-equine SCNT embryos; however, we did not observe positive effects on equine iSCNT embryo development. Among oocytes

  9. Migration and growth of protoplanetary embryos. I. Convergence of embryos in protoplanetary disks

    SciTech Connect

    Zhang, Xiaojia; Lin, Douglas N. C.; Liu, Beibei; Li, Hui

    2014-12-10

    According to the core accretion scenario, planets form in protostellar disks through the condensation of dust, coagulation of planetesimals, and emergence of protoplanetary embryos. At a few AU in a minimum mass nebula, embryos' growth is quenched by dynamical isolation due to the depletion of planetesimals in their feeding zone. However, embryos with masses (M{sub p} ) in the range of a few Earth masses (M {sub ⊕}) migrate toward a transition radius between the inner viscously heated and outer irradiated regions of their natal disk. Their limiting isolation mass increases with the planetesimals surface density. When M{sub p} > 10 M {sub ⊕}, embryos efficiently accrete gas and evolve into cores of gas giants. We use a numerical simulation to show that despite stream line interference, convergent embryos essentially retain the strength of non-interacting embryos' Lindblad and corotation torques by their natal disks. In disks with modest surface density (or equivalently accretion rates), embryos capture each other in their mutual mean motion resonances and form a convoy of super-Earths. In more massive disks, they could overcome these resonant barriers to undergo repeated close encounters, including cohesive collisions that enable the formation of massive cores.

  10. Non-invasive methods for embryo selection

    PubMed Central

    Sallam, HN; Sallam, NH; Sallam, SH

    2016-01-01

    Abstract With the widespread use of assisted reproduction, a simple and practical method for embryo selection is needed to optimize the chances of pregnancy while diminishing the incidence of multiple pregnancy and its accompanying problems. Many non-invasive methods for embryo selection have been proposed and some are more promising than others. This review summarizes these methods and attempts to evaluate them in the light of the best currently available evidence and to find out whether any of them is ripe for replacing or supplementing the time-honored method of morphological assessment. PMID:27909565

  11. Regulatory blueprint for a chordate embryo.

    PubMed

    Imai, Kaoru S; Levine, Michael; Satoh, Nori; Satou, Yutaka

    2006-05-26

    Ciona is an emerging model system for elucidating gene networks in development. Comprehensive in situ hybridization assays have identified 76 regulatory genes with localized expression patterns in the early embryo, at the time when naïve blastomeres are determined to follow specific cell fates. Systematic gene disruption assays provided more than 3000 combinations of gene expression profiles in mutant backgrounds. Deduced gene circuit diagrams describing the formation of larval tissues were computationally visualized. These diagrams constitute a blueprint for the Ciona embryo and provide a foundation for understanding the evolutionary origins of the chordate body plan.

  12. Scientists Create Part-Human, Part-Pig Embryo

    MedlinePlus

    ... 163262.html Scientists Create Part-Human, Part-Pig Embryo One goal of this stem cell research is ... have successfully used human stem cells to create embryos that are part-human, part-pig. Scientists said ...

  13. Cryopreservation of embryos and oocytes in human assisted reproduction.

    PubMed

    Konc, János; Kanyó, Katalin; Kriston, Rita; Somoskői, Bence; Cseh, Sándor

    2014-01-01

    Both sperm and embryo cryopreservation have become routine procedures in human assisted reproduction and oocyte cryopreservation is being introduced into clinical practice and is getting more and more widely used. Embryo cryopreservation has decreased the number of fresh embryo transfers and maximized the effectiveness of the IVF cycle. The data shows that women who had transfers of fresh and frozen embryos obtained 8% additional births by using their cryopreserved embryos. Oocyte cryopreservation offers more advantages compared to embryo freezing, such as fertility preservation in women at risk of losing fertility due to oncological treatment or chronic disease, egg donation, and postponing childbirth, and eliminates religious and/or other ethical, legal, and moral concerns of embryo freezing. In this review, the basic principles, methodology, and practical experiences as well as safety and other aspects concerning slow cooling and ultrarapid cooling (vitrification) of human embryos and oocytes are summarized.

  14. INFECTIVITY OF METARHIZIUM ANISOPLIAE IN GRASS SHRIMP EMBRYOS

    EPA Science Inventory

    Developing embryos of the estuarine grass shrimp, Palaemonetes pugio, were exposed to Metarhizium anisopliae conidiospores. Attachment of conidiospores was often followed by germination and outgrowth on embryo surface. Penetration of the embryonic envelopes by M. anisopliae allow...

  15. Sourcing human embryos for embryonic stem cell lines: problems & perspectives.

    PubMed

    Mehta, Rajvi H

    2014-11-01

    The ability to successfully derive human embryonic stem cells (hESC) lines from human embryos following in vitro fertilization (IVF) opened up a plethora of potential applications of this technique. These cell lines could have been successfully used to increase our understanding of human developmental biology, transplantation medicine and the emerging science of regenerative medicine. The main source for human embryos has been 'discarded' or 'spare' fresh or frozen human embryos following IVF. It is a common practice to stimulate the ovaries of women undergoing any of the assisted reproductive technologies (ART) and retrieve multiple oocytes which subsequently lead to multiple embryos. Of these, only two or maximum of three embryos are transferred while the rest are cryopreserved as per the decision of the couple. in case a couple does not desire to 'cryopreserve' their embryos then all the embryos remaining following embryo transfer can be considered 'spare' or if a couple is no longer in need of the 'cryopreserved' embryos then these also can be considered as 'spare'. But, the question raised by the ethicists is, "what about 'slightly' over-stimulating a woman to get a few extra eggs and embryos? The decision becomes more difficult when it comes to 'discarded' embryos. As of today, the quality of the embryos is primarily assessed based on morphology and the rate of development mainly judged by single point assessment. Despite many criteria described in the literature, the quality assessment is purely subjective. The question that arises is on the decision of 'discarding' embryos. What would be the criteria for discarding embryos and the potential 'use' of ESC derived from the 'abnormal appearing' embryos? This paper discusses some of the newer methods to procure embryos for the derivation of embryonic stem cell lines which will respect the ethical concerns but still provide the source material.

  16. Direct visualization of replication dynamics in early zebrafish embryos.

    PubMed

    Kuriya, Kenji; Higashiyama, Eriko; Avşar-Ban, Eriko; Okochi, Nanami; Hattori, Kaede; Ogata, Shin; Takebayashi, Shin-Ichiro; Ogata, Masato; Tamaru, Yutaka; Okumura, Katsuzumi

    2016-05-01

    We analyzed DNA replication in early zebrafish embryos. The replicating DNA of whole embryos was labeled with the thymidine analog 5-ethynyl-2'-deoxyuridine (EdU), and spatial regulation of replication sites was visualized in single embryo-derived cells. The results unveiled uncharacterized replication dynamics during zebrafish early embryogenesis.

  17. Embryo dignity: the status and juridical protection of the in vitro embryo.

    PubMed

    Raposo, Vera Lúcia; Osuna, Eduardo

    2007-12-01

    In the context of research and reproduction, the status of the human in vitro embryo ranges from being regarded as a person to being regarded as mere property. As regards the first view, one extreme of the spectrum for offering possible legal protection considers that the embryo constitutes a legal person from the moment of conception. For opponents of this view life is a continuum that runs from conception until death. In this process one of the most important stages is birth, the reason being that birth represents the transition between a potential person and a person. The term "embryo" is used to express the being that exists after fusion of the egg and a spermatozoon during the process of embryogenesis until it reaches eight weeks, after which time it is termed a foetus. The embryo's life is recognized as a constitutional value which deserves juridical protection, but not as a person. It only becomes a person with birth.

  18. Culture of Cells from Amphibian Embryos.

    ERIC Educational Resources Information Center

    Stanisstreet, Martin

    1983-01-01

    Describes a method for in vitro culturing of cells from amphibian early embryos. Such cells can be used to demonstrate such properties of eukaryote cells as cell motility, adhesion, differentiation, and cell sorting into tissues. The technique may be extended to investigate other factors. (Author/JN)

  19. Mouse embryo manipulations with OCT guidance

    NASA Astrophysics Data System (ADS)

    Garcia, Monica D.; Syed, Saba H.; Coughlin, Andrew J.; Wang, Shang; West, Jennifer L.; Larin, Kirill V.; Larina, Irina V.

    2014-03-01

    Optical coherence tomography (OCT) is a three-dimensional, non-invasive optical imaging technique that relies on low-coherence interferometry. OCT has the capability of imaging 2 - 3 mm into tissue, which enables imaging of deeper structures within the embryo with a relatively high spatial resolution (2 - 15μm). Within the past decade, OCT has been increasingly used as a live imaging tool for embryonic cardiovascular research in several animal models. Research in our lab has recently shown that OCT can be used in combination with embryo culture for the visualization of early mammalian cardiovascular development (E7.5 - E10.0). Here, we demonstrate that OCT can be used for the guided microinjection of gold-silica nanoshell suspension into the cardiovascular system in live embryos without deleterious effect. This approach shows a promising application for the OCT guided delivery of contrast agents, viral vectors, therapeutic or pharmacological agents, signaling molecules or dyes to specific organ systems or tissues in live embryos and demonstrates a great potential for gold-silica nanoshells as a contrast agent in embryonic studies.

  20. Lipidome signatures in early bovine embryo development.

    PubMed

    Sudano, Mateus J; Rascado, Tatiana D S; Tata, Alessandra; Belaz, Katia R A; Santos, Vanessa G; Valente, Roniele S; Mesquita, Fernando S; Ferreira, Christina R; Araújo, João P; Eberlin, Marcos N; Landim-Alvarenga, Fernanda D C

    2016-07-15

    Mammalian preimplantation embryonic development is a complex, conserved, and well-orchestrated process involving dynamic molecular and structural changes. Understanding membrane lipid profile fluctuation during this crucial period is fundamental to address mechanisms governing embryogenesis. Therefore, the aim of the present work was to perform a comprehensive assessment of stage-specific lipid profiles during early bovine embryonic development and associate with the mRNA abundance of lipid metabolism-related genes (ACSL3, ELOVL5, and ELOVL6) and with the amount of cytoplasmic lipid droplets. Immature oocytes were recovered from slaughterhouse-derived ovaries, two-cell embryos, and eight- to 16-cell embryos, morula, and blastocysts that were in vitro produced under different environmental conditions. Lipid droplets content and mRNA transcript levels for ACSL3, ELOVL5, and ELOVL6, monitored by lipid staining and quantitative polymerase chain reaction, respectively, increased at morula followed by a decrease at blastocyst stage. Relative mRNA abundance changes of ACSL3 were closely related to cytoplasmic lipid droplet accumulation. Characteristic dynamic changes of phospholipid profiles were observed during early embryo development and related to unsaturation level, acyl chain length, and class composition. ELOVL5 and ELOVL6 mRNA levels were suggestive of overexpression of membrane phospholipids containing elongated fatty acids with 16, 18, and 20 carbons. In addition, putative biomarkers of key events of embryogenesis, embryo lipid accumulation, and elongation were identified. This study provides a comprehensive description of stage-specific lipidome signatures and proposes a mechanism to explain its potential relationship with the fluctuation of both cytoplasmic lipid droplets content and mRNA levels of lipid metabolism-related genes during early bovine embryo development.

  1. Retarded Embryo Development 1 (RED1) regulates embryo development, seed maturation and plant growth in Arabidopsis.

    PubMed

    Du, Qian; Wang, Huanzhong

    2016-07-20

    Plant seeds accumulate large amounts of protein and carbohydrate as storage reserves during maturation. Thus, understanding the genetic control of embryo and seed development may provide bioengineering tools for yield improvement. In this study, we report the identification of Retarded Embryo Development 1 (RED1) gene in Arabidopsis, whose two independent T-DNA insertion mutant lines, SALK_085642 (red1-1) and SALK_022583 (red1-2), show a retarded embryo development phenotype. The embryogenesis process ceases at the late heart stage in red1-1 and at the bent-cotyledon stage in red1-2, respectively, resulting in seed abortion in both lines. The retarded embryo development and seed abortion phenotypes reverted to normal when RED1 complementation constructs were introduced into mutant plants. Small red1-2 homozygous plants can be successfully rescued by culturing immature seeds, indicating that seed abortion likely results from compromised tolerance to the desiccation process associated with seed maturation. Consistent with this observation, red1-2 seeds accumulate less protein, and the expression of two late embryo development reporter transgenes, LEA::GUS and β-conglycinin::GUS, was significantly weak and started relatively late in the red1-2 mutant lines compared to the wild type. The RED1 gene encodes a plant specific novel protein that is localized in the nucleus. These results indicate that RED1 plays important roles in embryo development, seed maturation and plant growth.

  2. Development of interspecies cloned embryos in yak and dog.

    PubMed

    Murakami, Masao; Otoi, Takeshige; Wongsrikeao, Pimprapar; Agung, Budiyanto; Sambuu, Rentsenkhand; Suzuki, Tatsuyuki

    2005-01-01

    Interspecies nuclear transfer (NT) could be an alternative to replicate animals when supply of recipient oocytes is limited or in vitro embryo production systems are incomplete. In the present study, embryonic development was assessed following interspecies NT of donor cumulus cells derived from yak and dog into the recipient ooplasm of domestic cow. The percentages of fusion and subsequent embryo development to the eight-cell stage of interspecies NT embryos were comparable to those of intraspecies NT embryos (cow-cow NT embryos). The percentage of development to blastocysts was significantly lower (p < 0.05) in yak-cow NT embryos than that in cow-cow NT embryos (10.9% vs. 39.8%). In dog-cow NT embryos, only one embryo (0.4%) developed to the blastocyst stage. These results indicate that interspecies NT embryos possess equally developmental competence to the eight-cell stage as intraspecies NT embryos, but the development to blastocysts is very low when dog somatic cells are used as the donor nuclei.

  3. Effect of embryo density on in vitro developmental characteristics of bovine preimplantative embryos with respect to micro and macroenvironments.

    PubMed

    Hoelker, M; Rings, F; Lund, Q; Phatsara, C; Schellander, K; Tesfaye, D

    2010-10-01

    To overcome developmental problems as a consequence of single embryo culture, the Well of the Well (WOW) culture system has been developed. In this study, we aimed to examine the effect of embryo densities with respect to both microenvironment and macroenvironment on developmental rates and embryo quality to get a deeper insight into developmentally important mechanisms. WOW diameter and depth significantly affected developmental rates (p < 0.05). WOWs with diameter of 500 μm reached significantly higher blastocyst rates (32.5 vs 21.1% vs 20.3%) compared to embryos cultured in WOWs of 300 μm diameter or plain cultured controls. Embryos cultured in WOWs with 700 μm depth reached significant higher developmental rates compared with embryos cultured in WOWs of 300 μm depth and control embryos (30.6 vs 22.6% vs 20.3%). Correlation of the embryo per WOW volume with developmental rates was higher (r(2) = 0.92, p = 0.0004) than correlation of WOW diameter or WOW depth with developmental rates. However, the embryo per WOW volume did not affect differential cell counts. An embryo per culture dish volume of 1 : 30 μl was identified to be optimal when the embryo per WOW volume was 1 : 0.27 μl increasing developmental rates up to the level of mass embryo production. Giving the opportunity to track each embryo over the complete culture period while keeping high developmental rates with normal mitotic dynamics, the results of this work will provide benefit for the single culture of embryos in human assisted reproduction, mammalian embryos with high economic interest as well as for scientific purpose.

  4. Tissue densities in developing avian embryos. [under acceleration stresses

    NASA Technical Reports Server (NTRS)

    Smith, A. H.; Abbott, U. K.; Morzenti, A.

    1984-01-01

    The density changes in the components of the incubated egg, the embryo, and the embryo's body parts were measured in the course of 21 days of incubation. In the first two-thirds of the incubation period there is a sequence of increasing density among egg contents: amniotic fluid, embryo, yolk, and albumin. As a result, the embryo is located at the bottom of the amniotic fluid, but at the top of the albumin. This position provides the embryo with mechanical protection and a proximity to the egg's air cell. The observed density changes and the asymmetry of these changes among various body parts of the embryo suggest a functional relationship. The density distributions among the body parts are particularly important in gravitational investigations of embryogenesis since they will produce forces tending to dislocate parts of the embryo.

  5. A fully automated robotic system for microinjection of zebrafish embryos.

    PubMed

    Wang, Wenhui; Liu, Xinyu; Gelinas, Danielle; Ciruna, Brian; Sun, Yu

    2007-09-12

    As an important embodiment of biomanipulation, injection of foreign materials (e.g., DNA, RNAi, sperm, protein, and drug compounds) into individual cells has significant implications in genetics, transgenics, assisted reproduction, and drug discovery. This paper presents a microrobotic system for fully automated zebrafish embryo injection, which overcomes the problems inherent in manual operation, such as human fatigue and large variations in success rates due to poor reproducibility. Based on computer vision and motion control, the microrobotic system performs injection at a speed of 15 zebrafish embryos (chorion unremoved) per minute, with a survival rate of 98% (n = 350 embryos), a success rate of 99% (n = 350 embryos), and a phenotypic rate of 98.5% (n = 210 embryos). The sample immobilization technique and microrobotic control method are applicable to other biological injection applications such as the injection of mouse oocytes/embryos and Drosophila embryos to enable high-throughput biological and pharmaceutical research.

  6. Turtle embryos move to optimal thermal environments within the egg.

    PubMed

    Zhao, Bo; Li, Teng; Shine, Richard; Du, Wei-Guo

    2013-08-23

    A recent study demonstrated that the embryos of soft-shelled turtles can reposition themselves within their eggs to exploit locally warm conditions. In this paper, we ask whether turtle embryos actively seek out optimal thermal environments for their development, as do post-hatching individuals. Specifically, (i) do reptile embryos move away from dangerously high temperatures as well as towards warm temperatures? and (ii) is such embryonic movement due to active thermoregulation, or (more simply) to passive embryonic repositioning caused by local heat-induced changes in viscosity of fluids within the egg? Our experiments with an emydid turtle (Chinemys reevesii) show that embryos avoid dangerously high temperatures by moving to cooler regions of the egg. The repositioning of embryos is an active rather than passive process: live embryos move towards a heat source, whereas dead ones do not. Overall, our results suggest that behavioural thermoregulation by turtle embryos is genuinely analogous to the thermoregulatory behaviour exhibited by post-hatching ectotherms.

  7. Oxamflatin Treatment Enhances Cloned Porcine Embryo Development and Nuclear Reprogramming*

    PubMed Central

    Mao, Jiude; Zhao, Ming-Tao; Whitworth, Kristin M.; Spate, Lee D.; Walters, Eric M.; O'Gorman, Chad; Lee, Kiho; Samuel, Melissa S.; Murphy, Clifton N.; Wells, Kevin; Rivera, Rocio M.

    2015-01-01

    Abstract Faulty epigenetic reprogramming of somatic nuclei is thought to be the main reason for low cloning efficiency by somatic cell nuclear transfer (SCNT). Histone deacetylase inhibitors (HDACi), such as Scriptaid, improve developmental competence of SCNT embryos in several species. Another HDACi, Oxamflatin, is about 100 times more potent than Scriptaid in the ability to inhibit nuclear-specific HDACs. The present study determined the effects of Oxamflatin treatment on embryo development, DNA methylation, and gene expression. Oxamflatin treatment enhanced blastocyst formation of SCNT embryos in vitro. Embryo transfer produced more pigs born and fewer mummies from the Oxamflatin-treated group compared to the Scriptaid-treated positive control. Oxamflatin also decreased DNA methylation of POU5F1 regulatory elements and centromeric repeat elements in day-7 blastocysts. When compared to in vitro–fertilized (IVF) embryos, the methylation status of POU5F1, NANOG, and centromeric repeat was similar in the cloned embryos, indicating these genes were successfully reprogrammed. However, compared to the lack of methylation of XIST in day-7 IVF embryos, a higher methylation level in day-7 cloned embryos was observed, implying that X chromosomes were activated in day-7 IVF blastocysts, but were not fully activated in cloned embryos, i.e., reprogramming of XIST was delayed. A time-course analysis of XIST DNA methylation on day-13, -15, -17, and -19 in vivo embryos revealed that XIST methylation initiated at about day 13 and was not completed by day 19. The methylation of the XIST gene in day-19 control cloned embryos was delayed again when compared to in vivo embryos. However, methylation of XIST in Oxamflatin-treated embryos was comparable with in vivo embryos, which further demonstrated that Oxamflatin could accelerate the delayed reprogramming of XIST gene and thus might improve cloning efficiency. PMID:25548976

  8. Variable imprinting of the MEST gene in human preimplantation embryos

    PubMed Central

    Huntriss, John D; Hemmings, Karen E; Hinkins, Matthew; Rutherford, Anthony J; Sturmey, Roger G; Elder, Kay; Picton, Helen M

    2013-01-01

    There is evidence that expression and methylation of the imprinted paternally expressed gene 1/mesoderm-specific transcript homologue (PEG1/MEST) gene may be affected by assisted reproductive technologies (ARTs) and infertility. In this study, we sought to assess the imprinting status of the MEST gene in a large cohort of in vitro-derived human preimplantation embryos, in order to characterise potentially adverse effects of ART and infertility on this locus in early human development. Embryonic genomic DNA from morula or blastocyst stage embryos was screened for a transcribed AflIII polymorphism in MEST and imprinting analysis was then performed in cDNA libraries derived from these embryos. In 10 heterozygous embryos, MEST expression was monoallelic in seven embryos, predominantly monoallelic in two embryos, and biallelic in one embryo. Screening of cDNA derived from 61 additional human preimplantation embryos, for which DNA for genotyping was unavailable, identified eight embryos with expression originating from both alleles (biallelic or predominantly monoallelic). In some embryos, therefore, the onset of imprinted MEST expression occurs during late preimplantation development. Variability in MEST imprinting was observed in both in vitro fertilization and intracytoplasmic sperm injection-derived embryos. Biallelic or predominantly monoallelic MEST expression was not associated with any one cause of infertility. Characterisation of the main MEST isoforms revealed that isoform 2 was detected in early development and was itself variably imprinted between embryos. To our knowledge, this report constitutes the largest expression study to date of genomic imprinting in human preimplantation embryos and reveals that for some imprinted genes, contrasting imprinting states exist between embryos. PMID:22763377

  9. Gene transfer into older chicken embryos by ex ovo electroporation.

    PubMed

    Luo, Jiankai; Yan, Xin; Lin, Juntang; Rolfs, Arndt

    2012-07-27

    The chicken embryo provides an excellent model system for studying gene function and regulation during embryonic development. In ovo electroporation is a powerful method to over-express exogenous genes or down-regulate endogenous genes in vivo in chicken embryos(1). Different structures such as DNA plasmids encoding genes(2-4), small interfering RNA (siRNA) plasmids(5), small synthetic RNA oligos(6), and morpholino antisense oligonucleotides(7) can be easily transfected into chicken embryos by electroporation. However, the application of in ovo electroporation is limited to embryos at early incubation stages (younger than stage HH20--according to Hamburg and Hamilton)(8) and there are some disadvantages for its application in embryos at later stages (older than stage HH22--approximately 3.5 days of development). For example, the vitelline membrane at later stages is usually stuck to the shall membrane and opening a window in the shell causes rupture of the vessels, resulting in death of the embryos; older embryos are covered by vitelline and allantoic vessels, where it is difficult to access and manipulate the embryos; older embryos move vigorously and is difficult to control the orientation through a relatively small window in the shell. In this protocol we demonstrate an ex ovo electroporation method for gene transfer into chicken embryos at late stages (older than stage HH22). For ex ovo electroporation, embryos are cultured in Petri dishes(9) and the vitelline and allantoic vessels are widely spread. Under these conditions, the older chicken embryos are easily accessed and manipulated. Therefore, this method overcomes the disadvantages of in ovo electroporation applied to the older chicken embryos. Using this method, plasmids can be easily transfected into different parts of the older chicken embryos(10-12).

  10. Pattern of Brachyury gene expression in starfish embryos resembles that of hemichordate embryos but not of sea urchin embryos.

    PubMed

    Shoguchi, E; Satoh, N; Maruyama, Y K

    1999-04-01

    Echinoderms, hemichordates and chordates are deuterostomes and share a number of developmental features. The Brachyury gene is responsible for formation of the notochord, the most defining feature of chordates, and thus may be a key to understanding the origin and evolution of the chordates. Previous studies have shown that the ascidian Brachyury (As-T and Ci-Bra) is expressed in the notochord and that a sea urchin Brachyury (HpTa) is expressed in the secondary mesenchyme founder cells. A recent study by [Tagawa et al. (1998)], however, revealed that a hemichordate Brachyury (PfBra) is expressed in a novel pattern in an archenteron invagination region and a stomodaeum invagination region in the gastrula. The present study demonstrated that the expression pattern of Brachyury (ApBra) of starfish embryos resembles that of PfBra in hemichordate embryos but not of HpTa in sea urchin embryos. Namely, ApBra is expressed in an archenteron invagination region and a stomodaeum invagination region.

  11. Microfluidic EmbryoSort technology: towards in flow analysis, sorting and dispensing of individual vertebrate embryos

    NASA Astrophysics Data System (ADS)

    Fuad, Nurul M.; Wlodkowic, Donald

    2013-12-01

    The demand to reduce the numbers of laboratory animals has facilitated the emergence of surrogate models such as tests performed on zebrafish (Danio rerio) or African clawed frog's (Xenopus levis) eggs, embryos and larvae. Those two model organisms are becoming increasingly popular replacements to current adult animal testing in toxicology, ecotoxicology and also in drug discovery. Zebrafish eggs and embryos are particularly attractive for toxicological analysis due their size (diameter 1.6 mm), optical transparency, large numbers generated per fish and very straightforward husbandry. The current bottleneck in using zebrafish embryos for screening purposes is, however, a tedious manual evaluation to confirm the fertilization status and subsequent dispensing of single developing embryos to multitier plates to perform toxicity analysis. Manual procedures associated with sorting hundreds of embryos are very monotonous and as such prone to significant analytical errors due to operator's fatigue. In this work, we present a proofof- concept design of a continuous flow embryo sorter capable of analyzing, sorting and dispensing objects ranging in size from 1.5 - 2.5 mm. The prototypes were fabricated in polymethyl methacrylate (PMMA) transparent thermoplastic using infrared laser micromachining. The application of additive manufacturing processes to prototype Lab-on-a-Chip sorters using both fused deposition manufacturing (FDM) and stereolithography (SLA) were also explored. The operation of the device was based on a revolving receptacle capable of receiving, holding and positioning single fish embryos for both interrogation and subsequent sorting. The actuation of the revolving receptacle was performed using a DC motor and/or microservo motor. The system was designed to separate between fertilized (LIVE) and non-fertilized (DEAD) eggs, based on optical transparency using infrared (IR) emitters and receivers.

  12. Microspore-derived embryos from Quercus suber anthers mimic zygotic embryos and maintain haploidy in long-term anther culture.

    PubMed

    Bueno, Maria A; Gomez, Arancha; Sepulveda, Federico; Seguí, José M; Testillano, Pilar S; Manzanera, José A; Risueño, Maria-Carmen

    2003-08-01

    Microspore-derived embryos produced from cork oak anther cultures after long-term incubations (up to 10-12 months) were analysed in order to determine the genetic variability and ploidy level stability, as well as morphology, developmental pattern and cellular organisation. Most of the embryos from long-term anther cultures were haploid (90.7%), corresponding to their microspore origin. The presence of a low percentage of diploid embryos (7.4%) was observed. Microsatellite analysis of haploid embryos, indicated different microspores origins of the same anther. In the diploid embryos, homozygosity for different alleles was detected from anther wall tissues, excluding the possibility of clonal origin. The maintenance of a high proportion of haploid embryos, in long-term anther cultures, is similar in percentage to that reported in embryos originating after 20 days of plating (Bueno et al. 1997). This suggests that no significant alterations in the ploidy level occurred during long incubations (up to 12 months). These results suggest that ploidy changes are rare in this in vitro system, and do not significantly increase during long-term cultures. Microscopical studies of the microspore embryos in various stages revealed a healthy and well developed anatomy with no aberrant or chimeric structures. The general morphology of embryos appearing at different times after plating, looked similar to that of earlier embryos, as well as the zygotic embryos, indicating that they represent high quality material for cork oak breeding.

  13. A simple and rapid flow cytometry-based assay to identify a competent embryo prior to embryo transfer

    PubMed Central

    Pallinger, Eva; Bognar, Zoltan; Bodis, Jozsef; Csabai, Timea; Farkas, Nelli; Godony, Krisztina; Varnagy, Akos; Buzas, Edit; Szekeres-Bartho, Julia

    2017-01-01

    Multiple pregnancy is a risk for prematurity and preterm birth. The goal of assisted reproduction is to achieve a single pregnancy, by transferring a single embryo. This requires improved methods to identify the competent embryo. Here, we describe such a test, based on flow cytometric determination of the nucleic acid (PI+) containing extracellular vesicle (EV) count in day 5 embryo culture media. 88 women undergoing IVF were included in the study. More than 1 embryos were transferred to most patients. In 58 women, the transfer resulted in clinical pregnancy, whereas in 30 women in implantation failure. In 112 culture media of embryos from the “clinical pregnancy” group, the number of PI+ EVs was significantly lower than in those of 49 embryos, from the “implantation failure” group. In 14 women, transfer of a single embryo resulted in a singleton pregnancy, or, transfer of two embryos in twin pregnancy. The culture media of 19 out of the 20 “confirmed competent” embryos contained a lower level of PI+ EVs than the cut off level, suggesting that the competent embryo can indeed be identified by low PI+ EV counts. We developed a noninvasive, simple, inexpensive, quick test, which identifies the embryos that are most likely to implant. PMID:28057937

  14. Cells, embryos and development in space

    NASA Technical Reports Server (NTRS)

    Krikorian, A. D.

    1984-01-01

    Work continues to focus on the demonstrable totipotency of cultured somatic cells of various higher plants and has examined the conditions which regulate this propensity to be controllably released. This was done with special reference to cells obtained from cultured explants of daylily and carrot. For purposes of identifying the variables in question, work was carried out almost exclusively in liquid media. The events that intervene between the aseptic isolation of tissue explants, the culture of small derived units and free cells and the propagation in large numbers of adventive or somatic embryos to plantlets were traced and certain definitive stages at which control is exercised were identified. In daylily, morphologically competent units are now propagated with a high degree of precision in rotated liquid cultures in bulk, and under the conditions of continuous neutralized gravity, the development progresses so that embryo-plantlets are obtained.

  15. Control of segment number in vertebrate embryos.

    PubMed

    Gomez, Céline; Ozbudak, Ertuğrul M; Wunderlich, Joshua; Baumann, Diana; Lewis, Julian; Pourquié, Olivier

    2008-07-17

    The vertebrate body axis is subdivided into repeated segments, best exemplified by the vertebrae that derive from embryonic somites. The number of somites is precisely defined for any given species but varies widely from one species to another. To determine the mechanism controlling somite number, we have compared somitogenesis in zebrafish, chicken, mouse and corn snake embryos. Here we present evidence that in all of these species a similar 'clock-and-wavefront' mechanism operates to control somitogenesis; in all of them, somitogenesis is brought to an end through a process in which the presomitic mesoderm, having first increased in size, gradually shrinks until it is exhausted, terminating somite formation. In snake embryos, however, the segmentation clock rate is much faster relative to developmental rate than in other amniotes, leading to a greatly increased number of smaller-sized somites.

  16. Photobiomodulation of early mouse embryo development

    NASA Astrophysics Data System (ADS)

    Sviridova-Chailakhyan, T. A.; Fakhranurova, L. I.; Simonova, N. B.; Khramov, R. N.; Manokhin, A. A.; Paskevich, S. I.; Chailakhyan, L. M.

    2008-04-01

    The effect of artificial sunlight (AS) from a xenon source and of converted AS with an additional orange-red luminescent (λ MAX=626 nm) component (AS+L) on the development of mouse zygotes was investigated. A plastic screen with a photoluminophore layer was used for production of this orange-red luminescent (L) component. A single short-term (15 min) exposure produced a long-term stable positive effect on early embryo development of mice, which persisted during several days. After exposure to AS+L, a stimulating influence on preimplantation development was observed, in comparison with the control group without AS exposure. The positive effects were as follows: increase in percent of embryos (P <= 0.05) developed to the blastocyst stage (96.2 %) with hatching from the zona pellucida (80.8 %) within 82-96 hours in vitro compared to the control (67.1 % and 28.8 %, respectively).

  17. Echinoderm eggs and embryos: procurement and culture.

    PubMed

    Foltz, Kathy R; Adams, Nikki L; Runft, Linda L

    2004-01-01

    The protocols outlined here hopefully will provide researchers with healthy, beautiful echinoderm oocytes, eggs, and embryos for experimental use. The large size of echinoderm oocytes and eggs, the ease with which they can be manipulated, and (in many species) their optical clarity, make them an ideal model system for studying not only the events specific to oocyte maturation and fertilization, but also for investigating more general questions regarding cell cycle regulation in an in vivo system. The quick rate at which development proceeds after fertilization to produce transparent embryos and larva makes the echinoderm an advantageous organism for studying deuterostome embryogenesis. Continued use of the echinoderms as model systems will undoubtedly uncover exciting answers to questions regarding fertilization, cell cycle regulation, morphogenesis, and how developmental events are controlled.

  18. Estimating limits for natural human embryo mortality.

    PubMed

    Jarvis, Gavin E

    2016-01-01

    Natural human embryonic mortality is generally considered to be high. Values of 70% and higher are widely cited. However, it is difficult to determine accurately owing to an absence of direct data quantifying embryo loss between fertilisation and implantation. The best available data for quantifying pregnancy loss come from three published prospective studies (Wilcox, Zinaman and Wang) with daily cycle by cycle monitoring of human chorionic gonadotrophin (hCG) in women attempting to conceive. Declining conception rates cycle by cycle in these studies indicate that a proportion of the study participants were sub-fertile. Hence, estimates of fecundability and pre-implantation embryo mortality obtained from the whole study cohort will inevitably be biased. This new re-analysis of aggregate data from these studies confirms the impression that discrete fertile and sub-fertile sub-cohorts were present. The proportion of sub-fertile women in the three studies was estimated as 28.1% (Wilcox), 22.8% (Zinaman) and 6.0% (Wang). The probability of conceiving an hCG pregnancy (indicating embryo implantation) was, respectively, 43.2%, 38.1% and 46.2% among normally fertile women, and 7.6%, 2.5% and 4.7% among sub-fertile women. Pre-implantation loss is impossible to calculate directly from available data although plausible limits can be estimated. Based on this new analysis and a model for evaluating reproductive success and failure it is proposed that a plausible range for normal human embryo and fetal mortality from fertilisation to birth is 40-60%.

  19. Developmental toxicity of cartap on zebrafish embryos.

    PubMed

    Zhou, Shengli; Dong, Qiaoxiang; Li, Shaonan; Guo, Jiangfeng; Wang, Xingxing; Zhu, Guonian

    2009-12-13

    Cartap is a widely used insecticide which belongs to a member of nereistoxin derivatives and acts on nicotinic acetylcholine receptor site. Its effects on aquatic species are of grave concern. To explore the potential developmental toxicity of cartap, zebrafish embryos were continually exposed, from 0.5 to 144h post-fertilization, to a range of concentrations of 25-1000microg/l. Results of the experiment indicated that cartap concentrations of 100microg/l and above negatively affected embryo survival and hatching success. Morphological analysis uncovered a large suite of abnormalities such as less melanin pigmentation, wavy notochord, crooked trunk, fuzzy somites, neurogenesis defects and vasculature defects. The most sensitive organ was proved to be the notochord which displayed defects at concentrations as low as 25microg/l. Both sensitivity towards exposure and localization of the defect were stage specific. To elucidate mechanisms concerning notochord, pigmentation, and hatching defects, enzyme assay, RT Q-PCR, and different exposure strategies were performed. For embryos with hatching failure, chorion was verified not to be digested, while removing cartap from exposure at early pre-hatching stage could significantly increase the hatching success. However, cartap was proved, via vitro assay, to have no effect on proteolytic activity of hatching enzyme. These findings implied that the secretion of hatching enzyme might be blocked. We also revealed that cartap inhibited the activity of melanogenic enzyme tyrosinase and matrix enzyme lysyl oxidase and induced expression of their genes. These suggested that cartap could impaired melanin pigmentation of zebrafish embryos through inhibiting tyrosinase activity, while inhibition of lysyl oxidase activity was responsible for notochord undulation, which subsequently caused somite defect, and at least partially responsible for defects in vasculature and neurogenesis.

  20. Blastocoel expansion in the preimplantation mouse embryo

    SciTech Connect

    Manejwala, F.M.

    1989-01-01

    Since cAMP can regulate fluid transport in many types of epithelia, the mechanism of fluid transport and the role of cAMP in the mouse blastocyst were examined. Results described here indicate an increase in the ability of mouse embryos to elevate cAMP levels in response to activators of adenylate cyclase, which is the enzyme that synthesizes cAMP, during the transition from morula to blastocyst. In addition, a positive correlation is observed between the increase in activatable adenylate cyclase and the presence of a blastocoel. Moreover, elevating intracellular cAMP in nascent cavitating embryos stimulates the rate of fluid transport in the blastocoel. Accumulation of fluid in the blastocoel is a function of the tight permeability seal around the embryo and the vectorial flow of ions into the blastocoel. Results reported here indicate that extracellular Na{sup +} and Cl{sup {minus}} are required for expansion of the blastocoel in nascent cavitating blastocysts. Sodium uptake into the embryo is carrier-mediated and probably occurs through multiple routes which include the Na{sup +}-channel and the Na{sup +}/H{sup +} exchanger. Chloride uptake is non-carrier mediated and may occur by a paracellular route. In addition, cAMP, which stimulates blastocoel expansion, also stimulates uptake of {sup 22}Na{sup +}. This effect may be mediated by a cAMP-dependent protein kinase, since inhibition of this enzyme inhibits both the cAMP-stimulated rate of blastocoel expansion and {sup 22}Na{sup +} uptake.

  1. Cryopreservation of ilex immature zygotic embryos.

    PubMed

    Mroginski, Luis; Dolce, Natalia; Sansberro, Pedro; Luna, Claudia; Gonzalez, Ana; Rey, Hebe

    2011-01-01

    Tropical Ilex species have recalcitrant seeds. This chapter describes protocols for long-term conservation of Ilex brasiliensis, I. brevicuspis, I. dumosa, I. microdonta, I. integerrima, I. paraguariensis, I. pseudoboxus, I. taubertiana, and I. theezans through cryopreservation of zygotic rudimentary embryos at the heart developmental stage. The embryos are aseptically removed from the seeds and precultured (7 days) in the dark at 27±2°C on solidified quarter-strength Murashige and Skoog medium with 3% sucrose and 0.1 mg/L zeatin. The embryos are then encapsulated in 3% calcium alginate beads and pretreated at 24-h intervals in liquid medium supplemented with progressively increasing sucrose concentrations (0.5, 0.75, and 1 M). The beads are dehydrated for 5 h with silica gel to 25% water content (fresh weight basis) and then placed in sterile 5-mL cryovials. Then the beads are either plunged rapidly in liquid nitrogen where they are kept for 1 h (rapid cooling), or cooled at 1°C/min to -30°C and then immersed in liquid nitrogen for 1 h (slow cooling). After cryopreservation, the beads are rewarmed by immersion of the cryovials for 1 min in a water bath at 30°C. Finally, the beads are transferred onto culture medium (1/4MS, 3% sucrose, and 0.1 mg/L zeatin, solidified with 0.8% agar) and incubated in a growth room at 27±2°C under a 14-h light (116 μmol/m2/s) and 10-h dark photoperiod. Maximum recovery percentages between 15 and 83% (depending on the species and the treatment) were obtained with the cryopreserved embryos.

  2. Galileo and the embryos: religion and science in parliamentary debate over research on human embryos.

    PubMed

    Mulkay, Michael

    1995-08-01

    Confrontation between science and religion was a significant feature of the lengthy public appraisal of research on human embryos in Britain during the 1880s. The series of formal debates over embryo research in the House of Lords is chosen as a particularly appropriate setting to study this confrontation. It is shown that religious opposition to embryo research was repeatedly attacked in these debates by means of a stereotyped contrast between religious and scientific styles of thought. Leading figures in the movement for embryo research attempted to discredit their opponents by claiming that, whereas their own case was built upon reasoned assessment of the facts, the other side relied on religious dogma, clerical authority and faith. It is shown that, although there were genuine differences between those critical of embryo research on religious grounds and those supporting such research on grounds furnished by scientists, this account of the differences is inaccurate: dogma, reliance on authority and faith were as characteristic of the discourse associated with science as they were of that associated with religion. It is argued that these features were not generated by the presence of religious or scientific beliefs as such, but by the struggle between advocates of science and religion for intellectual and moral dominance.

  3. Human-animal interaction, stress, and embryo production in Bos indicus embryo donors under tropical conditions.

    PubMed

    Macedo, Gustavo Guerino; Zúccari, Carmem Estefânia Serra Neto; de Abreu, Urbano Gomes Pinto; Negrão, João Alberto; da Costa e Silva, Eliane Vianna

    2011-08-01

    This study investigated the effect of human-animal interaction (HAI) and the stress response on the quality of embryo production in superovulated Nelore (Bos indicus) cattle, under tropical conditions. Thirty-two females underwent a superovulation protocol for 5 days. Cortisol concentrations were determined in blood plasma collected on days 0, 4, and 5. Artificial insemination was performed on days 4 and 5, and nonsurgical embryo flushing on day 11. Embryo production and viability were determined. Human stimulation, animal behaviors, accidents, and handling time were recorded to assess HAI. Cattle age was negatively correlated with accidents, frequency of aversive behaviors, and negative stimuli by stockperson during transit through corral compartments to receive superovulation treatments. The factor analysis revealed two distinct groups. The first group was called stressed and had higher cortisol concentration than the nonstressed group, 16.0 ± 2.1 and 12.5 ± 1.0 ng/mL, respectively. Comparisons between these groups showed that the frequency of voice emissions by the stockperson and the number of accidents were higher in the stressed group, and also, the mean handling time was longer in the stressed group than for the nonstressed. As a result, viability rate of the embryos was 19% lower in the stressed group (P < 0.05). This indicates that intensive negative HAI is likely related to stress, which affects embryo production in a superovulation program.

  4. Characterization of embryo-specific genes

    SciTech Connect

    Sung, R.

    1992-06-12

    The objective of the proposed research is to characterize the function and regulation of a set of embryonic genes which are expressed in the embryos, not in the plants. 22 cDNA clones were isolated from a cDNA library we constructed using mRNAS of -carrot somatic embryos. These cDNA clones identified mRNA species that are present in the somatic and zygotic embryos, but not in adult plants. The sequence of all 22cDNA clones were determined; genomic clones for three cDNA clones, DC8, DC59, and DC49 were isolated and gene sequences determined. DC8, DC49, and several other genes identified by the cDNA sequences belong to the category of late embryogenesis abundant protein genes, Lea. The function of these gens have not yet been determined, but they share common structural features, are regulated by ABA and are speculated to play a role in seed desiccation.

  5. Human embryo cloning prohibited in Hong Kong.

    PubMed

    Liu, Athena

    2005-12-01

    Since the birth of Dolly (the cloned sheep) in 1997, debates have arisen on the ethical and legal questions of cloning-for-biomedical-research (more commonly termed "therapeutic cloning") and of reproductive cloning using human gametes. Hong Kong enacted the Human Reproductive Technology Ordinance (Cap 561) in 2000. Section 15(1)(e) of this Ordinance prohibits the "replacing of the nucleus of a cell of an embryo with a nucleus taken from any other cell," i.e., nucleus substitution. Section 15(1)(f) prohibits the cloning of any embryo. The scope of the latter, therefore, is arguably the widest, prohibiting all cloning techniques such as cell nucleus replacement, embryo splitting, parthenogenesis, and cloning using stem cell lines. Although the Human Reproductive Technology Ordinance is not yet fully operative, this article examines how these prohibitions may adversely impact on basic research and the vision of the Hong Kong scientific community. It concludes that in light of recent scientific developments, it is time to review if the law offers a coherent set of policies in this area.

  6. Role of melatonin in embryo fetal development.

    PubMed

    Voiculescu, S E; Zygouropoulos, N; Zahiu, C D; Zagrean, A M

    2014-01-01

    Melatonin is an indoleamine produced by the pineal gland and secreted in a circadian manner. In the past few decades, research over this topic has been enhanced. Melatonin has many important roles in the human physiology: regulator of the circadian rhythms, sleep inducer, antioxidant, anticarcinogenic. This paper reviews the involvement of melatonin in embryo fetal development. The pineal gland develops completely postpartum, so both the embryo and the fetus are dependent on the maternal melatonin provided transplacentally. Melatonin appears to be involved in the normal outcome of pregnancy beginning with the oocyte quality and finishing with the parturition. Its pregnancy night-time concentrations increase after 24 weeks of gestation, with significantly high levels after 32 weeks. Melatonin receptors are widespread in the embryo and fetus since early stages. There is solid evidence that melatonin is neuroprotective and has a positive effect on the outcome of the compromised pregnancies. In addition, chronodisruption leads to a reproductive dysfunction. Thus, the influence of melatonin on the developing human fetus may not be limited to the entertaining of circadian rhythmicity, but further studies are needed.

  7. In vivo photoacoustic imaging of mouse embryos

    NASA Astrophysics Data System (ADS)

    Laufer, Jan; Norris, Francesca; Cleary, Jon; Zhang, Edward; Treeby, Bradley; Cox, Ben; Johnson, Peter; Scambler, Pete; Lythgoe, Mark; Beard, Paul

    2012-06-01

    The ability to noninvasively image embryonic vascular anatomy in mouse models is an important requirement for characterizing the development of the normal cardiovascular system and malformations in the heart and vascular supply. Photoacoustic imaging, which can provide high resolution non invasive images of the vasculature based upon optical absorption by endogenous hemoglobin, is well suited to this application. In this study, photoacoustic images of mouse embryos were obtained ex vivo and in vivo. The images show intricate details of the embryonic vascular system to depths of up to 10 mm, which allowed whole embryos to be imaged in situ. To achieve this, an all-optical photoacoustic scanner and a novel time reversal image reconstruction algorithm, which provide deep tissue imaging capability while maintaining high spatial resolution and contrast were employed. This technology may find application as an imaging tool for preclinical embryo studies in developmental biology as well as more generally in preclinical and clinical medicine for studying pathologies characterized by changes in the vasculature.

  8. Genetic analysis of embryo dormancy. Final report

    SciTech Connect

    Galau, G.

    1998-09-01

    Primary dormancy is the inability of mature seed to immediately germinate until specific environmental stimuli are perceived that predict that future conditions will support plant growth and seed set. The analysis of abscisic acid deficient and insensitive mutants, in particular in Arabidopsis, suggests that embryo abscisic acid may be directly involved in the development of primary dormancy. Other studies implicate the continued accumulation of LEA proteins as inhibiting germination in dormant embryos. The results of these physiological, molecular and genetic approaches are complex and equivocal. There is a real need for approaches that test the separate nature of vivipary inhibition and primary dormancy and deliberately seed to decouple and dissect them. These approaches should be of help in understanding both late embryo development and primary dormancy. The approach taken here is to directly isolate mutants of Arabidopsis that appear to be deficient only in primary dormancy, that is fresh seed that germinate rapidly without the normally-required cold-stratification. The authors have isolated at least 8 independent, rapidly germinating RGM mutants of Arabidopsis. All others aspects of plant growth and development appear normal in these lines, suggesting that the rgm mutants are defective only in the establishment or maintenance of primary dormancy. At least one of these may be tagged with T-DNA. In addition, about 50 RGM isolates have been recovered from EMS-treated seed.

  9. Stimulus-triggered enhancement of chilling tolerance in zebrafish embryos

    PubMed Central

    Szabó, Katalin; Budai, Csilla; Losonczi, Eszter; Bernáth, Gergely; Csenki-Bakos, Zsolt; Urbányi, Béla; Pribenszky, Csaba; Horváth, Ákos; Cserepes, Judit

    2017-01-01

    Background Cryopreservation of zebrafish embryos is still an unsolved problem despite market demand and massive efforts to preserve genetic variation among numerous existing lines. Chilled storage of embryos might be a step towards developing successful cryopreservation, but no methods to date have worked. Methods In the present study, we applied a novel strategy to improve the chilling tolerance of zebrafish embryos by introducing a preconditioning hydrostatic pressure treatment to the embryos. In our experiments, 26-somites and Prim-5 stage zebrafish embryos were chilled at 0°C for 24 hours after preconditioning. Embryo survival rate, ability to reach maturation and fertilizing capacity were tested. Results Our results indicate that applied preconditioning technology made it possible for the chilled embryos to develop normally until maturity, and to produce healthy offspring as normal, thus passing on their genetic material successfully. Treated embryos had a significantly higher survival and better developmental rate, moreover the treated group had a higher ratio of normal morphology during continued development. While all controls from chilled embryos died by 30 day-post-fertilization, the treated group reached maturity (~90–120 days) and were able to reproduce, resulting in offspring in expected quantity and quality. Conclusions Based on our results, we conclude that the preconditioning technology represents a significant improvement in zebrafish embryo chilling tolerance, thus enabling a long-time survival. Furthermore, as embryonic development is arrested during chilled storage this technology also provides a solution to synchronize or delay the development. PMID:28166301

  10. Optimizing the culture environment and embryo manipulation to help maintain embryo developmental potential.

    PubMed

    Swain, Jason E; Carrell, Doug; Cobo, Ana; Meseguer, Marcos; Rubio, Carmen; Smith, Gary D

    2016-03-01

    With increased use of comprehensive chromosome screening (CCS), the question remains as to why some practices do not experience the same high levels of clinical success after implementation of the approach. Indeed, the debate surrounding the efficacy and usefulness of blastocyst biopsy and CCS continues. Importantly, several variables impact the success of an assisted reproductive technology cycle. Transfer of a euploid embryo is but one factor in an intricate system that requires numerous steps to occur successfully. Certainly, the culture environment and the manipulations of the embryo during its time in the laboratory can impact its reproductive potential. Environmental stressors ranging from culture media to culture conditions and even culture platform can impact biochemical, metabolic, and epigenetic patterns that can affect the developing cell independent of chromosome number. Furthermore, accompanying procedures, such as biopsy and vitrification, are complex and, when performed improperly, can negatively impact embryo quality. These are areas that likely still carry room for improvement within the IVF laboratory.

  11. Effects of embryo-derived exosomes on the development of bovine cloned embryos

    PubMed Central

    Qu, Pengxiang; Qing, Suzhu; Liu, Ruiqi; Qin, Hongyu; Wang, Weiwei; Qiao, Fang; Ge, Hui; Liu, Jun; Zhang, Yong; Wang, Yongsheng

    2017-01-01

    The developmental competence of in vitro cultured (IVC) embryos is markedly lower than that of their in vivo counterparts, suggesting the need for optimization of IVC protocols. Embryo culture medium is routinely replaced three days after initial culture in bovine, however, whether this protocol is superior to continuous nonrenewal culture method under current conditions remains unclear. Using bovine somatic cell nuclear transfer (SCNT) embryos as the model, our results showed that compared with routine renewal treatment, nonrenewal culture system significantly improved blastocyst formation, blastocyst quality (increased total cell number, decreased stress and apoptosis, enhanced Oct-4 expression and ratio of ICM/TE), as well as following development to term. Existence and function of SCNT embryo-derived exosomes were then investigated to reveal the cause of impaired development induced by culture medium replacement. Exosomes were successfully isolated through differential centrifugation and identified by both electron microscopy and immunostaining against exosomal membrane marker CD9. Supplementation of extracted exosomes into freshly renewed medium significantly rescued not only blastocyst formation and quality (in vitro development), but also following growth to term (in vivo development). Notably, ratio of ICM/TE and calving rate were enhanced to a similar level as that in nonrenewal group. In conclusion, our results for the first time indicate that 1: bovine SCNT embryos can secrete exosomes into chemically defined culture medium during IVC; 2: secreted exosomes are essential for SCNT blastocyst formation, blastocyst quality, and following development to term; 3: removal of exosomes induced by culture medium replacement impairs SCNT embryo development, which can be avoided by nonrenewal culture procedure or markedly recovered by exosome supplementation. PMID:28350875

  12. Can Coxiella burnetii be transmitted by embryo transfer in goats?

    PubMed

    Alsaleh, A; Fieni, F; Rodolakis, A; Bruyas, J F; Roux, C; Larrat, M; Chatagnon, G; Pellerin, J L

    2013-10-01

    The detection of significant bacterial loads of Coxiella burnetii in flushing media and tissue samples from the genital tracts of nonpregnant goats represents a risk factor for in utero infection and transmission during embryo transfer. The aim of this study was to investigate (1) whether cells of early goat embryos isolated from in vivo-fertilized goats interact with C. burnetii in vitro, (2) whether the embryonic zona pellucida (ZP) protects early embryo cells from infection, and (3) the efficacy of the International Embryo Transfer Society (IETS) washing protocol for bovine embryos. The study was performed in triple replicate: 12 donor goats, certified negative by ELISA and polymerase chain reaction, were synchronized, superovulated, and subsequently inseminated by Q fever-negative males. Sixty-eight embryos were collected 4 days later by laparotomy. Two-thirds of the resulting ZP-intact and ZP-free 8- to 16-cell embryos (9-9, 11-11, and 4-4 in replicates 1, 2, and 3, respectively) were placed in 1 mL minimum essential medium containing 10(9)C. burnetii CBC1 (IASP, INRA Tours). After overnight incubation at 37 °C and 5% CO2, the embryos were washed according to the IETS procedure. In parallel, the remaining third ZP-intact and ZP-free uninfected embryos (3-3, 5-5, and 2-2 in replicates 1, 2, and 3, respectively) were subjected to the same procedures, but without C. burnetii, thus serving as controls. The 10 washing fluids for all batches of each replicate were collected and centrifuged for 1 hour at 13,000 × g. The washed embryos and pellets were tested by polymerase chain reaction. Coxiella burnetii DNA was found in all batches of ZP-intact and ZP-free infected embryos after 10 successive washes. It was also detected in the first five washing fluids for ZP-intact embryos and in the first eight washing fluids for ZP-free embryos. None of the control batches (embryos and washing fluids) were found to contain bacterial DNA. These results clearly indicate that

  13. Fiber optic light-scattering measurement system for evaluation of embryo viability: light-scattering characteristics from live mouse embryo

    NASA Astrophysics Data System (ADS)

    Itoh, Harumi; Arai, Tsunenori; Kikuchi, Makoto

    1997-06-01

    We measured angular distribution of the light scattering from live mouse embryo with 632.8nm in wavelength to evaluate the embryo viability. We aim to measure the mitochondrial density in human embryo which have relation to the embryo viability. We have constructed the light scattering measurement system to detect the mitochondrial density non-invasively. We have employed two optical fibers for the illumination and sensing to change the angle between these fibers. There were two dips on the scattering angular distribution from the embryo. These dips existed on 30 and 85 deg. We calculated the scattering angular pattern by Mie theory to fit the measured scattering estimated scattering size and density. The best fitting was obtained when the particle size and density were 0.9 micrometers and 1010 particles per ml, respectively. These values coincided with the approximated values of mitochondrial in the embryo. The measured light scattering may mainly originated from mitochondria in spite of the existence of the various scattering particles in the embryo. Since our simple scattering measurement may offer the mitochondrial density in the embryo, it might become the practical method of human embryo on in vitro fertilization-embryo transfer.

  14. In vitro culture of embryos of the guppy, Poecilia reticulata.

    PubMed

    Martyn, Ulrike; Weigel, Detlef; Dreyer, Christine

    2006-03-01

    The rich variation in adult color patterns of male guppies (Poecilia reticulata) has attracted the attention of geneticists and ecologists for almost a century. Studies on their embryogenesis, however, have been limited by the fact that guppies are live bearers. We have observed normal development after explantation of guppy embryos from the ovary of pregnant females at various times after last parturition, and found that development of each batch of eggs is slightly asynchronous, most likely due to asynchronous fertilization. We have cultured explanted embryos in vitro and continuously observed their development. Although embryos explanted a few days after fertilization survived up to 4 weeks in culture, they did not complete their development. In contrast, embryos explanted at late stages of gestation could hatch and develop to fertile adults. Our embryo culture techniques overcome some of the limitations of using livebearers as study objects, and they allow continuous observation of and accessibility to live embryos at all stages.

  15. Developmental arrest in grass shrimp embryos exposed to selected toxicants

    SciTech Connect

    Wilson, J.E.H.

    1998-12-31

    Excised embryos of the grass shrimp (Palaemonetes pugio) were exposed to single pulse concentrations of selected pollutants for 4 days. The following toxicity endpoints were monitored: rate of embryonic development, embryo mortality, and types of embryo malformation. Each endpoint exhibited concentration--response relationships which were modified by the embryonic age at which exposure commenced. Developmental retardation of up to 3 days was effected by phenol at 0.01% (V/V) and complete developmental arrest occurred at 0.05% and 0.1% (V/V). Similarly for methylene chloride, developmental retardation of 1003 days were observed at 0.1% (V/V) depending on the age of the embryos at the start of the tests. The morphological abnormalities of the embryos are described. The ecological significance of these findings and implications for the development of short-term toxicity tests using grass shrimp embryos are discussed.

  16. Interference Function of Crystalline Embryo Model of Amorphous Metals. I

    NASA Astrophysics Data System (ADS)

    Hamada, Tadashi; Fujita, Francisco Eiichi

    1982-07-01

    A simple and possible structural model of amorphous metals based on the concept of crystalline embryos is proposed. The quasi-crystalline clusters are supposed to exist in the liquid state, be enhanced during supercooling, and be frozen as the crystalline embryos in the amorphous state by rapid quenching. A model assembly of atoms containing the crystalline embryos and the boundary regions is constructed, and the pair correlation function and the interference function are calculated. The interference function of the b.c.c. embryo model is in good agreement with experimental ones. It is concluded that the structure of the boundary connecting the embryos plays an essential role as well as the ordered part in the embryos in the diffraction phenomena of the amorphous structures. The importance of chemical clusters and metalloid atoms is also suggested and discussed.

  17. Bovine embryo recovery by filtration of non-surgical flushings.

    PubMed

    Pugh, A; Trounson, A O; Aarts, M H; McPhee, S

    1980-04-01

    A simple filtration system has been developed for the rapid collection of bovine embryos from large fluid volumes such as non-surgical uterine flushings. The technique utilizes a nylon plankton net sieve of 56 mum pore size and was evaluated on the non-surgical flushings of 18 superovulated cows. Approximately 500 ml of flushings from each uterine horn was collected in sedimentation flasks and two aliquots of 20 ml removed from the bottom of the flask after standing for 20 min, and searched for embryos. The remainder of the flushings was passed through the sieve and the sieve examined for embryos. Seven days (Day 7) after insemination, 53.3% (40 75 ) of embryos were found on the sieve or 47.2% of all normal embryos. On Day 12,28% (7 25 ) of eggs were found on the sieve, all of which were unfertilized or degenerate. All embryos were located within 10 min of starting filtration.

  18. Fatty acid breakdown in developing embryos of Brassica napus L.

    PubMed

    Chia, T; Rawsthorne, S

    2000-12-01

    Developing Brassica napus embryos are primarily concerned with the accumulation of storage products, namely oil, starch and protein. The presence of fatty acid catabolic pathways in the background of this biosynthetic activity was investigated. Enzymes involved in the process of lipid mobilization, such as malate synthase and isocitrate lyase, are detectable towards the late stages of embryo development. [(14)C]Acetate feeding experiments also reveal that fatty acid catabolism becomes increasingly functional as the embryo matures.

  19. Efficient Production and Cellular Characterization of Sheep Androgenetic Embryos

    PubMed Central

    Zacchini, Federica; Czernik, Marta; Iuso, Domenico; Toschi, Paola; di Egidio, Fiorella; Scapolo, Pier Augusto; Ptak, Grazyna

    2011-01-01

    Abstract The production of androgenetic embryos in large animals is a complex procedure. Androgenetic embryos have been produced so far only in cattle and sheep using pronuclear transfer (PT) between zygotes derived from in vitro fertilization (IVF) of previously enucleated oocytes. PT is required due to the poor developmental potential of androgenotes derived from IVF of enucleated oocytes. Here we compare the developemt to blastocyst of androgenetic embryos produced by the standard pronuclear transfer and by fertilization of oocytes enucleated in Ca2+/Mg2+-free medium, without pronuclear transfer. The enucleation in Ca2+/Mg2+-free medium abolished almost completely the manipulation-induced activation, significantly improving the development to blastocyst of the androgenetic embryos (IVF followed by PT; 18.6%: IVF only; 17.7%, respectively). Karyotype analysis of IVF revealed a similar proportion of diploid embryos in androgenetic and control blastocysts (35% and 36%, respectively), although mixoploid blastocysts were frequently observed in both groups (64%). Androgenotes had lower total cell numbers than control and parthenogenetic embryos, but more cells in ICM cells comparing to parthenogenotes (30.42 vs. 17.15%). Higher expression of the pluripotency-associated gene NANOG, and trophoblastic-specific gene CDX2, were also observed in androgenotes compared to parthenogenotes and controls. The global methytion profile of androgenetic embryos was comparable to controls, but was lower than parthenogenetic embryos. The cell composition and methylation pattern we have detected in monoparental sheep monoparental embryos are unprecedented, and differ considerably from the standard reference mouse embryos. Altogether, these finding indicate significant differences across species in the molecular mechanisms regulating early development of monoparental embryos, and highlights the need to study postimplantation development of androgenetic embryos in sheep. PMID:22043807

  20. Injuries in pacu embryos (Piaractus mesopotamicus) after freezing and thawing.

    PubMed

    Neves, Patrícia Ribeiro; Ribeiro, Ricardo Pereira; Streit, Danilo Pedro; Natali, Maria Raquel M; Fornari, Darci Carlos; Santos, Alexandra Inês; Godoy, Leandro C

    2014-02-01

    Although the sperm cryopreservation of freshwater and marine teleosts has been feasible for years, the cryopreservation of some fish embryos still remains elusive. Thus, the objective of this experiment was to analyze the embryo morphology after freezing and thawing 40 embryos of Piaractus mesopotamicus immersed into methanol and ethylene glycol, both at 7, 10 and 13% plus 0.1 M sucrose for 10 min. Soon after thawing, three embryos were treated with historesin, stained with hematoxylin-eosin and analyzed under an optical microscope. From every treatment, one palette containing embryos was thawed and incubated, but none of the eggs hatched. Samples containing two embryos were immersed into 10% methanol or 10% ethylene glycol both in association with sucrose, and embryos immersed into only water or sucrose solution were frozen, processed and analyzed using scanning electron microscopy (SEM). In both cases, the control group was immersed into only water. Although the embryos had the chorion, vitello, yolk syncytial layer and blastoderm, all of them were found altered under the optical microscope and by SEM. The chorion was irregular and injured; there was no individuality in the yolk granules; the yolk syncytial layer had an irregular shape, thickness and size; the blastoderm showed injuries in the nucleus shape and sometimes was absent; the blastoderm was located in atypical areas and absent in some embryos. In conclusion, no treatment was effective in preserving the embryos, and none of the embryos avoided injury from intracellular ice formation. These morphological injuries during the freezing process made the P. mesopotamicus embryos unfeasible for hatching.

  1. Digital microfluidic processing of mammalian embryos for vitrification.

    PubMed

    Pyne, Derek G; Liu, Jun; Abdelgawad, Mohamed; Sun, Yu

    2014-01-01

    Cryopreservation is a key technology in biology and clinical practice. This paper presents a digital microfluidic device that automates sample preparation for mammalian embryo vitrification. Individual micro droplets manipulated on the microfluidic device were used as micro-vessels to transport a single mouse embryo through a complete vitrification procedure. Advantages of this approach, compared to manual operation and channel-based microfluidic vitrification, include automated operation, cryoprotectant concentration gradient generation, and feasibility of loading and retrieval of embryos.

  2. Twin delivery following 12 years of human embryo cryopreservation: case report.

    PubMed

    Revel, A; Safran, A; Laufer, N; Lewin, A; Reubinov, B E; Simon, A

    2004-02-01

    It is uncertain how long IVF units can keep frozen embryos. Few data exist on success of embryo transfer for embryos that have been cryopreserved for many years. We report the delivery of healthy twins following the transfer of embryos cryopreserved for 12 years. To the best of our knowledge, this is the longest reported successful human embryo freezing.

  3. Mouse embryos' fusion for the tetraploid complementation assay.

    PubMed

    Gertsenstein, Marina

    2015-01-01

    Production of the germline-competent chimeras using genetically modified ES cell lines is an essential step in the establishment of novel mouse models. In addition chimeras provide a powerful tool to study the cell lineage and to analyze complex phenotypes of mutant mice. Mouse chimeras with tetraploid embryos are used to rescue extraembryonic defects, to analyze an impact of gene function on specific lineage, to study the interaction between embryonic and extraembryonic tissues, and to produce mutant embryos and mice for the phenotype analysis. Tetraploid embryos are generated by the fusion of two blastomeres of the mouse embryo. The applications of tetraploid complementation assay and the protocol are described below.

  4. In vitro fertilization and embryo development in pigs.

    PubMed

    Abeydeera, L R

    2001-01-01

    Considerable progress has been made in the in vitro production of pig embryos using improved methods for in vitro maturation (IVM) and fertilization (IVF). Despite the progress, polyspermic penetration remains a problem for in vitro-matured oocytes. Variation among boars, ejaculates and IVF protocols used in different laboratories appears to influence the incidence of polyspermy. Recent studies indicate that oviduct cells and their secretions play a role in reducing polyspermy. Very early attempts to culture in vivo-derived pig embryos met with little success and most were arrested at the four-cell stage. At present, many culture media are available that can overcome the four-cell block and support development to the blastocyst stage. In contrast, blastocyst development of in vitro-produced (IVP) embryos in these culture media varies significantly. Significant differences in morphology and numbers of cells have been observed in in vitro-produced blastocysts compared with in vivo-derived blastocysts. Surgical transfer of in vitro-produced embryos to recipient animals has resulted in acceptable pregnancy rates with moderate litter sizes. Although several systems are available for the generation of in vitro-produced embryos, the problems of polyspermy and poor embryo survival prevent large-scale production of embryos. Further research should be directed to improve oocyte and embryo quality, and to develop methods to minimize polyspermy through development of better IVM, IVF and embryo culture techniques.

  5. Embryo fossilization is a biological process mediated by microbial biofilms

    PubMed Central

    Raff, Elizabeth C.; Schollaert, Kaila L.; Nelson, David E.; Donoghue, Philip C. J.; Thomas, Ceri-Wyn; Turner, F. Rudolf; Stein, Barry D.; Dong, Xiping; Bengtson, Stefan; Huldtgren, Therese; Stampanoni, Marco; Chongyu, Yin; Raff, Rudolf A.

    2008-01-01

    Fossilized embryos with extraordinary cellular preservation appear in the Late Neoproterozoic and Cambrian, coincident with the appearance of animal body fossils. It has been hypothesized that microbial processes are responsible for preservation and mineralization of organic tissues. However, the actions of microbes in preservation of embryos have not been demonstrated experimentally. Here, we show that bacterial biofilms assemble rapidly in dead marine embryos and form remarkable pseudomorphs in which the bacterial biofilm replaces and exquisitely models details of cellular organization and structure. The experimental model was the decay of cleavage stage embryos similar in size and morphology to fossil embryos. The data show that embryo preservation takes place in 3 distinct steps: (i) blockage of autolysis by reducing or anaerobic conditions, (ii) rapid formation of microbial biofilms that consume the embryo but form a replica that retains cell organization and morphology, and (iii) bacterially catalyzed mineralization. Major bacterial taxa in embryo decay biofilms were identified by using 16S rDNA sequencing. Decay processes were similar in different taphonomic conditions, but the composition of bacterial populations depended on specific conditions. Experimental taphonomy generates preservation states similar to those in fossil embryos. The data show how fossilization of soft tissues in sediments can be mediated by bacterial replacement and mineralization, providing a foundation for experimentally creating biofilms from defined microbial species to model fossilization as a biological process. PMID:19047625

  6. Preimplantation death of xenomitochondrial mouse embryo harbouring bovine mitochondria.

    PubMed

    Kawahara, Manabu; Koyama, Shiori; Iimura, Satomi; Yamazaki, Wataru; Tanaka, Aiko; Kohri, Nanami; Sasaki, Keisuke; Takahashi, Masashi

    2015-09-29

    Mitochondria, cellular organelles playing essential roles in eukaryotic cell metabolism, are thought to have evolved from bacteria. The organization of mtDNA is remarkably uniform across species, reflecting its vital and conserved role in oxidative phosphorylation (OXPHOS). Our objectives were to evaluate the compatibility of xenogeneic mitochondria in the development of preimplantation embryos in mammals. Mouse embryos harbouring bovine mitochondria (mtB-M embryos) were prepared by the cell-fusion technique employing the haemagglutinating virus of Japan (HVJ). The mtB-M embryos showed developmental delay at embryonic days (E) 3.5 after insemination. Furthermore, none of the mtB-M embryos could implant into the maternal uterus after embryo transfer, whereas control mouse embryos into which mitochondria from another mouse had been transferred developed as well as did non-manipulated embryos. When we performed quantitative PCR (qPCR) of mouse and bovine ND5, we found that the mtB-M embryos contained 8.3% of bovine mitochondria at the blastocyst stage. Thus, contamination with mitochondria from another species induces embryonic lethality prior to implantation into the maternal uterus. The heteroplasmic state of these xenogeneic mitochondria could have detrimental effects on preimplantation development, leading to preservation of species-specific mitochondrial integrity in mammals.

  7. In vitro production of embryos in South American camelids.

    PubMed

    Trasorras, V; Giuliano, S; Miragaya, M

    2013-01-10

    Studies in reproductive biotechnology techniques have been minimal in South American camelids (SAC). Complex reproductive characteristics of these species contribute to slow progress. Nevertheless, some techniques, such as in vitro fertilization, intracytoplasmic sperm injection and nuclear transfer have been applied and have produced advances in knowledge on embryo environment and in vitro conditions necessary for development. Embryo production may have a high impact in both domestic and wild camelids population. Studies addressed to improve in vitro embryo production and oocyte collection could be a potential key to develop IVF and embryo production as a routine procedure in camelids.

  8. Myosin II Dynamics during Embryo Morphogenesis

    NASA Astrophysics Data System (ADS)

    Kasza, Karen

    2013-03-01

    During embryonic morphogenesis, the myosin II motor protein generates forces that help to shape tissues, organs, and the overall body form. In one dramatic example in the Drosophila melanogaster embryo, the epithelial tissue that will give rise to the body of the adult animal elongates more than two-fold along the head-to-tail axis in less than an hour. This elongation is accomplished primarily through directional rearrangements of cells within the plane of the tissue. Just prior to elongation, polarized assemblies of myosin II accumulate perpendicular to the elongation axis. The contractile forces generated by myosin activity orient cell movements along a common axis, promoting local cell rearrangements that contribute to global tissue elongation. The molecular and mechanical mechanisms by which myosin drives this massive change in embryo shape are poorly understood. To investigate these mechanisms, we generated a collection of transgenic flies expressing variants of myosin II with altered motor function and regulation. We found that variants that are predicted to have increased myosin activity cause defects in tissue elongation. Using biophysical approaches, we found that these myosin variants also have decreased turnover dynamics within cells. To explore the mechanisms by which molecular-level myosin dynamics are translated into tissue-level elongation, we are using time-lapse confocal imaging to observe cell movements in embryos with altered myosin activity. We are utilizing computational approaches to quantify the dynamics and directionality of myosin localization and cell rearrangements. These studies will help elucidate how myosin-generated forces control cell movements within tissues. This work is in collaboration with J. Zallen at the Sloan-Kettering Institute.

  9. Crystalline embryos at ice-vapor interfaces

    NASA Technical Reports Server (NTRS)

    Bartley, D. L.

    1976-01-01

    The nucleation of small monolayer ice-like clusters at the basal and prism ice-vapor interfaces is considered. It is found that the basal surfaces prefer triangular embryos with an orientation that reverses from layer to layer, whereas the most stable clusters on the prism surfaces are rectangular in configuration. At any given saturation ratio, the preferred prism clusters are found to have a critical energy of formation significantly lower than that of the basal clusters, basically because of differences in cluster corner free energies.

  10. Patterning of angiogenesis in the zebrafish embryo.

    PubMed

    Childs, Sarah; Chen, Jau-Nian; Garrity, Deborah M; Fishman, Mark C

    2002-02-01

    Little is known about how vascular patterns are generated in the embryo. The vasculature of the zebrafish trunk has an extremely regular pattern. One intersegmental vessel (ISV) sprouts from the aorta, runs between each pair of somites, and connects to the dorsal longitudinal anastomotic vessel (DLAV). We now define the cellular origins, migratory paths and cell fates that generate these metameric vessels of the trunk. Additionally, by a genetic screen we define one gene, out of bounds (obd), that constrains this angiogenic growth to a specific path. We have performed lineage analysis, using laser activation of a caged dye and mosaic construction to determine the origin of cells that constitute the ISV. Individual angioblasts destined for the ISVs arise from the lateral posterior mesoderm (LPM), and migrate to the dorsal aorta, from where they migrate between somites to their final position in the ISVs and dorsal longitudinal anastomotic vessel (DLAV). Cells of each ISV leave the aorta only between the ventral regions of two adjacent somites, and migrate dorsally to assume one of three ISV cell fates. Most dorsal is a T-shaped cell, based in the DLAV and branching ventrally; the second constitutes a connecting cell; and the third an inverted T-shaped cell, based in the aorta and branching dorsally. The ISV remains between somites during its ventral course, but changes to run mid-somite dorsally. This suggests that the pattern of ISV growth ventrally and dorsally is guided by different cues. We have also performed an ENU mutagenesis screen of 750 mutagenized genomes and identified one mutation, obd that disrupts this pattern. In obd mutant embryos, ISVs sprout precociously at abnormal sites and migrate anomalously in the vicinity of ventral somite. The dorsal extent of the ISV is less perturbed. Precocious sprouting can be inhibited in a VEGF morphant, but the anomalous site of origin of obd ISVs remains. In mosaic embryos, obd somite causes adjacent wild

  11. Fusion of blastomeres in mouse embryos under the action of femtosecond laser radiation. Efficiency of blastocyst formation and embryo development

    NASA Astrophysics Data System (ADS)

    Osychenko, A. A.; Zalesskii, A. D.; Krivokharchenko, A. S.; Zhakhbazyan, A. K.; Ryabova, A. V.; Nadtochenko, V. A.

    2015-05-01

    Using the method of femtosecond laser surgery we study the fusion of two-cell mouse embryos under the action of tightly focused femtosecond laser radiation with the fusion efficiency reaching 60%. The detailed statistical analysis of the efficiency of blastomere fusion and development of the embryo up to the blastocyst stage after exposure of the embryos from different mice to a femtosecond pulse is presented. It is shown that the efficiency of blastocyst formation essentially depends on the biological characteristics of the embryo, namely, the strain and age of the donor mouse. The possibility of obtaining hexaploid embryonal cells using the methods of femtosecond laser surgery is demonstrated.

  12. Avian genetic resource banking: can fish embryos yield any clues for bird embryos?

    PubMed

    Hagedorn, M

    2006-02-01

    Cryopreservation of avian germplasm is becoming better understood and more commonly practiced. However, one area that would be of great benefit for genome resource banking is the preservation of avian embryos. Little is know about the cryobiology of avian embryos, and they have never been successfully cryopreserved. However, it is likely that they share many of the challenges of other yolk-filled multicompartmental embryos. For example, the fish embryo has 1) a large overall size, resulting in a low surface-to-volume ratio, which retards water and cryoprotectant efflux/influx; 2) large-sized cells, such as the yolk, which could increase the likelihood of membrane disruption by intracellular ice formation; 3) compartments, such as the blastoderm and yolk, with differing permeability properties; and 4) susceptibility to chilling injury. Both the avian and fish systems share many physical and anatomical properties, and it is predicted that some of the same permeability barriers would exist in both as well. Although the systems are similar, some of the goals, and thus the practices, to protect the genome may be quite different. One of these major goals in avian developmental biology is to produce chicken:chicken transgenic animals, especially those with germ line transmission. Producing efficient germ line transmissions and being able to cryopreserve these transmissions would be extremely beneficial to both basic and agricultural science. This could be accomplished through the cryopreservation of embryonic gonadal tissue followed by grafting into a host. The gonadal/tail-graft system would provide an advantage for cryopreservation because it is small (in comparison with the whole embryo), has fairly uniform tissue, and contains the essential primordial germ line cells capable of recreating the genetic line of interest. Moreover, because the chicken is such a robust model for most other avian species, the cryopreservation of the gonadal/tail-graft may potentially open

  13. Sex and PRNP genotype determination in preimplantation caprine embryos.

    PubMed

    Guignot, F; Perreau, C; Cavarroc, C; Touzé, J-L; Pougnard, J-L; Dupont, F; Beckers, J-F; Rémy, B; Babilliot, J-M; Bed'Hom, B; Lamorinière, J M; Mermillod, P; Baril, G

    2011-08-01

    The objective of this study was to test the accuracy of genotype diagnosis after whole amplification of DNA extracted from biopsies obtained by trimming goat embryos and to evaluate the viability of biopsied embryos after vitrification/warming and transfer. Whole genome amplification (WGA) was performed using Multiple Displacement Amplification (MDA). Sex and prion protein (PRNP) genotypes were determined. Sex diagnosis was carried out by PCR amplification of ZFX/ZFY and Y chromosome-specific sequences. Prion protein genotype determination was performed on codons 142, 154, 211, 222 and 240. Embryos were collected at day 7 after oestrus and biopsied either immediately after collection (blastocysts and expanded blastocysts) or after 24 h of in vitro culture (compacted morulae). Biopsied embryos were frozen by vitrification. Vitrified whole embryos were kept as control. DNA of biopsies was extracted and amplified using MDA. Sex diagnosis was efficient for 97.4% of biopsies and PRNP genotyping was determined in 78.7% of biopsies. After embryo transfer, no significant difference was observed in kidding rate between biopsied and vitrified control embryos, whereas embryo survival rate was different between biopsied and whole vitrified embryos (p = 0.032). At birth, 100% of diagnosed sex and 98.2% of predetermined codons were correct. Offspring PRNP profiles were in agreement with parental genotype. Whole genome amplification with MDA kit coupled with sex diagnosis and PRNP genotype predetermination are very accurate techniques to genotype goat embryos before transfer. These novel results allow us to plan selection of scrapie-resistant genotypes and kid sex before transfer of cryopreserved embryo.

  14. Studies of In Vitro Embryo Culture of Guppy (Poecilia reticulata).

    PubMed

    Liu, LiLi; Lee, Ki-Young

    2014-09-01

    Different with other fishes, the guppies (Poecilia reticulata) is ovoviviparity, which retain their fertilized eggs within the follicle throughout gestation. The synchronously growing diplotene oocytes store nutrients in droplets and yolk, before their maturation and fertilization. The lecithotrophic strategy of development entails the provisioning of embryos with resources from the maternal yolk deposit rather than from a placenta, it allows the extracorporeal culture of guppy embryo. Studies on their early development of live bearers like the guppy including lineage tracing and genetic manipulations, have been limited. Therefore, to optimize conditions of embryo in vitro culture, explanted embryos from pregnant females were incubated in embryo medium (L-15 medium, supplemented with 5, 10, 15, 20% fetal bovine serum, respectively). We investigated whether the contents of FBS in vitro culture medium impact the development of embryos, and whether they would hatch in vitro. Our study found that in 5% of FBS of the medium, although embryos developed significantly slower in vitro than in the ovary, it was impossible to exactly quantify the developmental delay in culture, due to the obvious spread in developmental stage within each batch of eggs, and embryos can only be maintained until the early-eyed. And although in culture with 20% FBS the embryos can sustain rapid development of early stage, but cannot be cultured for the entire period of their embryonic development and ultimately died. In the medium with 10% and 15% FBS, the embryos seems well developed, even some can continue to grow after follicle ruptures until it can be fed. We also observed that embryonic in these two culture conditions were significantly different in development speed, in 15% it is faster than 10%. But 10% FBS appears to be more optimizing condition than 15% one on development process of embryos and survival rate to larvae stage.

  15. Culture of bovine embryos on a polydimethylsiloxane (PDMS) microwell plate.

    PubMed

    Akagi, Satoshi; Hosoe, Misa; Matsukawa, Kazutsugu; Ichikawa, Akihiko; Tanikawa, Tamio; Takahashi, Seiya

    2010-08-01

    We fabricated a polydimethylsiloxane (PDMS)-based microwell plate (PDMS-MP) containing 100 microwells with a rounded bottom and examined whether it can be used for culture of individual in vitro fertilized (IVF) embryos or parthenogenetically activated zona-free embryos in cattle. In Experiment 1, we examined the in vitro developmental ability of IVF embryos cultured individually on PDMS-MP. After IVF, 20 embryos were transferred into 100 microl drops on PDMS-MP and cultured individually in each well of PDMS-MP (PDMS group). After 7 days of culture, the embryos in the PDMS group developed to the blastocyst stage at the same rate of those in the control group cultured in a group of 20 embryos without PDMS-MP. There were no differences in total number of cells and the ratio of inner cell mass to total cells between the PDMS and control groups. In Experiment 2, we examined the in vitro developmental ability of parthenogenetically activated zona-free bovine embryos cultured individually on PDMS-MP. The zona-free embryos were cultured individually in each well of a PDMS-MP or in each well produced by pressing a darning needle onto the bottom of a culture dish (WOW group). After 7 days of culture, the blastocyst formation rate and cell number of blastocysts in the PDMS group did not differ from those of the zona-intact embryos in the control group. Also, there were no differences in the blastocyst formation rate and cell number of blastocysts between the WOW and PDMS groups. These results suggest that the culture system using PDMS-MP is useful for individual embryos or zona-free embryos in cattle.

  16. Macroevolutionary developmental biology: Embryos, fossils, and phylogenies.

    PubMed

    Organ, Chris L; Cooper, Lisa Noelle; Hieronymus, Tobin L

    2015-10-01

    The field of evolutionary developmental biology is broadly focused on identifying the genetic and developmental mechanisms underlying morphological diversity. Connecting the genotype with the phenotype means that evo-devo research often considers a wide range of evidence, from genetics and morphology to fossils. In this commentary, we provide an overview and framework for integrating fossil ontogenetic data with developmental data using phylogenetic comparative methods to test macroevolutionary hypotheses. We survey the vertebrate fossil record of preserved embryos and discuss how phylogenetic comparative methods can integrate data from developmental genetics and paleontology. Fossil embryos provide limited, yet critical, developmental data from deep time. They help constrain when developmental innovations first appeared during the history of life and also reveal the order in which related morphologies evolved. Phylogenetic comparative methods provide a powerful statistical approach that allows evo-devo researchers to infer the presence of nonpreserved developmental traits in fossil species and to detect discordant evolutionary patterns and processes across levels of biological organization.

  17. Twinning of amphibian embryos by centrifugation

    NASA Technical Reports Server (NTRS)

    Black, S. D.

    1984-01-01

    In the frog Xenopus laevis, the dorsal structures of the embryonic body axis normally derive from the side of the egg opposite the side of sperm entry. However, if the uncleaved egg is inclined at lg or centrifuged in an inclined position, this topographic relationship is overridden: the egg makes its dorsal axial structures according to its orientation in the gravitational/centrifugal field, irrespective of the position of sperm entry. Certain conditions of centrifugation cause eggs to develop into conjoined twins with two sets of axial structures. A detailed analysis of twinning provided some insight into experimental axis orientation. First, as with single-axis embryos, both axes in twins are oriented according to the direction of centrifugation. One axis forms at the centripetal side of the egg and the other forms at the centrifugal side, even when the side of sperm entry is normal to the centrifugal force vector. Second, if eggs are centrifuged to give twins, but are inclined at lg to prevent post-centrifugation endoplasmic redistributions, only single-axis embryos develop. Thus, a second redistribution is required for high-frequency secondary axis formation. This can be accomplished by lg (as in the single centrifugations) or by a second centrifugation directed along the egg's animal-vegetal axis.

  18. Epigenetics in fertilization and preimplantation embryo development.

    PubMed

    Rivera, Rocio Melissa; Ross, Jason Wayne

    2013-12-01

    Epigenetic reprogramming of the parental genomes upon fertilization is required for proper embryonic development. It has long been appreciated that asymmetric distribution of histone modifications as well as differences in the level of DNA methylation exist between the parental pronuclei in mammalian zygotes and during preimplantation development. The speed at which the paternal genome is demethylated after entering the oocyte and the fact that rapid demethylation occurs in the absence of DNA replication have led many to hypothesize that a DNA demethylase must exist. However, such an enzyme has not been found. That the genome of mammalian preimplantation embryos undergo a wave of global demethylation was first reported 25 years ago but only in the past three years has data surfaced that can partially explain the elusive nature of this phenomenon. In addition to the global reorganization of the methylation and histone modification patterns, oocyte development prior to germinal vesicle breakdown involves the production of numerous small RNA, including miRNA. Despite their presence, miRNA functional activity is thought to be limited in the mature mouse oocyte. Additionally, molecular signatures in the 3' untranslated region of maternally expressed transcripts may impact mRNA stability during the transcriptionally quiescent period following germinal vesicle breakdown and prior to the maternal to zygote transition. In this review, we reference some of the recent works which attempt to shed light into the importance of the dynamic epigenetic landscape observed during oocyte maturation and preimplantation embryo development in mammals.

  19. Manipulating claudin expression in avian embryos.

    PubMed

    Collins, Michelle M; Ryan, Aimee K

    2011-01-01

    Since the discovery of Claudin-1 and -2 by Tsukita and colleagues in the late 1990s [Furuse et al. J Cell Biol 141:1539-50,1998], claudin family members have been found to have critical roles in maintaining the integrity of epithelial and endothelial tight junctions [Furuse and Moriwaki Ann N Y Acad Sci 1165:58-61, 2009; Morita et al. Proc Natl Acad Sci USA 96:511-6, 1999; Tsukita and Furuse Ann N Y Acad Sci 915:129-35, 2000; Turksen and Troy J Cell Sci 117:2435-47, 2004]. The properties of distinct claudin family members in tight junction permeability and specificity have been extensively studied in vitro using cell culture models. In vivo, claudin family members are dynamically regulated during embryogenesis and alterations in their expression patterns can have detrimental effects on the formation and physiological function of the tissues in which they are expressed. The chick embryo provides an excellent system to dissect the roles of specific family members in vivo and to explore the effects of modulating claudin expression during the epithelial-to-mesenchymal and mesenchymal-to-epithelial transitions that are associated with tissue morphogenesis and differentiation. We are using the chick embryo to understand the roles of the claudin family of tight junction proteins during gastrulation and left-right patterning during embryogenesis. Here, we describe methodologies for manipulating claudin gene expression in specific target tissues during chick embryogenesis.

  20. Embryo cryopreservation and in vitro culture of preimplantation embryos in Campbell's hamster (Phodopus campbelli).

    PubMed

    Amstislavsky, Sergei; Brusentsev, Eugeny; Kizilova, Elena; Igonina, Tatyana; Abramova, Tatyana; Rozhkova, Irina

    2015-04-01

    The aims of this study were to compare different protocols of Campbell's hamster (Phodopus campbelli) embryos freezing-thawing and to explore the possibilities of their in vitro culture. First, the embryos were flushed from the reproductive ducts 2 days post coitum at the two-cell stage and cultured in rat one-cell embryo culture medium (R1ECM) for 48 hours. Most (86.7%) of the two-cell embryos developed to blastocysts in R1ECM. Second, the embryos at the two- to eight-cell stages were flushed on the third day post coitum. The eight-cell embryos were frozen in 0.25 mL straws according to standard procedures of slow cooling. Ethylene glycol (EG) was used either as a single cryoprotectant or in a mixture with sucrose. The survival of frozen-thawed embryos was assessed by double staining with fluorescein diacetate and propidium iodide. The use of EG as a single cryoprotectant resulted in fewer alive embryos when compared with control (fresh embryos), but combined use of EG and sucrose improved the survival rate after thawing. Furthermore, granulocyte-macrophage colony-stimulating factor rat (2 ng/mL) improved the rate of the hamster frozen-thawed embryo development in vitro by increasing the final cell number and alleviating nuclear fragmentation. Our data show the first attempt in freezing and thawing Campbell's hamster embryos and report the possibility of successful in vitro culture for this species in R1ECM supplemented with granulocyte-macrophage colony-stimulating factor.

  1. Early embryo development in Fucus distichus is auxin sensitive

    NASA Technical Reports Server (NTRS)

    Basu, Swati; Sun, Haiguo; Brian, Leigh; Quatrano, Ralph L.; Muday, Gloria K.

    2002-01-01

    Auxin and polar auxin transport have been implicated in controlling embryo development in land plants. The goal of these studies was to determine if auxin and auxin transport are also important during the earliest stages of development in embryos of the brown alga Fucus distichus. Indole-3-acetic acid (IAA) was identified in F. distichus embryos and mature tissues by gas chromatography-mass spectroscopy. F. distichus embryos accumulate [(3)H]IAA and an inhibitor of IAA efflux, naphthylphthalamic acid (NPA), elevates IAA accumulation, suggesting the presence of an auxin efflux protein complex similar to that found in land plants. F. distichus embryos normally develop with a single unbranched rhizoid, but growth on IAA leads to formation of multiple rhizoids and growth on NPA leads to formation of embryos with branched rhizoids, at concentrations that are active in auxin accumulation assays. The effects of IAA and NPA are complete before 6 h after fertilization (AF), which is before rhizoid germination and cell division. The maximal effects of IAA and NPA are between 3.5 and 5 h AF and 4 and 5.5 h AF, respectively. Although, the location of the planes of cell division was significantly altered in NPA- and IAA-treated embryos, these abnormal divisions occurred after abnormal rhizoid initiation and branching was observed. The results of this study suggest that auxin acts in the formation of apical basal patterns in F. distichus embryo development.

  2. CULTURING RABIES (HYDROPHOBIA) VIRUSES IN A DEVELOPING CHICKEN EMBRYO

    DTIC Science & Technology

    This report states that 19 successive passages of the rabies virus strain 83 through the brain of a chicken embryo and 6 passages through the yolk...sac are feasible. In the process of cultivation of the rabies virus in the organism of chicken embryo there was a variation-fixation of it; shortening

  3. Permeability barriers to embryo cryopreservation of Pectinophora gossypiella (Lepidoptera: Gelechiidae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The aim of this study was to develop a method to cryopreserve the embryos of the pink bollworm moth, Pectinophora gossypiella (Saunders). Previously developed dipteran cryopreservation protocols were not directly adaptable to use with the embryos of this lepidopteran species. Physiochemical and ele...

  4. An Arabidopsis thaliana embryo arrest mutant exhibiting germination potential

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The ability to initiate radicle elongation, or germination potential, occurs in developing embryos before the completion of seed maturation. Green embryos after walking-stick stage in developing Arabidopsis thaliana seeds germinate when excised from seeds and incubated in MS media containing 1 % suc...

  5. Storage oil breakdown during embryo development of Brassica napus (L.).

    PubMed

    Chia, Tansy Y P; Pike, Marilyn J; Rawsthorne, Stephen

    2005-05-01

    In this study it is shown that at least 10% of the major storage product of developing embryos of Brassica napus (L.), triacylglycerol, is lost during the desiccation phase of seed development. The metabolism of this lipid was studied by measurements of the fate of label from [1-(14)C]decanoate supplied to isolated embryos, and by measurements of the activities of enzymes of fatty acid catabolism. Measurements on desiccating embryos have been compared with those made on embryos during lipid accumulation and on germinating seedlings. Enzymes of beta-oxidation and the glyoxylate cycle, and phosphoenolpyruvate carboxykinase were present in embryos during oil accumulation, and increased in activity and abundance as the seeds matured and became desiccated. Although the activities were less than those measured during germination, they were at least comparable to the in vivo rate of fatty acid synthesis in the embryo during development. The pattern of labelling, following metabolism of decanoate by isolated embryos, indicated a much greater involvement of the glyoxylate cycle during desiccation than earlier in oil accumulation, and showed that much of the (14)C-label from decanoate was released as CO(2) at both stages. Sucrose was not a product of decanoate metabolism during embryo development, and therefore lipid degradation was not associated with net gluconeogenic activity. These observations are discussed in the context of seed development, oil yield, and the synthesis of novel fatty acids in plants.

  6. Developmental ability of trophoblast stem cells in uniparental mouse embryos.

    PubMed

    Ogawa, H; Shindo, N; Kumagai, T; Usami, Y; Shikanai, M; Jonwn, K; Fukuda, A; Kawahara, M; Sotomaru, Y; Tanaka, S; Arima, T; Kono, T

    2009-05-01

    Neither parthenogenetic (PG) nor androgenetic (AG) mouse embryos survive after day 9.5 of pregnancy, owing to the inadequate growth of extraembryonic tissues, including the placenta. At day 9.5 of pregnancy, the placental structures are poorly developed in PG embryos, while trophoblast giant cells are abundant at the implantation site in AG embryos. These findings suggest that both parental genomes are required for placental development. To gain further insight into the trophoblast lineage in PG and AG embryos, we attempted to derive trophoblast stem (TS)-like cell lines from uniparental embryos. Furthermore, we sought to assess their ability to differentiate into cells of the trophoblast lineage by using gene expression analysis. Three cell lines that expressed marker genes for undifferentiated TS cells (Cdx2 and Errbeta) were derived from AG embryos. Under differentiation conditions, these cells expressed the trophoblast giant cell-specific genes, but did not express the spongiotrophoblast-specific genes. In contrast, none of the four cell lines from PG embryos expressed marker genes for undifferentiated TS cells, but they expressed Oct3/4, a marker gene for embryonic stem cells. Immunohistochemical analysis indicated that PG blastocysts expressed Oct3/4 and Cdx2 specifically in inner cell mass and the trophectoderm respectively. These results suggest that PG embryos do not possess TS cells, because of the lack of the developmental ability of trophoblast cells.

  7. Cryopreservation of embryos of Lucilia sericata (Diptera: Calliphoridae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Embryos of Lucilia (Phaenicia) sericata (Meigen) (Diptera: Calliphoridae), the green blowfly, were successfully cryopreserved by vitrification in liquid nitrogen and stored for 8 yr. Embryos incubated at 19 deg. C for 17 h after oviposition were found to be the most appropriate stage to cryopreserve...

  8. The human embryo in the Christian tradition: a reconsideration.

    PubMed

    Jones, D A

    2005-12-01

    Recent claims that the Christian tradition justifies destructive research on human embryos have drawn upon an article by the late Professor Gordon Dunstan which appeared in this journal in 1984. Despite its undoubted influence, this article was flawed and seriously misrepresented the tradition of Christian reflection on the moral status of the human embryo.

  9. Growth hormone inhibits apoptosis in in vitro produced bovine embryos.

    PubMed

    Kölle, Sabine; Stojkovic, Miodrag; Boie, Gudrun; Wolf, Eckhard; Sinowatz, Fred

    2002-02-01

    Growth hormone (GH) has recently been shown to exert distinct effects on the differentiation and metabolism of early embryos. Up to now, however, it is not clear whether GH is able to modulate apoptosis during early embryogenesis. Differential cell staining of 8-day-old bovine embryos cultured with 100 ng bovine recombinant GH (rbGH) per ml medium (synthetic oviduct fluid-polyvinylalcohol) demonstrated that GH significantly increased the number of inner cell mass (ICM) and trophectoderm cells in bovine expanded blastocysts. As shown by terminal deoxynucleotidyl transferase mediated dUTP labeling (TUNEL) supplementation of bGH decreased the percentage of 8-day-old embryos showing at least one apoptotic cell from 58 to 21%. The percentage of apoptotic cells in one blastocyst was significantly (P < 0.01) reduced from 4.6 to 1.1% by GH treatment. Incubation of the embryos with 150 mM vanillylnonanamide induced apoptosis in all embryos. Whereas in control embryos 14% of the embryonic cells were TUNEL-positive, the percentage of apoptotic cells declined to 2.7% in the GH treated embryos. Expression of immunoreactive bcl-2 in blastocysts was not affected by GH treatment. Synthesis of the bax protein which is known to promote apoptosis was reduced in embryos cultured with GH. Our results suggest that GH acts as survival factor during in vitro culture and reduces apoptosis by altering the bax to bcl-2 ratio during early embryogenesis.

  10. Freezing injuries in the embryos of Piaractus mesopotamicus.

    PubMed

    Fornari, Darci Carlos; Ribeiro, Ricardo Pereira; Streit, Danilo Pedro; Vargas, Lauro; Barrero, Nelson M Lopera; de Moraes, Gentil Vanini

    2011-11-01

    Cryopreservation of mammal embryos has been technically feasible for many years, but morphological injuries still persist in fish embryos during cryopreservation. Thus, the objective of the present study was to describe these freezing injuries in Piaractus mesopotamicus embryos. Two hundred and twenty-five embryos were collected at the post-gastrula stage and assigned into four treatments of sucrose at 8.5, 17.0, 25.0 or 34.0% plus 9.0% methanol. The control was prepared with distilled water only. The gradual decrease in the temperature was 0.5°C/min. After the seeding stage, the fish embryos were stored in liquid nitrogen at -33°C. Thereafter, they were thawed for evaluating per cent hatching, and the samples collected from every treatment were submitted to scanning electron microscopy for morphological analysis. The micrographic images showed that there was substantial alterations in embryo morphology under the highest concentrations of sucrose. These solutions did not prevent the formation of ice crystals, which lead to deformities and killed the embryos, but the observed reduced level of morphological structure in these embryos when treated with 17.0% sucrose plus 9.0% methanol is a compelling argument for additional studies.

  11. The gastrocoel roof plate in embryos of different frogs.

    PubMed

    Sáenz-Ponce, Natalia; Santillana-Ortiz, Juan-Diego; del Pino, Eugenia M

    2012-02-01

    The morphology of the gastrocoel roof plate and the presence of cilia in this structure were examined in embryos of four species of frogs. Embryos of Ceratophrys stolzmanni (Ceratophryidae) and Engystomops randi (Leiuperidae) develop rapidly, provide comparison for the analysis of gastrocoel roof plate development in the slow-developing embryos of Epipedobates machalilla (Dendrobatidae) and Gastrotheca riobambae (Hemiphractidae). Embryos of the analyzed frogs develop from eggs of different sizes, and display different reproductive and developmental strategies. In particular, dorsal convergence and extension and archenteron elongation begin during gastrulation in embryos of rapidly developing frogs, as in Xenopus laevis. In contrast, cells that involute during gastrulation are stored in the large circumblastoporal collar that develops around the closed blastopore in embryos of slow-developing frogs. Dorsal convergence and extension only start after blastopore closure in slow-developing frog embryos. However, in the neurulae, a gastrocoel roof plate develops, despite the accumulation of superficial mesodermal cells in the circumblastoporal collar. Embryos of all four species develop a ciliated gastrocoel roof plate at the beginning of neurulation. Accordingly, fluid-flow across the gastrocoel roof plate is likely the mechanism of left-right asymmetry patterning in these frogs, as in X. laevis and other vertebrates. A ciliated gastrocoel roof plate, with a likely origin as superficial mesoderm, is conserved in frogs belonging to four different families and with different modes of gastrulation.

  12. Use of blue crab (Callinectes sapidus) embryos for toxicity testing

    SciTech Connect

    Lee, R.; O`Malley, K.

    1995-12-31

    After fertilization, blue crab embryos develop in egg sacs attached to the female pleopods, often referred to as the sponge. Lipovitellin and lipid droplets in the egg sacs provide energy and nutrition for the developing embryos. Embryos were removed from the sponge and transferred to 24 well culture plates containing sea water with or without toxicants, Each well contained 10 embryos. After 7 to 10 days, embryos hatched to swimming zoea. The effects of toxicants at various concentrations on hatching were determined and the EC{sub 50} calculated. For example, the EC{sub 50} for tributyltin, fenvalerate and mercuric chloride were 50, 30 and 90 ng/liter, respectively. The hatching success of control embryos ranged from 95 to 98%. Formation of the heart, eyespot formation, appendage formation and utilization rate of lipovitellin were also effected by exposure to toxicants. At a low concentration of mercuric ion (30ng/liter) the heart formed, but there was no heart beat. Eyespot formation was abnormal in the presence of high concentrations of cadmium (2 {micro}g/liter) and zinc (5 {micro}g/liter), Crab embryos offer many advantages for toxicity testing of pure compounds or mixtures in water, including toxicity testing of sediment pore water. The crab embryos may also serve as models to understand the effect of specific toxicants on the heart and eye spots of crustaceans.

  13. Morphological and cytogenetic assessment of cleavage and blastocyst stage embryos.

    PubMed

    Fragouli, E; Alfarawati, S; Spath, K; Wells, D

    2014-02-01

    Morphological assessments are the main way in which fertility clinics select in vitro generated embryo(s) for transfer to the uterus. However, it is widely acknowledged that the microscopic appearance of an embryo is only weakly correlated with its viability. Furthermore, the extent to which morphology is affected by aneuploidy, a genetic defect common in human preimplantation embryos, remains unclear. Aneuploidy is of great relevance to embryo selection as it represents one of the most important causes of implantation failure and miscarriage. The current study aimed to examine whether morphological appearance can assist in identifying embryos at risk of aneuploidy. Additionally, the data produced sheds light on how chromosomal anomalies impact development from the cleavage to the blastocyst stage. A total of 1213 embryos were examined. Comprehensive chromosome analysis was combined with well-established criteria for the assessment of embryo morphology. At the cleavage stage, chromosome abnormalities were common even amongst embryos assigned the best morphological scores, indicating that aneuploidy has little effect on microscopic appearance at fixed time points up until Day 3 of development. However, at the blastocyst stage aneuploidies were found to be significantly less common among embryos of optimal morphological quality, while such abnormalities were overrepresented amongst embryos considered to be of poor morphology. Despite the link between aneuploidy and blastocyst appearance, many chromosomally abnormal embryos were able to achieve the highest morphological scores. In particular, blastocysts affected by forms of aneuploidy with the greatest capacity to produce clinical pregnancies (e.g. trisomy 21) were indistinguishable from euploid embryos. The sex ratio was seen to be equal throughout preimplantation development. Interestingly, however, females were overrepresented amongst the fastest growing cleavage-stage embryos, whereas a sex-related skew in the

  14. Embryo apoptosis identification: Oocyte grade or cleavage stage?

    PubMed Central

    Bakri, Noraina Mohd; Ibrahim, Siti Fatimah; Osman, Nurul Atikah; Hasan, Nurhaslina; Jaffar, Farah Hanan Fathihah; Rahman, Zulaiha Abdul; Osman, Khairul

    2015-01-01

    Apoptosis is a programed cell death that is vital for tissue homeostasis. However, embryo apoptosis had been known to be related to embryo fragmentation which should be avoided in in vitro fertilization (IVF). The purpose of this study was to evaluate the relationship of embryo apoptosis with the grade of immature oocytes and cleavage stage of in vitro produced (IVP) cattle embryos. This study consisted of 345 oocytes collected through ovary slicing. Immature oocytes were graded as A, B and C. This grading was based on cumulus cell thickness and compactness. All oocytes then underwent an in vitro maturation (IVM) procedure. An IVF was done 24 h after IVM culture. Prior to staining, stage of cleaved embryos was determined and classified as either 2, 4, 8 or >8-cell embryo stage. Apoptosis status of cleaved IVP embryos was determined by using annexin V-FITC staining technique at 48 and 72 h post insemination (hpi). Apoptosis status for each embryo was classified as either early or late. The result showed that there was no significant difference (p > 0.05) of apoptosis status among grade A, B and C embryos. All grades of oocytes showed embryo apoptosis where 1.5% late apoptosis for grade A, 4.5% and 10.4% of early and late apoptosis for grade B and grade C. Early apoptosis was not seen in grade A embryo. We also noted no significant difference (p > 0.05) of apoptosis status between 2, 4, 8 and >8-cell embryo stage. Early apoptosis was also not seen in >8-cell stage. Even though there were no differences in apoptosis expression between the three classes, the cleavage rate of grade A oocytes was significantly higher (p < 0.01) than grade B and grade C. In conclusion, the apoptosis expression in the embryo can occur regardless of the oocyte quality and the cleavage stage of the embryo produced. PMID:26858565

  15. Human oocyte cryopreservation: a valid alternative to embryo cryopreservation?

    PubMed

    Tucker, Michael; Morton, Paula; Liebermann, Juergen

    2004-04-05

    Embryo cryopreservation has become an ethical necessity due to the way human in vitro fertilization (IVF) infertility therapy has developed. Limited embryonic implantation has by necessity driven IVF therapy to adopt ways to maximize the harvest of oocytes following ovarian hyperstimulation with its attendant risks. Collection of more oocytes has allowed more embryos to be generated to compensate for poor embryonic viability, often leading to transfer of multiple embryos to increase per transfer pregnancy rates. In an era of improving embryonic viability and prevailing trend toward single embryo transfers, production of excessive numbers of surplus embryos appears increasingly inappropriate. At which stage embryo cryopreservation can be undertaken most effectively remains controversial. Embryo cryopreservation nevertheless represents the current solution to the problem of excessive embryo production, but inherently raises ethical concerns for certain couples uncomfortable with what they might perceive to be "experimental" cryostorage, who in extreme circumstances may even choose to limit the number of oocytes inseminated to obviate the production of spare embryos. On a more practical level, cryostored embryos are co-owned by two people who may separate, and as such the embryos then face an uncertain fate, commonly decided in courts of law. Oocyte cryopreservation, if consistent and successful, offers a way to avoid the above complications of routine IVF therapy. Oocytes may need to be cryostored in the event of unforeseen non-production of sperm during IVF therapy, allowing a more measured consideration of donor sperm use or other means of sperm retrieval. Beyond IVF for infertility therapy using a couple's own gametes, oocyte cryopreservation provides a wonderful opportunity to optimize donor oocyte cryo-banking, reducing costs and improving convenience. Meanwhile, frozen embryo donation is an approach that many couples are uncomfortable with, and allows only for

  16. High incubation temperatures enhance mitochondrial energy metabolism in reptile embryos

    PubMed Central

    Sun, Bao-Jun; Li, Teng; Gao, Jing; Ma, Liang; Du, Wei-Guo

    2015-01-01

    Developmental rate increases exponentially with increasing temperature in ectothermic animals, but the biochemical basis underlying this thermal dependence is largely unexplored. We measured mitochondrial respiration and metabolic enzyme activities of turtle embryos (Pelodiscus sinensis) incubated at different temperatures to identify the metabolic basis of the rapid development occurring at high temperatures in reptile embryos. Developmental rate increased with increasing incubation temperatures in the embryos of P. sinensis. Correspondingly, in addition to the thermal dependence of mitochondrial respiration and metabolic enzyme activities, high-temperature incubation further enhanced mitochondrial respiration and COX activities in the embryos. This suggests that embryos may adjust mitochondrial respiration and metabolic enzyme activities in response to developmental temperature to achieve high developmental rates at high temperatures. Our study highlights the importance of biochemical investigations in understanding the proximate mechanisms by which temperature affects embryonic development. PMID:25749301

  17. Toxicity test of xanthone from mangosteen on zebrafish embryos

    NASA Astrophysics Data System (ADS)

    Noordin, Muhammad Akram Mohd; Noor, Mahanem Mat; Kamaruddin, Wan Mohd Aizat Wan; Lazim, Azwan Mat; Fazry, Shazrul

    2016-11-01

    Xanthone is a chemical compound identified in mangosteen pericarp. A previous study showed that xanthone has anti-proliferating effect on cancer cells. In this study we investigate the toxicity level of xanthone in zebrafish embryo to for future reference on other animal model. We employed Fish Embryo Toxicity (FET) assay to determine the toxicity level of different concentrations of xanthone. Embryos were observed at 24, 48 and 72 hours post fertilization (hpf) under microscope at 4× magnification. The extract showed toxicity effect on embryo at concentrations of 250, 125 and 62.5 µg/mL. Concentrations at 15.63, 7.81 and 3.91 µg / mL of xanthone did not harm the embryos and showed 100% of survival.

  18. Sex determination of duck embryos: observations on syrinx development

    USGS Publications Warehouse

    Wilson, Robert E.; Sonsthagen, Sarah A.; Franson, J. Christian

    2013-01-01

    Ducks exhibit sexual dimorphism in vocal anatomy. Asymmetrical ossification of the syrinx (bulla syringealis) is discernable at about 10 days of age in male Pekin duck (Anas platyrhynchos domestica) embryos, but information is lacking on the early development of the bulla in wild ducks. To evaluate the reliability of this characteristic for sexing developing embryos, we examined the syrinx of dead embryos and compared results with molecular sexing techniques in high arctic nesting Common Eiders (Somateria mollissima). Embryos 8 days or older were accurately (100%) sexed based on the presence/absence of a bulla, 2 days earlier than Pekin duck. The use of the tracheal bulla can be a valuable technique when sex identification of embryos or young ducklings is required.

  19. Heartbeat, embryo communication and hatching synchrony in snake eggs

    PubMed Central

    Aubret, Fabien; Blanvillain, Gaëlle; Bignon, Florent; Kok, Philippe J. R.

    2016-01-01

    Communication is central to life at all levels of complexity, from cells to organs, through to organisms and communities. Turtle eggs were recently shown to communicate with each other in order to synchronise their development and generate beneficial hatching synchrony. Yet the mechanism underlying embryo to embryo communication remains unknown. Here we show that within a clutch, developing snake embryos use heart beats emanating from neighbouring eggs as a clue for their metabolic level, in order to synchronise development and ultimately hatching. Eggs of the water snake Natrix maura increased heart rates and hatched earlier than control eggs in response to being incubated in physical contact with more advanced eggs. The former produced shorter and slower swimming young than their control siblings. Our results suggest potential fitness consequences of embryo to embryo communication and describe a novel driver for the evolution of egg-clustering behaviour in animals. PMID:26988725

  20. Chill sensitivity of honey bee, Apis mellifera, embryos.

    PubMed

    Collins, Anita M; Mazur, Peter

    2006-08-01

    Improved methods for preservation of honey bee, Apis mellifera L., germplasm would be very welcome to beekeeping industry queen breeders. The introduction of two parasites and the emergence of an antibiotic resistant disease have increased demands for resistant stock. Techniques for artificial insemination of queens are available, and semen has been cryopreserved with limited success. However, cryopreservation of embryos for rearing queens would mesh well with current practices and also provide drones (haploid males). Eggs at five ages between twenty-four hours and sixty-two hours were exposed to 0, -6.6, and/or -15 degrees C for various times, and successful hatch measured. Honey bee embryos show chill sensitivity as do other insect embryos, and the rate of chill injury increases dramatically with decrease in holding temperature. The 48 h embryos in both groups showed the greatest tolerance to chilling, although 44 h embryos were only slightly less so.

  1. Comparative pathogenicity of avian encephalomyelitis viruses in chicken embryos.

    PubMed

    Miyamae, T

    1975-07-01

    Multiplications of wild, various embryo-adapting and completely embryo-adapted avian encephalomyelitis (AE) viruses in chicken embryos were compared by the fluorescent-antibody technique (FAT). With a wild AE virus, viral antigens were randomly seen in the central nervous system (CNS), appearing least often in the cerebellum. Other organs seldom became test positive, except for heart and kidney. Even with 4 chicken brain-passaged viruses in the process of embryo adaptation, there was little augmentation of antigens except in the alimentary tract. However, the 2 midpassage viruses showed a peculiar localization of antigens in the white matter of the lumbosacral cord, together with the appearance of test-positive spinal ganglion cells. With 2 strains of embryo-adapted AE virus, the antigens appeared first in the spinal ganglion cells and secondly in the lumbosacral cord and then spread to the cerebrum. Subsequently, clinical signs of AE were evident. This peculiar invasion order was a prominent feature.

  2. Cryopreservation of coconut (Cocos nucifera L.) zygotic embryos by vitrification.

    PubMed

    Sajini, K K; Karun, A; Amamath, C H; Engelmann, F

    2011-01-01

    The present study investigates the effect of preculture conditions, vitrification and unloading solutions on survival and regeneration of coconut zygotic embryos after cryopreservation. Among the seven plant vitrification solutions tested, PVS3 was found to be the most effective for regeneration of cryopreserved embryos. The optimal protocol involved preculture of embryos for 3 days on medium with 0.6 M sucrose, PVS3 treatment for 16 h, rapid cooling and rewarming and unloading in 1.2 M sucrose liquid medium for 1.5 h. Under these conditions, 70-80 survival (corresponding to size enlargement and weight gain) was observed with cryopreserved embryos and 20-25 percent of the plants regenerated (showing normal shoot and root growth) from cryopreserved embryos were established in pots.

  3. Effect of supplementation of different growth factors in embryo culture medium with a small number of bovine embryos on in vitro embryo development and quality.

    PubMed

    Ahumada, C J; Salvador, I; Cebrian-Serrano, A; Lopera, R; Silvestre, M A

    2013-03-01

    When embryos are cultured individually or in small groups, blastocyst yield efficiency and quality are usually reduced. The aim of this work was to investigate the effect of supplementation of the embryo culture medium (CM) with several growth factors (GFs) on embryo development and apoptosis rate when a reduced number of embryos were in vitro cultured. Two experimental studies (ES) were carried out. In ES 1, five treatments were tested to study the effect of GF on embryo development: Control (∼30 to 50 embryos cultured in 500 μl of CM); Control 5 (Five embryos cultured in 50 μl microdrops of CM), without addition of GF in either of the two control groups; epidermal GF (EGF); IGF-I; and transforming GF-α (TGF-α) (Five embryos were cultured in 50 μl microdrops of CM with 10 ng/ml EGF, 10 ng/ml IGF-I or 10 ng/ml TGF-α, respectively). In ES 2, following the results obtained in ES 1, four different treatments were tested to study their effect on embryo development and quality (number of cells per blastocyst and apoptotic rate): Control; Control 5; EGF, all three similar to ES 1; EGF + IGF-I group (five embryos cultured in 50 μl microdrops of CM with 10 ng/ml EGF and 10 ng/ml IGF-I). In both ESs, it was observed that a higher proportion of embryos cultured in larger groups achieved blastocyst stage than embryos cultured in reduced groups (22.6% v. 14.0%, 12.6% and 5.3% for Control v. Control 5, IGF-I, TGF-α groups in ES 1, and 24.9% v. 17.1% and 19.0% for Control v. Control 5 and EGF in ES 2, respectively; P < 0.05), with the exception of embryos cultured in medium supplemented with EGF (18.5%) or with EGF + IGF-I (23.5%), in ES 1 and ES 2, respectively. With regard to blastocyst quality, embryos cultured in reduced groups and supplemented with EGF, alone or combined with IGF-I, presented lower apoptosis rates than embryos cultured in reduced groups without GF supplementation (11.6% and 10.5% v. 21.9% for EGF, EGF + IGF-I and Control 5 groups, respectively; P

  4. Embryotoxic effects of chlorobutanol in cultured mouse embryos.

    PubMed

    Smoak, I W

    1993-03-01

    Chlorobutanol (CB) is a commonly used preservative which is added to numerous pharmaceutical preparations, and it is the active ingredient in certain oral sedatives and topical anesthetics. Chlorobutanol has demonstrated adverse effects in adult tissues, but CB has not been previously investigated for its effect on the developing whole embryo. The method of whole-embryo culture was used in this study to expose mouse embryos during two stages of organogenesis to CB at final concentrations of 0 (control), 10, 25, 50, 100, and 200 micrograms/ml. Embryos were evaluated for heart rate (HR), malformations, and somite number, and embryos and visceral yolk sacs (VYSs) were assayed for total protein content as a measure of overall growth. Neurulating (3-6 somite) embryos were malformed and growth retarded by exposure to CB concentrations > or = 25 micrograms/ml, with decreased VYS growth at > or = 50 micrograms/ml and decreased HR at > or = 100 micrograms/ml CB. Early limb-bud stage (20-25 somite) embryos were malformed at CB concentrations > or = 50 micrograms/ml and growth retarded at > or = 100 micrograms/ml, with decreased VYS growth at 200 micrograms/ml and decreased HR at > or = 100 micrograms/ml CB. Thus, CB produces dysmorphogenesis in mouse embryos in vitro, and neurulating embryos are somewhat less sensitive than early limb-bud stage embryos. The concentrations of CB that interfere with normal embryonic development are within the range of human blood levels measured following multiple doses of CB. Preparations containing CB should be used with caution during pregnancy, particularly when repeated dosing may allow accumulation of CB to potentially embryotoxic levels.

  5. PEI1, an embryo-specific zinc finger protein gene required for heart-stage embryo formation in Arabidopsis.

    PubMed Central

    Li, Z; Thomas, T L

    1998-01-01

    We used virtual subtraction, a new gene isolation strategy, to isolate several genes of interest that are expressed in Arabidopsis embryos. These genes have demonstrated biological properties or have the potential to be involved in important biological processes. One gene isolated by virtual subtraction is PEI. It encodes a protein containing a Cys3His zinc finger domain associated with a number of animal and fungal transcription factors. In situ hybridization results showed that PEI1 is expressed throughout the embryo from globular to late cotyledon stage. Transgenic Arabidopsis plants expressing a PEI1 antisense gene produced white seeds in which embryo development did not progress through heart stage. Aberrant embryos failed to form cotyledons, but the embryonic root appeared to be normal. Aberrant embryos did not turn green, and the expression of genes involved in photomorphogenesis was drastically attenuated. In culture, aberrant embryos did not form true leaves, but root formation was apparently normal. These results suggest that PEI1 is an embryo-specific transcription factor that plays an important role during Arabidopsis embryogenesis, functioning primarily in the apical domain of the embryo. PMID:9501112

  6. Inoculation of somatic embryos of sweet potato with an arbuscular mycorrhizal fungus improves embryo survival and plantlet formation.

    PubMed

    Bressan, W; de Carvalho, C H; Sylvia, D M

    2000-08-01

    Responses of somatic embryos of sweet potato (Ipomoea batata (L.) Poir., cv. White Star) at different developmental stages to in vitro inoculation with Glomus etunicatum (Becker and Gerdemann) (isolate INVAM FL329) were evaluated. Somatic embryos were grown in glass tubes containing sterilized vermiculite and sand. A layer of natrosol plus White's medium was used as a carrier for arbuscular mycorrhizal (AM) fungal spores. Survival of embryos inoculated with AM fungi was significantly (P < 0.05) greater than that of noninoculated embryos at the rooted-cotyledonary-torpedo and rooted-elongated-torpedo developmental stages. Mycorrhizae significantly (P < 0.05) increased plantlet formation only when inoculation occurred at the rooted-elongated-torpedo developmental stage. The growth stage at which the embryos were inserted into the glass tubes exerted a significant influence upon plantlet formation, and plantlet formation was further enhanced by inoculation with G. etunicatum. Plantlet formation was greatest at the rooted-elongated-torpedo stage. These results demonstrate that inoculation of somatic embryos with AM fungi improves embryo survival and plantlet formation, and could enhance use of somatic embryos as synthetic seeds.

  7. Accurate and noninvasive embryos screening during in vitro fertilization (IVF) assisted by Raman analysis of embryos culture medium Accurate and noninvasive embryos screening during IVF

    NASA Astrophysics Data System (ADS)

    Shen, A. G.; Peng, J.; Zhao, Q. H.; Su, L.; Wang, X. H.; Hu, J. M.; Yang, J.

    2012-04-01

    In combination with morphological evaluation tests, we employ Raman spectroscopy to select higher potential reproductive embryos during in vitro fertilization (IVF) based on chemical composition of embryos culture medium. In this study, 57 Raman spectra are acquired from both higher and lower quality embryos culture medium (ECM) from 10 patients which have been preliminarily confirmed by clinical assay. Data are fit by using a linear combination model of least squares method in which 12 basis spectra represent the chemical features of ECM. The final fitting coefficients provide insight into the chemical compositions of culture medium samples and are subsequently used as criterion to evaluate the quality of embryos. The relative fitting coefficients ratios of sodium pyruvate/albumin and phenylalanine/albumin seem act as key roles in the embryo screening, attaining 85.7% accuracy in comparison with clinical pregnancy. The good results demonstrate that Raman spectroscopy therefore is an important candidate for an accurate and noninvasive screening of higher quality embryos, which potentially decrease the time-consuming clinical trials during IVF.

  8. Treating cloned embryos, but not donor cells, with 5-aza-2'-deoxycytidine enhances the developmental competence of porcine cloned embryos.

    PubMed

    Huan, Yan Jun; Zhu, Jiang; Xie, Bing Teng; Wang, Jian Yu; Liu, Shi Chao; Zhou, Yang; Kong, Qing Ran; He, Hong Bin; Liu, Zhong Hua

    2013-10-01

    The efficiency of cloning by somatic cell nuclear transfer (SCNT) has remained low. In most cloned embryos, epigenetic reprogramming is incomplete, and usually the genome is hypermethylated. The DNA methylation inhibitor 5-aza-2'-deoxycytidine (5-aza-dC) could improve the developmental competence of cow, pig, cat and human SCNT embryos in previous studies. However, the parameters of 5-aza-dC treatment among species are different, and whether 5-aza-dC could enhance the developmental competence of porcine cloned embryos has still not been well studied. Therefore, in this study, we treated porcine fetal fibroblasts (PFF) that then were used as donor nuclei for nuclear transfer or fibroblast-derived reconstructed embryos with 5-aza-dC, and the concentration- and time-dependent effects of 5-aza-dC on porcine cloned embryos were investigated by assessing pseudo-pronucleus formation, developmental potential and pluripotent gene expression of these reconstructed embryos. Our results showed that 5-aza-dC significantly reduced the DNA methylation level in PFF (0 nM vs. 10 nM vs. 25 nM vs. 50 nM, 58.70% vs. 37.37% vs. 45.43% vs. 39.53%, P<0.05), but did not improve the blastocyst rate of cloned embryos derived from these cells. Treating cloned embryos with 25 nM 5-aza-dC for 24 h significantly enhanced the blastocyst rate compared with that of the untreated group. Furthermore, treating cloned embryos, but not donor cells, significantly promoted pseudo-pronucleus formation at 4 h post activation (51% for cloned embryos treated, 34% for donor cells treated and 36% for control, respectively, P<0.05) and enhanced the expression levels of pluripotent genes (Oct4, Nanog and Sox2) up to those of in vitro fertilized embryos during embryo development. In conclusion, treating cloned embryos, but not donor cells, with 5-aza-dC enhanced the developmental competence of porcine cloned embryos by promotion of pseudo-pronucleus formation and improvement of pluripotent gene expression.

  9. The problem of periodic patterns in embryos.

    PubMed

    Cooke, J

    1981-10-07

    A segmented body-plan has developed at least twice during metazoan evolution: in the lineage including annelids and arthropods, where the segment is the unit of body structure, and in the ancestors of vertebrates, where a primary segmentation of the middle, mesodermal cell layer of the embryo imposes a spatially periodic character upon derivatives of other layers. The mechanism controlling the development of these periodic patterns has the property that the number of the serially homologous structures formed within each species is largely independent of the linear dimension, or scale, at which pattern formation occurs in individual cases. In this they contrast with other patterns of dispersed, homologous structures occurring in animal epidermis and dermis. The performance of various classes of model for the control of number in vertebrates somite formation are compared, in the light of experimentally and naturally observable properties of this aspect of pattern.

  10. Transcriptional Memory in the Drosophila Embryo.

    PubMed

    Ferraro, Teresa; Esposito, Emilia; Mancini, Laure; Ng, Sam; Lucas, Tanguy; Coppey, Mathieu; Dostatni, Nathalie; Walczak, Aleksandra M; Levine, Michael; Lagha, Mounia

    2016-01-25

    Transmission of active transcriptional states from mother to daughter cells has the potential to foster precision in the gene expression programs underlying development. Such transcriptional memory has been specifically proposed to promote rapid reactivation of complex gene expression profiles after successive mitoses in Drosophila development [1]. By monitoring transcription in living Drosophila embryos, we provide the first evidence for transcriptional memory in animal development. We specifically monitored the activities of stochastically expressed transgenes in order to distinguish active and inactive mother cells and the behaviors of their daughter nuclei after mitosis. Quantitative analyses reveal that there is a 4-fold higher probability for rapid reactivation after mitosis when the mother experienced transcription. Moreover, memory nuclei activate transcription twice as fast as neighboring inactive mothers, thus leading to augmented levels of gene expression. We propose that transcriptional memory is a mechanism of precision, which helps coordinate gene activity during embryogenesis.

  11. Mammalian diversity: gametes, embryos and reproduction.

    PubMed

    Behringer, Richard R; Eakin, Guy S; Renfree, Marilyn B

    2006-01-01

    The class Mammalia is composed of approximately 4800 extant species. These mammalian species are divided into three subclasses that include the monotremes, marsupials and eutherians. Monotremes are remarkable because these mammals are born from eggs laid outside of the mother's body. Marsupial mammals have relatively short gestation periods and give birth to highly altricial young that continue a significant amount of 'fetal' development after birth, supported by a highly sophisticated lactation. Less than 10% of mammalian species are monotremes or marsupials, so the great majority of mammals are grouped into the subclass Eutheria, including mouse and human. Mammals exhibit great variety in morphology, physiology and reproduction. In the present article, we highlight some of this remarkable diversity relative to the mouse, one of the most widely used mammalian model organisms, and human. This diversity creates challenges and opportunities for gamete and embryo collection, culture and transfer technologies.

  12. Microwave effects on isolated chick embryo hearts

    SciTech Connect

    Caddemi, A.; Tamburello, C.C.; Zanforlin, L.; Torregrossa, M.V.

    1986-01-01

    This study was designed to examine the effects of microwaves on the electric activity of hearts as a means of elucidating interactive mechanisms of nonionizing radiation with cardiac tissue. Experiments were performed on isolated hearts of 9-12-day-old chick embryos placed in small petri dishes. Oxygenated isotonic Ringer's solution at 37 degrees C permitted heart survival. Samples were irradiated at 2.45 GHz with a power density of 3 mW/cm2. The heart signal was detected with a glass micropipet inserted into the sinoatrial node and examined by means of a Berg-Fourier analyzer. Pulsed microwaves caused the locking of the heartbeat to the modulation frequency, whereas continuous wave irradiation might have induced slight bradycardia. Pulsed fields induced stimulation or regularization of the heartbeat in arrhythmia, fibrillation, or arrest of the heart.

  13. Phosphoproteome Analysis of Drosophila melanogaster Embryos

    PubMed Central

    Zhai, Bo; Villén, Judit; Beausoleil, Sean A.; Mintseris, Julian; Gygi, Steven P.

    2011-01-01

    Protein phosphorylation is a key regulatory event in most cellular processes and development. Mass spectrometry-based proteomics provides a framework for the large-scale identification and characterization of phosphorylation sites. Here, we used a well-established phosphopeptide enrichment and identification strategy including the combination of strong cation exchange chromatography, immobilized metal affinity chromatography, and high-accuracy mass spectrometry instrumentation to study phosphorylation in developing Drosophila embryos. In total, 13 720 different phosphorylation sites were discovered from 2702 proteins with an estimated false-discovery rate (FDR) of 0.63% at the peptide level. Because of the large size of the data set, both novel and known phosphorylation motifs were extracted using the Motif-X algorithm, including those representative of potential ordered phosphorylation events. PMID:18327897

  14. [Cultural diversity in gamete and embryos donation].

    PubMed

    Epelboin, S

    2014-09-01

    Through gamete and embryo donation have successively emerged new ways of designing individuals who, in turn, have generated mutations in the concept of parenthood. A debate is open to the society, which often raises ideological cleavages. Indeed, donation practices mobilize the conflicting interests of donor of gametes, the recipient couple, child, whose origins are complex, although his filiation is legally clear. Its place in the family genealogy can be examined in relation to other societies, which admit plural concepts called "classificatory" kinship. They set up role partition between parents and educators. Setting anthropological perspective provides a broadening of the reflection to answer questions from the donation practices, including genealogical questions of revelation to the child of his conception, his incorporation in family and social group and the importance of compensation of donation.

  15. Directional freezing of spermatozoa and embryos.

    PubMed

    Arav, Amir; Saragusty, Joseph

    2013-01-01

    Directional freezing is based on a simple thermodynamic principle whereby the sample is moved through a predetermined temperature gradient at a velocity that determines the cooling rate. Directional freezing permits a precise and uniform cooling rate in small- and large-volume samples. It avoids supercooling and reduces mechanical damage caused by crystallisation. Directional solidification was used to date for slow and rapid freezing, as well as for vitrification of oocytes and embryos by means of the minimum drop size technique: small drops are placed on a microscope slide that is moved at high velocity from the hot base to the cold base. Sperm samples from a wide range of domestic and wild animals were successfully cryopreserved using the directional freezing method. The bovine sexed semen industry may benefit from the increased survival of spermatozoa after directional freezing.

  16. The transcriptome of the sea urchin embryo.

    PubMed

    Samanta, Manoj P; Tongprasit, Waraporn; Istrail, Sorin; Cameron, R Andrew; Tu, Qiang; Davidson, Eric H; Stolc, Viktor

    2006-11-10

    The sea urchin Strongylocentrotus purpuratus is a model organism for study of the genomic control circuitry underlying embryonic development. We examined the complete repertoire of genes expressed in the S. purpuratus embryo, up to late gastrula stage, by means of high-resolution custom tiling arrays covering the whole genome. We detected complete spliced structures even for genes known to be expressed at low levels in only a few cells. At least 11,000 to 12,000 genes are used in embryogenesis. These include most of the genes encoding transcription factors and signaling proteins, as well as some classes of general cytoskeletal and metabolic proteins, but only a minor fraction of genes encoding immune functions and sensory receptors. Thousands of small asymmetric transcripts of unknown function were also detected in intergenic regions throughout the genome. The tiling array data were used to correct and authenticate several thousand gene models during the genome annotation process.

  17. Selection for rapid embryo development correlates with embryo exposure to maternal androgens among passerine birds

    USGS Publications Warehouse

    Schwabl, H.; Palacios, M.G.; Martin, T.E.

    2007-01-01

    Greater offspring predation favors evolution of faster development among species. We hypothesized that greater offspring predation exerts selection on mothers to increase levels of anabolic androgens in egg yolks to achieve faster development. Here, we tested whether (1) concentrations of yolk androgens in passerine species were associated with offspring predation and (2) embryo and nestling development rates were associated with yolk androgen concentrations. We examined three androgens that increase in potency along the synthesis pathway: androstenedione (A4) to testosterone (T) to 5??- dihydrotestosterone (5??-DHT). Concentrations of none of these steroids were related to clutch size; only A4 was allometrically related to egg volume. Species that experience greater predation showed higher yolk concentrations of T and 5??-DHT. Higher concentrations of T and particularly 5??-DHT were strongly correlated with faster development during the embryo period and less so during the nestling period. Development rates were most strongly correlated with 5??-DHT, suggesting that potency increases along the androgen synthesis pathway and that effects are mediated by the androgen receptor pathway. These results are consistent with the hypothesis that selection for faster development by time-dependent offspring mortality may be achieved epigenetically by varying embryo exposure to maternal anabolic steroids. ?? 2007 by The University of Chicago. All rights reserved.

  18. From embryo sac to oil and protein bodies: embryo development in the model legume Medicago truncatula.

    PubMed

    Wang, Xin-Ding; Song, Youhong; Sheahan, Michael B; Garg, Manohar L; Rose, Ray J

    2012-01-01

    • The cell and developmental biology of zygotic embryogenesis in the model legume Medicago truncatula has received little attention. We studied M. truncatula embryogenesis from embryo sac until cotyledon maturation, including oil and protein body biogenesis. • We characterized embryo development using light and electron microscopy, measurement of protein and lipid fatty acid accumulation and by profiling the expression of key seed storage genes. • Embryo sac development in M. truncatula is of the Polygonum type. A distinctive multicellular hypophysis and suspensor develops before the globular stage and by the early cotyledon stage, the procambium connects the developing apical meristems. In the storage parenchyma of cotyledons, ovoid oil bodies surround protein bodies and the plasma membrane. Four major lipid fatty acids accumulate as cotyledons develop, paralleling the expression of OLEOSIN and the storage protein genes, VICILIN and LEGUMIN. • Zygotic embryogenesis in M. truncatula features the development of a distinctive multicellular hypophysis and an endopolyploid suspensor with basal transfer cell. A clear procambial connection between the apical meristems is evident and there is a characteristic arrangement of oil bodies in the cotyledons and radicle. Our data help link embryogenesis to the genetic regulation of oil and protein body biogenesis in legume seed.

  19. Ion/water channels for embryo implantation barrier.

    PubMed

    Liu, Xin-Mei; Zhang, Dan; Wang, Ting-Ting; Sheng, Jian-Zhong; Huang, He-Feng

    2014-05-01

    Successful implantation involves three distinct processes, namely the embryo apposition, attachment, and penetration through the luminal epithelium of the endometrium to establish a vascular link to the mother. After penetration, stromal cells underlying the epithelium differentiate and surround the embryo to form the embryo implantation barrier, which blocks the passage of harmful substances to the embryo. Many ion/water channel proteins were found to be involved in the process of embryo implantation. First, ion/water channel proteins play their classical role in establishing a resting membrane potential, shaping action potentials and other electrical signals by gating the flow of ions across the cell membrane. Second, most of ion/water channel proteins are regulated by steroid hormone (estrogen or progesterone), which may have important implications to the embryo implantation. Last but not least, these proteins do not limit themselves as pure channels but also function as an initiator of a series of consequences once activated by their ligand/stimulator. Herein, we discuss these new insights in recent years about the contribution of ion/water channels to the embryo implantation barrier construction during early pregnancy.

  20. Shaping the norms that regulate international commerce of embryos.

    PubMed

    Gard, Julie A; Stringfellow, David A

    2014-01-01

    As various embryo technologies in livestock were developed and evolved to a state of usefulness over the past 40 years, scientists with a specific interest in infectious diseases sought to determine the epidemiologic consequences of movement, especially international movement, of increasing numbers of embryos. Many of the foundational studies in this area were reported in Theriogenology, beginning in the 1970s and especially throughout the 1980s and 1990s. Unquestionably, Theriogenology has been a widely used venue for dissemination of basic information on this subject, which ultimately led to the development of the now universally accepted techniques for certification of embryo health. Today it is well-recognized that movement in commerce of embryos, especially in vivo-derived embryos, is a very low-risk method for exchange of animal germ plasm. This paper chronicles the evolution of strategies for health certification of embryos. An overview is provided of the calculated efforts of practitioners, scientists, and regulators to organize, forge necessary partnerships, stimulate needed research, provide purposeful analysis of the results, and, through these processes, guarantee the universal acceptance of efficient protocols for certifying the health of embryos intended for movement in international commerce.

  1. Should we be promoting embryo transfer at blastocyst stage?

    PubMed

    Maheshwari, Abha; Hamilton, Mark; Bhattacharya, Siladitya

    2016-02-01

    Improved laboratory standards and better culture media have made extended culture to blastocyst stage a reality to identify embryos with maximum implantation potential. The strategy of extended culture has become more popular across the world at a time when regulatory bodies have emphasized the need to increase the uptake of elective single embryo transfer, minimize complications associated with multiple births and aim for a healthy singleton live-birth as the preferred outcome in IVF. New data on perinatal outcomes suggest that pregnancies after embryo transfer at blastocyst stage are associated with a higher risk of preterm delivery, large for gestational age babies, monozygotic twins and altered sex ratio compared with those following embryo transfers at cleavage stage. In addition, concerns have been raised of increased congenital anomalies and epigenetic modifications with embryo transfer at blastocyst stage. Twenty-four years on from the first embryo transfer at blastocyst stage, we examine the reasons for extended embryo culture, evaluate the risks and benefits of this strategy and suggest the need to reconsider this policy in the interests of fetal safety.

  2. Zika Virus Induced Mortality and Microcephaly in Chicken Embryos.

    PubMed

    Goodfellow, Forrest T; Tesla, Blanka; Simchick, Gregory; Zhao, Qun; Hodge, Thomas; Brindley, Melinda A; Stice, Steven L

    2016-11-15

    The explosive spread of the Zika virus (ZIKV) through South and Central America has been linked to an increase in congenital birth defects, specifically microcephaly. Representative rodent models for investigating infections include direct central nervous system (CNS) injections late in pregnancy and transplacental transmission in immunodeficient mice. Microcephaly in humans may be the result of infection occurring early in pregnancy, therefore recapitulating that the human course of ZIKV infection should include normal embryo exposed to ZIKV during the first trimester. In ovo development of the chicken embryo closely mirrors human fetal neurodevelopment and, as a comparative model, could provide key insights into both temporal and pathophysiological effects of ZIKV. Chick embryos were directly infected early and throughout incubation with ZIKV isolated from a Mexican mosquito in January 2016. High doses of virus caused embryonic lethality. In a subset of lower dosed embryos, replicating ZIKV was present in various organs, including the CNS, throughout development. Surviving ZIKV-infected embryos presented a microcephaly-like phenotype. Chick embryos were longitudinally monitored by magnetic resonance imaging that documented CNS structural malformations, including enlarged ventricles (30% increase) and stunted cortical growth (decreased telencephalon by 18%, brain stem by 32%, and total brain volume by 18%), on both embryonic day 15 (E15) and E20 of development. ZIKV-induced microcephaly was observed with inoculations of as few as 2-20 viral particles. The chick embryo model presented ZIKV embryonic lethal effects and progressive CNS damage similar to microcephaly.

  3. Is embryo research and preimplantation genetic diagnosis ethical?

    PubMed

    Beyleveld, D

    2000-09-11

    The legal position in the UK on embryo research and preimplantation genetic diagnosis (PGD) is outlined and contrasted with the position in other EU countries. The "gradualist" position of the UK on the moral status of the embryo is defended on the basis of an argument that precaution must be applied in proportion to the degree to which the embryo has developed to display components of agency, on the assumption that mortality is categorically binding and requires agents to be granted rights and that it cannot be known with certainty that the embryo is not an agent. The extent to which this argument, when combined with vicarious protections that the embryo should receive in order to protect the rights of other agents, limits embryo research and PGD, is discussed. It is concluded that the complexities that attend deliberation about the moral problems attending embryo research and PGD are such that the proper response to these problems is via the procedures of political democracy to achieve accountable answers rather than "correct" answers. This allows for a variety of judgements.

  4. Nuclear reprogramming of cloned embryos produced in vitro.

    PubMed

    Han, Y M; Kang, Y K; Koo, D B; Lee, K K

    2003-01-01

    Despite the fact that cloned animals derived from somatic cells have been successfully generated in a variety of mammalian species, there are still many unsolved problems with current cloning technology. Somatic cell nuclear transfer has shown several developmental aberrancies, including a high rate of abortion during early gestation and increased perinatal death. One cause of these developmental failures of cloned embryos may reside in the epigenetic reprogramming of somatic donor genome. In mammals, DNA methylation is an essential process in the regulation of transcription during embryonic development and is generally associated with gene silencing. A genome-wide demethylation may be a prerequisite for the formation of pluripotent stem cells that are important for later development. We analyzed methylation patterns in cloned bovine embryos to monitor the epigenetic reprogramming process of donor genomic DNA. Aberrant methylation profiles of cloned bovine embryos were observed in various genomic regions, except in single-copy gene sequences. The overall genomic methylation status of cloned embryos was quite different from that of normal embryos produced in vitro or in vivo. These results suggest that the developmental failures of cloned embryos may be due to incomplete epigenetic reprogramming of donor genomic DNA. We expect that advances in understanding the molecular events for reprogramming of donor genome will contribute to clarify the developmental defects of cloned embryos.

  5. Human embryos cultured in vitro to 14 days

    PubMed Central

    2017-01-01

    We know a great deal about the development of the mammalian embryo until the time that the blastocyst implants into the uterus. With model organisms such as the mouse, we have also developed a considerable understanding of development immediately around gastrulation as embryos can be recovered at this stage for short-term in vitro culture. However, the intervening period of development remained a ‘black box’ because it takes place as the blastocyst is implanting into the uterus. Over the past 6 years, techniques pioneered and developed in Magdalena Zernicka-Goetz's laboratory for the in vitro culture of embryos through these implantation stages have opened up this box, affording the first glimpse of embryonic development through these previously hidden stages. Remarkably, the techniques developed with mouse embryos are equally applicable to human embryos, ushering the very first opportunities for studying our own development throughout this time. Here, I outline how the culture methods were developed, paving the way to culture of the human embryo to the point of gastrulation, an accomplishment recognized as the People's Choice for the Scientific Breakthrough of 2016 in Science magazine. I also discuss the new ethical challenges raised by the possibility of extending the time limits for human embryo culture. PMID:28123056

  6. Diffusion of small molecules into medaka embryos improved by electroporation

    PubMed Central

    2013-01-01

    Background Diffusion of small molecules into fish embryos is essential for many experimental procedures in developmental biology and toxicology. Since we observed a weak uptake of lithium into medaka eggs we started a detailed analysis of its diffusion properties using small fluorescent molecules. Results Contrary to our expectations, not the rigid outer chorion but instead membrane systems surrounding the embryo/yolk turned out to be the limiting factor for diffusion into medaka eggs. The consequence is a bi-phasic uptake of small molecules first reaching the pervitelline space with a diffusion half-time in the range of a few minutes. This is followed by a slow second phase (half-time in the range of several hours) during which accumulation in the embryo/yolk takes place. Treatment with detergents improved the uptake, but strongly affected the internal distribution of the molecules. Testing electroporation we could establish conditions to overcome the diffusion barrier. Applying this method to lithium chloride we observed anterior truncations in medaka embryos in agreement with its proposed activation of Wnt signalling. Conclusions The diffusion of small molecules into medaka embryos is slow, caused by membrane systems underneath the chorion. These results have important implications for pharmacologic/toxicologic techniques like the fish embryo test, which therefore require extended incubation times in order to reach sufficient concentrations in the embryos. PMID:23815821

  7. Danio rerio embryos on Prozac - Effects on the detoxification mechanism and embryo development.

    PubMed

    Cunha, V; Rodrigues, P; Santos, M M; Moradas-Ferreira, P; Ferreira, M

    2016-09-01

    In the past decade the presence of psychopharmaceuticals, including fluoxetine (FLU), in the aquatic environment has been associated with the increasing trend in human consumption of these substances. Aquatic organisms are usually exposed to chronic low doses and, therefore, risk assessments should evaluate the effects of these compounds in non-target organisms. Teleost fish possess an array of active defence mechanisms to cope with the deleterious effects of xenobiotics. These include ABC transporters, phase I and II of cellular detoxification and oxidative stress enzymes. Hence, the present study aimed at characterising the effect of FLU on embryo development of the model teleost zebrafish (Danio rerio) concomitantly with changes in the detoxification mechanisms during early developmental phases. Embryos were exposed to different concentrations of FLU (0.0015, 0.05, 0.1, 0.5 and 0.8μM) for 80hours post fertilization. Development was screened and the impact in the transcription of key genes, i.e., abcb4, abcc1, abcc2, abcg2, cyp1a, cyp3a65, gst, sod, cat, ahr, pxr, pparα, pparβ, pparγ, rxraa, rxrab, rxrbb, rxrga, rxrgb, raraa, rarab, rarga evaluated. In addition, accumulation assays were performed to measure the activity of ABC proteins and antioxidant enzymes (CAT and Cu/ZnSOD) after exposure to FLU. Embryo development was disrupted at the lowest FLU concentration tested (0.0015μM), which is in the range of concentrations found in WWTP effluents. Embryos exposed to higher concentrations of FLU decreased Cu/Zn SOD, and increased CAT (0.0015 and 0.5μM) enzymatic activity. Exposure to higher concentrations of FLU decreased the expression of most genes belonging to the detoxification system and upregulated cat at 0.0015μM of FLU. Most of the tested concentrations downregulated pparα, pparβ, pparγ, and raraa, rxraa, rxrab, rxrbb rxrgb and ahr gene expression while pxr was significantly up regulated at all tested concentrations. In conclusion, this study

  8. The impact of the hailstone embryos on simulated surface precipitation

    NASA Astrophysics Data System (ADS)

    Kovačević, Nemanja; Ćurić, Mladjen

    2013-10-01

    Hailstorms cause significant damage to agriculture and property in many areas of the world. Therefore, it is useful to describe the size spectrum of hail and the mechanisms of formation in more detail. One important point in the formation of hail is the role of hailstone embryos, and an understanding of their mechanism would significantly improve our understanding of the evolution of hail, as well as the predicted amount of accumulated hail on the ground. We used a cloud-resolving mesoscale model to investigate the influence of the hailstone embryos on the measured ground precipitation. In this model, both types of the hailstone embryos (graupel and frozen raindrops) are incorporated. Therefore, the model predicts the mass and number concentration of the six microphysical elements - raindrops, ice crystals, snow, graupel, frozen raindrops and hail. The cloud droplet number concentration was prescribed. Thus, the primary goal of this sensitivity study was to examine the influence of hailstone embryos on the measured ground precipitation and the duration of precipitation. Thus, we performed a numerical comparison of the two microphysical schemes, one with hailstone embryos and the other without them. The sensitivity study indicated that the microphysical scenario with hailstone embryos leads to a greater increase in accumulated hail compared with the scheme without hailstone embryos. The time of hail occurrence on the ground occurs during the early stages of cloud life in the experiment without hailstone embryos. In the second case, the hail occurrence on the ground was delayed for the later stages of cloud life, which is much more realistic and in agreement with the measurements. The use of a model with hailstone embryos leads to a better description of the evolution of hail and a more accurate prediction of the accumulated hail on the ground.

  9. The avian embryo responding to microgravity of space flight

    NASA Technical Reports Server (NTRS)

    Hullinger, Ronald L.

    1993-01-01

    Of all the many potential and real microenvironmental influences, only gravity would appear to have remained relatively constant and ubiquitous for developing organisms. Histo- and organogenesis as well as differential growth of the embryo and fetus may have evolved with a constant environmental factor of gravity. Chick embryos of 2-day and 9-day stages of incubation were flown in an incubator on the Space Shuttle during a 9-day mission. Significant differences in embryo response to this microgravity environment were observed. This paper offers an analysis and suggests mechanisms which may contribute to these results.

  10. The scourge: moral implications of natural embryo loss.

    PubMed

    Ord, Toby

    2008-07-01

    It is often claimed that from the moment of conception embryos have the same moral status as adult humans. This claim plays a central role in many arguments against abortion, in vitro fertilization, and stem cell research. In what follows, I show that this claim leads directly to an unexpected and unwelcome conclusion: that natural embryo loss is one of the greatest problems of our time and that we must do almost everything in our power to prevent it. I examine the responses available to those who hold that embryos have full moral status and conclude that they cannot avoid the force of this argument without giving up this key claim.

  11. Genetic modification of preimplantation embryos: toward adequate human research policies.

    PubMed

    Dresser, Rebecca

    2004-01-01

    Citing advances in transgenic animal research and setbacks in human trials of somatic cell genetic interventions, some scientists and others want to begin planning for research involving the genetic modification of human embryos. Because this form of genetic modification could affect later-born children and their offspring, the protection of human subjects should be a priority in decisions about whether to proceed with such research. Yet because of gaps in existing federal policies, embryo modification proposals might not receive adequate scientific and ethical scrutiny. This article describes current policy shortcomings and recommends policy actions designed to ensure that the investigational genetic modification of embryos meets accepted standards for research on human subjects.

  12. Automatic zebrafish heartbeat detection and analysis for zebrafish embryos.

    PubMed

    Pylatiuk, Christian; Sanchez, Daniela; Mikut, Ralf; Alshut, Rüdiger; Reischl, Markus; Hirth, Sofia; Rottbauer, Wolfgang; Just, Steffen

    2014-08-01

    A fully automatic detection and analysis method of heartbeats in videos of nonfixed and nonanesthetized zebrafish embryos is presented. This method reduces the manual workload and time needed for preparation and imaging of the zebrafish embryos, as well as for evaluating heartbeat parameters such as frequency, beat-to-beat intervals, and arrhythmicity. The method is validated by a comparison of the results from automatic and manual detection of the heart rates of wild-type zebrafish embryos 36-120 h postfertilization and of embryonic hearts with bradycardia and pauses in the cardiac contraction.

  13. Competing Views of Embryos for the Twenty-First Century: Textbooks and Society

    ERIC Educational Resources Information Center

    Maienschein, Jane; Wellner, Karen

    2013-01-01

    It might seem that an embryo is an embryo, and that there would be a fact of the matter. That seems especially true with respect to the way embryos are presented in textbooks, including high school biology textbooks. This paper looks at three co-existing, competing, and often conflicting views of embryos. Then with a close study of twentieth…

  14. 9 CFR 98.18 - Shipment of embryos to the United States.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Shipment of embryos to the United... IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.18 Shipment of embryos to the United States. (a)...

  15. 9 CFR 98.18 - Shipment of embryos to the United States.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Shipment of embryos to the United... IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.18 Shipment of embryos to the United States. (a)...

  16. 9 CFR 98.18 - Shipment of embryos to the United States.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Shipment of embryos to the United... IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.18 Shipment of embryos to the United States. (a)...

  17. 9 CFR 98.18 - Shipment of embryos to the United States.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Shipment of embryos to the United... IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.18 Shipment of embryos to the United States. (a)...

  18. 9 CFR 98.18 - Shipment of embryos to the United States.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Shipment of embryos to the United... IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.18 Shipment of embryos to the United States. (a)...

  19. Polyethylene glycol-induced fusion of two-cell mouse embryo blastomeres

    SciTech Connect

    Spindle, A.

    1981-01-01

    Polyethylene glycol (PEG) was found to be an effective fusion-inducing agent for early mouse embryo blastomeres. A brief exposure of zona-intact 2-cell embryos to 40% PEG induced fusion of blastomeres in > 80% of embryos, and the treatment did not interfere with subsequent development of embryos to the blastocyst stage.

  20. The effect of sevoflurane on developing A/J strain mouse embryos using a whole-embryo culture system--the incidence of cleft lip in culture embryos.

    PubMed

    Yamada, Morimasa; Yamamoto, Naoki; Ohgami, Saori; Kanazawa, Mayuko; Harada, Jun; Ohno, Norikazu; Natsume, Nagato

    2014-03-01

    A/J strain mice have a high spontaneous incidence of cleft lip (ICL) and/or palate. The primary palate-related effects of sevoflurane on developing A/J strain mouse embryos (embryos) were studied using a whole-embryo culture (WEC) system. This system could separate the direct effects of sevoflurane from those that are maternally mediated. A total of 205 10.5-d embryos were cultured for 24 h in either a control group (control gas: 95% O2 and 5% CO2) or sevoflurane-administered groups (1/4, 1/2, and 1 minimum alveolar concentration (MAC) with control gas) for 8 h. After 16 h, 11.5-d culture embryos were examined in terms of crown-rump length, number of somites, and protein content. Crown-rump length in the 1 MAC was significantly shorter than in the control group (p < 0.05). Protein content in the 1/2 MAC (p < 0.05) and 1 MAC (p < 0.001) was significantly lower than in the control group. The ICL showed no significant differences between each group. (The ICL rose with an increase in the sevoflurane concentration, but this was not significant). The positive findings in this study indicate that a WEC system is useful for studying the mechanisms of ICL (teratogenicity) associated with sevoflurane.

  1. Haeckel's Embryos and Evolution: Setting the Record Straight.

    ERIC Educational Resources Information Center

    Wells, Jonathan

    1999-01-01

    Argues that Ernst Haeckel's drawings of vertebrate embryos (1891), which have been widely used in biology textbooks to illustrate his "Biogenetic Law", are factually flawed. Discusses the problems with Haeckel's drawings and his theory. Contains 14 references. (WRM)

  2. Embryo research: the ethical geography of the debate.

    PubMed

    Khushf, G

    1997-10-01

    Three basic political positions on embryo research will be identified as libertarian, conservative, and social-democratic. The Human Embryo Research Panel will be regarded as an expression of the social-democratic position. A taxonomy of the ethical issues addressed by the Panel will then be developed at the juncture of political and ethical modes of reflection. Among the arguments considered will be those for the separability of the abortion and embryo research debates; arguments against the possibility of the preembryo being a person, especially arguments associated with totipotency and the significance of the primitive streak; and the various reasons for regulating embryo research, including those associated with respect for the preembryo, the protection of traditional views of human procreation, and the prevention of commercialization.

  3. Immature seeds and embryos of Medicago truncatula cultured in vitro.

    PubMed

    Ochatt, Sergio J

    2011-01-01

    Legumes are an important source of proteins and lipids for food and feed. In addition, they are -environmentally friendly because of their capacity to fix nitrogen through a symbiosis with Rhizobium that permits them to produce abundant proteins even in the absence of nitrogen fertilization. Seed development in plants follows three chronological steps (1) seed coat differentiation, embryo morphogenesis and endosperm development; (2) embryo maturation with storage accumulation and (3) dehydration and the acquisition of desiccation tolerance. Finally, germination occurs when the environmental conditions become favourable. Working with the model legume Medicago truncatula, an in vitro protocol was developed for the culture of immature embryos that permits their development in a way comparable to that observed in plants.In this chapter, the usefulness of this system for investigating embryo development in legumes is outlined.

  4. Environmental influences on the production of pre-implantation embryos.

    PubMed

    Diercks, Ann-Kathrin; Schwab, Anna; Rittgen, Werner; Kruspel, Andreas; Heuss, Edgar; Schenkel, Johannes

    2010-06-01

    Generation and cryopreservation of transgenic mice depend on reliable and continuous production of pre-implantation embryos. To suppress circannual and circadian rhythms driving the physiological and sexual behaviour of free living animals, laboratory animals are housed under standardized conditions. It remains to be elucidated if the artificial climate can cover all environmental effects. Here, we report that the humidity in an animal facility affects the embryo yield. The weather at the location of the facility, especially the temperature, influences the climate within an animal facility; weather peaks are obviously covered in part only, even if the facility is equipped with a powerful air-conditioning supply. Subsequently, external weather changes interact with the environment within the facility, influencing the production of embryos. Furthermore, noise and/or vibrations as generated by construction works, negatively affect the embryo yield.

  5. Targeted mutagenesis in sea urchin embryos using TALENs.

    PubMed

    Hosoi, Sayaka; Sakuma, Tetsushi; Sakamoto, Naoaki; Yamamoto, Takashi

    2014-01-01

    Genome editing with engineered nucleases such as zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) has been reported in various animals. We previously described ZFN-mediated targeted mutagenesis and insertion of reporter genes in sea urchin embryos. In this study, we demonstrate that TALENs can induce mutagenesis at specific genomic loci of sea urchin embryos. Injection of TALEN mRNAs targeting the HpEts transcription factor into fertilized eggs resulted in the impairment of skeletogenesis. Sequence analyses of the mutations showed that deletions and/or insertions occurred at the HpEts target site in the TALEN mRNAs-injected embryos. The results suggest that targeted gene disruption using TALENs is feasible in sea urchin embryos.

  6. The moral status of the embryo post-Dolly.

    PubMed

    Stanton, Catherine; Harris, John

    2005-04-01

    Cameron and Williamson have provided a provocative and timely review of the ethical questions prompted by the birth of Dolly. The question Cameron and Williamson seek to address is "In the world of Dolly, when does a human embryo acquire respect?". Their initial discussion sets the scene by providing a valuable overview of attitudes towards the embryo, summarising various religious, scientific, and philosophical viewpoints. They then ask, "What has Dolly changed?" and identify five changes, the first being that fertilisation is no longer required to create an embryo. Following this analysis they then ask when an embryo created other than by fertilisation begins to acquire respect. This paper explores the ethical and legal issues highlighted by Cameron and Williamson's paper.

  7. Cell adhesion in zebrafish embryos is modulated by March 8.

    PubMed

    Kim, Mi Ha; Rebbert, Martha L; Ro, Hyunju; Won, Minho; Dawid, Igor B

    2014-01-01

    March 8 is a member of a family of transmembrane E3 ubiquitin ligases that have been studied mostly for their role in the immune system. We find that March 8 is expressed in the zebrafish egg and early embryo, suggesting a role in development. Both knock-down and overexpression of March 8 leads to abnormal development. The phenotype of zebrafish embryos and Xenopus animal explants overexpressing March 8 implicates impairment of cell adhesion as a cause of the effect. In zebrafish embryos and in cultured cells, overexpression of March 8 leads to a reduction in the surface levels of E-cadherin, a major cell-cell adhesion molecule. Experiments in cell culture further show that E-cadherin can be ubiquitinated by March 8. On the basis of these observations we suggest that March 8 functions in the embryo to modulate the strength of cell adhesion by regulating the localization of E-cadherin.

  8. Algorithms for automatic segmentation of bovine embryos produced in vitro

    NASA Astrophysics Data System (ADS)

    Melo, D. H.; Nascimento, M. Z.; Oliveira, D. L.; Neves, L. A.; Annes, K.

    2014-03-01

    In vitro production has been employed in bovine embryos and quantification of lipids is fundamental to understand the metabolism of these embryos. This paper presents a unsupervised segmentation method for histological images of bovine embryos. In this method, the anisotropic filter was used in the differents RGB components. After pre-processing step, the thresholding technique based on maximum entropy was applied to separate lipid droplets in the histological slides in different stages: early cleavage, morula and blastocyst. In the postprocessing step, false positives are removed using the connected components technique that identify regions with excess of dye near pellucid zone. The proposed segmentation method was applied in 30 histological images of bovine embryos. Experiments were performed with the images and statistical measures of sensitivity, specificity and accuracy were calculated based on reference images (gold standard). The value of accuracy of the proposed method was 96% with standard deviation of 3%.

  9. SSR markers for Quercus suber tree identification and embryo analysis.

    PubMed

    Gómez, A; Pintos, B; Aguiriano, E; Manzanera, J A; Bueno, M A

    2001-01-01

    Three Quercus simple sequence repeat (SSR) markers were amplified by polymerase chain reaction (PCR) from nuclear DNA extracts of trees and in vitro-induced haploid embryos from anther cultures of Quercus suber L. These markers were sufficiently polymorphic to identify 10 of 12 trees located in two Spanish natural areas. The same loci have been analyzed in anther-derived haploid embryos showing the parental tree allele segregation. All the alleles were present in the haploid progeny. The presence of diverse alleles in embryos derived from the same anther demonstrated that they were induced on multiple microspores or pollen grains and they were not clonally propagated. Also, diploid cultures and mixtures of haploid-diploid tissues were obtained. The origin of such cultures, either somatic or gametic, was elucidated by SSR markers. All the embryos showed only one allele, corroborating a haploid origin. Allelic composition of the haploid progeny permitted parental identification among all analyzed trees.

  10. vEmbryo In Silico Models: Predicting Vascular Developmental Toxicity

    EPA Science Inventory

    The cardiovascular system is the first to function in the vertebrate embryo, reflecting the critical need for nutrient delivery and waste removal during organogenesis. Blood vessel development occurs by complex interacting signaling networks, including extra-cellular matrix remod...

  11. [Contribution of embryo vitrification procedure to ART efficiency].

    PubMed

    Sifer, C

    2014-10-01

    This work aims to show, from data available in the literature and our own experience, how embryos' vitrification change and/or improve the management of infertile couples. In all, 652 cycles of frozen-thawed embryo transfers (FET) following vitrification were prospectively included and compared with 1126 FETs from slow freezing (SF) method. Primary end points were the (i) survival rate (SR) (% of embryos with>50% post-thaw intact blastomeres) and (ii) intact survival rate (ISR) (% of embryos with 100% post-thaw intact blastomeres). Secondary end point was the clinical pregnancy rate (CPR) defined as the presence of an intra uterine gestational sac with positive foetal heart beat. In all, 1097 and 2408 embryos have been thawed following vitrification and SF, respectively. We observed a highly significant increase of SR and ISR respectively when thawing concerned vitrified embryos rather than those from SF method (97.0% vs. 72.7%, P<10(-4); 91.5% vs. 49.8%, P<10(-4)). Furthermore, CPR were of 26.5% (73/652) and of 18.1% (204/1126) following FETs performed after vitrification or SF and thawing (P=0.0002), respectively. At the blastocyst stage, ISR was significantly improved following vitrification compared to SF (94.5% vs. 21.4%, P<10(-4)). In the study period, vitrification (i) reduced the mean number of fresh transferred embryos (1.5 vs. 1.6; P=0.08) and (ii) increased the rate of FETs at the blastocyst stage when compared with the control period (18.1% vs 2.5%., P<10(-4)). Embryo vitrification preserves all embryos from an ART cycle because of its excellent results regarding ISR at all stages of embryo development. This procedure allows a significant increase of pregnancy rates after thawing. In addition, there is a trend for increasing ART cycles performed using extended culture embryo and vitrification. The expected improvement of the cumulative birth rate at the blastocyst stage following vitrification remains to be demonstrated in a prospective randomized study.

  12. Defective Chromatin Structure in Somatic Cell Cloned Mouse Embryos*

    PubMed Central

    Zhang, Miao; Wang, Fengchao; Kou, Zhaohui; Zhang, Yu; Gao, Shaorong

    2009-01-01

    Epigenetic reprogramming plays a central role in the development of cloned embryos generated by somatic cell nuclear transfer, and it is believed that aberrant reprogramming leads to the abnormal development of most cloned embryos. Recent studies show that trimethylation of H3K27 (H3K27me3) contributes to the maintenance of embryonic stem cell pluripotency because the differentiation genes are always occupied by nucleosomes trimethylated at H3K27, which represses gene expression. Here, we provide evidence that differential H3K27me3 modification exists between normal fertilization-produced blastocysts and somatic cell nuclear transfer cloned blastocysts; H3K27me3 was specifically found in cells of the inner cell mass (ICM) of normal blastocysts, whereas there was no modification of H3K27me3 in the ICM of cloned blastocysts. Subsequently, we demonstrated that the differentiation-related genes, which are marked by H3K27me3 in embryonic stem cells, were expressed at significantly higher levels in cloned embryos than in normal embryos. The polycomb repressive complex 2 (PRC2) component genes (Eed, Ezh2, and Suz12), which are responsible for the generation of H3K27me3, were expressed at lower levels in the cloned embryos. Our results suggest that reduced expression of PRC2 component genes in cloned embryos results in defective modification of H3K27me3 to the differentiation-related genes in pluripotent ICM cells. This results in premature expression of developmental genes and death of somatic cloned embryos shortly after implantation. Taken together, these studies suggest that H3K27me3 might be an important epigenetic marker with which to evaluate the developmental potential of cloned embryos. PMID:19602512

  13. Rice embryos can express heat-shock genes under anoxia.

    PubMed

    Mocquot, B; Ricard, B; Pradet, A

    1987-01-01

    Heat-shock proteins (hsps) are induced by a number of oxidative stresses. The proposal that the reduction products of oxygen initiate hsp induction was tested in rice embryos, capable of coleoptile growth under oxygen-free conditions. In such embryos, hsps could be detected by both in vivo labeling and in vitro translation of RNA using the reticulocyte lysate system. It is therefore improbable that the mechanism for hsp induction involves oxygen.

  14. [The human embryo after Dolly: new practices for new times].

    PubMed

    de Miguel Beriain, Iñigo

    2008-01-01

    The possiblity of cloning human beings introduced a lot of issues in our ethical and legal frameworks. In this paper, we will put the focus into the necessary changes in the concept of embryo that our legal systems will have to implement in order to face the new situation. The description of the embryo as a group of cells able to develop into a human being will be defended here as the best way of doing so.

  15. [Research with human embryo stem cells. Foundations and judicial limits].

    PubMed

    Eser, Albin; Koch, Hans-Georg

    2004-01-01

    Research with human embryos, and particularly, the use for scientific purposes of human embryonic stem cells has given raise to different sort of problems at the international level. One of the most strict regulation in this field, is this lecture Professors Albin Eser and Hans-Georg Koch analyse the german legal framework in relation with the use of embryos and human embryonic stem cells for scientific purposes.

  16. Spemann's organizer and self-regulation in amphibian embryos

    PubMed Central

    De Robertis, Edward M.

    2008-01-01

    In 1924, Spemann and Mangold demonstrated the induction of Siamese twins in transplantation experiments with salamander eggs. Recent work in amphibian embryos has followed their lead and uncovered that cells in signalling centres that are located at the dorsal and ventral poles of the gastrula embryo communicate with each other through a network of secreted growth-factor antagonists, a protease that degrades them, a protease inhibitor and bone-morphogenic-protein signals. PMID:16482093

  17. American Society for Reproductive Medicine: defining embryo donation.

    PubMed

    2009-12-01

    Building families through adoption of children has been supported by human society throughout history. Building families through reproductive donation of surplus embryos, in contrast, has become an option only since the dawn of assisted reproductive technologies. The ethical appropriateness of patients donating embryos to other patients for family building, or for research, including stem cell research, is well established and has been affirmed by this body and many others.

  18. Study Progress on Tissue Culture of Maize Mature Embryo

    NASA Astrophysics Data System (ADS)

    Wang, Hongzhen; Cheng, Jun; Cheng, Yanping; Zhou, Xioafu

    It has been paid more and more attention on maize tissue culture as it is a basic work in maize genetic transformation, especially huge breakthrough has been made in maize tissue culture utilizing mature embryos as explants in the recent years. This paper reviewed the study progress on maize tissue culture and plant regeneration utilizing mature embryos as explants from callus induction, subculture, plant regeneration and browning reduction and so on.

  19. The effects of experimentally induced adelphophagy in gastropod embryos.

    PubMed

    Thomsen, Olaf; Collin, Rachel; Carrillo-Baltodano, Allan

    2014-01-01

    Adelphophagy, development where embryos grow large by consuming morphologically distinct nutritive embryos or their own normal siblings is widespread but uncommon among animal phyla. Among invertebrates it is particularly common in some families of marine gastropods and segmented worms, but rare or unknown in other closely related families. In calyptraeid gastropods phylogenetic analysis indicates that adelphophagy has arisen at least 9 times from species with planktotrophic larval development. This pattern of frequent parallel evolution of adelphophagy suggests that the embryos of planktotrophic species might be predisposed to evolve adelphophagy. Here we used embryos of three species of planktotrophic calyptraeids, one from each of three major genera in the family (Bostrycapulus, Crucibulum, and Crepidula), to answer the following 3 questions: (1) Can embryos of species with planktotrophic development benefit, in terms of pre-hatching growth, from the ingestion of yolk and tissue from experimentally damaged siblings? (2) Does ingestion of this material from damaged siblings increase variation in pre-hatching size? and (3) Does this experimentally induced adelphophagy alter the allometry between the velum and the shell, increasing morphological similarity to embryos of normally adelphophagic species? We found an overall increase in shell length and velum diameter when embryos feed on damaged siblings within their capsules. There was no detectable increase in variation in shell length or velum diameter, or changes in allometry. The overall effect of our treatment was small compared to the embryonic growth observed in naturally adelphophagic development. However each embryo in our experiment probably consumed less than one sibling on average, whereas natural adelphophages often each consume 10-30 or more siblings. These results suggest that the ability to consume, assimilate, and benefit from yolk and tissue of their siblings is widespread across calyptraeids.

  20. Effect of different methods of cryopreservation on the cytoskeletal integrity of dromedary camel (Camelus dromedarius) embryos.

    PubMed

    Skidmore, J A; Schoevers, E; Stout, T A E

    2009-07-01

    This study examined the effect of different methods of cryopreservation on the cytoskeletal integrity of camel embryos. A total of 32 embryos were recovered on Days 6 and 7 after ovulation and measured before being frozen using either a conventional slow-cooling technique (n=12: six Day 6 and six Day 7 embryos) or vitrification (n=12: four Day 6 and eight Day 7). The remaining 8 'control' embryos (four Day 6 and four Day 7) were not cryopreserved but instead incubated in holding medium for 30 min. After thawing, warming or incubation, the embryos were stained with 4,6-diamino-2-phenylindole dihydrochloride (DAPI) to identify dead cells. Subsequently, the embryos were fixed in 4% paraformaldehyde, permeabilized and labelled with Alexa Fluor 488-Phalloidin to enable assessment of cytoskeleton integrity. Vitrified-warmed embryos contained a significantly higher percentage of dead cells than either conventionally frozen embryos or controls (P<0.05). Although the proportion of dead cells in conventionally frozen embryos tended to be higher than in controls, the difference was not significant (P> or =0.07). Whereas embryo size did not affect the number of dead cells in conventionally frozen embryos, vitrified-warmed embryos >300 microm in diameter had a significantly higher percentage of dead cells than embryos < or =300 microm (P=0.01). Cytoskeleton integrity was also affected by both freezing method and embryo diameter. All 8 control embryos had a Grade I cytoskeleton, compared with only 2/24 (8.3%) frozen or vitrified embryos. Of the 8 slow-frozen or vitrified embryos with a Grade III cytoskeleton post-thaw, 7 had been vitrified and 6 were larger (Day 7) embryos. These results indicate that while both slow-freezing and vitrification of camel embryos lead to cytoskeleton disruption and cell death, embryo quality is better preserved by slow-freezing.

  1. Effects of Pollutants on Eggs, Embryos and Larvae of Amphibian Species

    DTIC Science & Technology

    1976-04-01

    development of Xenopus Embryos 19 7. Effect of unsymmetrical dimethylhydrazine on development of Xenopus embryos 20 8. Anencephaly caused by exposure...control larva and a larva exposed to UDMH for 15 days commencing at gastrula. Fig. 8. ANENCEPHALY CAUSED BY EXPOSURE OF XENOPUS EMBRYOS TO UNSYM- METRICAL... Anencephaly - Absence of the cerebral and cerebellar hemispheres, with only a rudimentary brain stem. Embryo - The developing frog is an embryo ýrom

  2. Neutron induced bystander effect among zebrafish embryos

    NASA Astrophysics Data System (ADS)

    Ng, C. Y. P.; Kong, E. Y.; Kobayashi, A.; Suya, N.; Uchihori, Y.; Cheng, S. H.; Konishi, T.; Yu, K. N.

    2015-12-01

    The present paper reported the first-ever observation of neutron induced bystander effect (NIBE) using zebrafish (Danio rerio) embryos as the in vivo model. The neutron exposure in the present work was provided by the Neutron exposure Accelerator System for Biological Effect Experiments (NASBEE) facility at the National Institute of Radiological Sciences (NIRS), Chiba, Japan. Two different strategies were employed to induce NIBE, namely, through directly partnering and through medium transfer. Both results agreed with a neutron-dose window (20-50 mGy) which could induce NIBE. The lower dose limit corresponded to the threshold amount of neutron-induced damages to trigger significant bystander signals, while the upper limit corresponded to the onset of gamma-ray hormesis which could mitigate the neutron-induced damages and thereby suppress the bystander signals. Failures to observe NIBE in previous studies were due to using neutron doses outside the dose-window. Strategies to enhance the chance of observing NIBE included (1) use of a mono-energetic high-energy (e.g., between 100 keV and 2 MeV) neutron source, and (2) use of a neutron source with a small gamma-ray contamination. It appeared that the NASBEE facility used in the present study fulfilled both conditions, and was thus ideal for triggering NIBE.

  3. Eggs and embryos from the Cambrian.

    PubMed

    Morris, S C

    1998-08-01

    The early evolution of metazoans is a major focus of biological attention, but is the historical record revealed in the Cambrian "explosion" an accurate reflection of original events? The key questions concern the nature of the earliest animals and when they originated. One widely-mooted suggestion is that planktotrophic larvae, typified by the annelidan trochophore and echinoid pluteus, existed long before the metazoan radiations evident in the Cambrian fossil record. This idea could be consistent for recent evidence of divergence times, based on molecular "clocks," of phyla appearing well before the Cambrian. Now a surprising new discovery of eggs with blastomeres and embryos with well-defined anatomy from the Cambrian (c.530 Myr ago) of China and Siberia promises to extend the arena of debate. In one case a convincing ontogeny can be traced from eggs to adult tube-dwelling cnidarians. In the other example a possible protostome, unhatched and wrapped around the egg, shows segmentation and possibly nascent sclerites. In both, these cases development is direct, i.e., there is no evidence for any planktotrophic larval stage. The implications for our perceptions of both the Cambrian 'explosion' and metazoan phylogeny could be considerable.

  4. Embryo mechanics: balancing force production with elastic resistance during morphogenesis.

    PubMed

    Davidson, Lance A

    2011-01-01

    Morphogenesis requires the spatial and temporal control of embryo mechanics, including force production and mechanical resistance to those forces, to coordinate tissue deformation and large-scale movements. Thus, biomechanical processes play a key role in directly shaping the embryo. Additional roles for embryo mechanics during development may include the patterning of positional information and to provide feedback to ensure the success of morphogenetic movements in shaping the larval body and organs. To understand the multiple roles of mechanics during development requires familiarity with engineering principles of the mechanics of structures, the viscoelastic properties of biomaterials, and the integration of force and stress within embryonic structures as morphogenesis progresses. In this chapter, we review the basic engineering principles of biomechanics as they relate to morphogenesis, introduce methods for quantifying embryo mechanics and the limitations of these methods, and outline a formalism for investigating the role of embryo mechanics in birth defects. We encourage the nascent field of embryo mechanics to adopt standard engineering terms and test methods so that studies of diverse organisms can be compared and universal biomechanical principles can be revealed.

  5. [Characteristics of morphogenesis of the Japanese quail embryos during microgravity

    NASA Technical Reports Server (NTRS)

    Dadasheva, O. A.; Gur'eva, T. S.; Sychev, V. N.; Jehns, G.; Jahns, G. (Principal Investigator)

    1998-01-01

    Experiments performed in the period of 1995-1996 cooperatively with US investigators within the MIR/SHUTTLE and MIR/NASA space science projects continued exploration of avian embryogenesis in microgravity. Evaluation of Japanese quail embryos incubated in spaceflight microgravity showed that for the most part they were normally developed and compliant with duration of incubation. One of the major morphometric characteristics of embryo are its mass and size. Comparative analysis of body mass values in the space and laboratory and synchronous control groups pointed to a slight retardation. Body length of space embryos mimicked their mass curve. Data on the dynamics of mass and length of Japanese quail embryos support the well-known theory according to which growth and formation are distinguished by equifinality. No differences were revealed by the investigations of individual parts of embryonic bodies in the space and control groups. However, this finding was true only with regard to the embryos that had no developmental abnormalities. A part of embryos had defective eyes (microphtalmia), limbs (twisted fingers), and beaks.

  6. Resveratrol prevents nicotine-induced teratogenesis in cultured mouse embryos.

    PubMed

    Lin, Chunmei; Yon, Jung-Min; Jung, A Young; Lee, Jong Geol; Jung, Ki Youn; Kang, Jong-Koo; Lee, Beom Jun; Yun, Young Won; Nam, Sang-Yoon

    2012-11-01

    Nicotine, a major toxic component in tobacco smoke, leads to severe embryonic damage during organogenesis in embryos. We investigated whether resveratrol would positively influence nicotine-induced teratogenesis in mouse embryos (embryonic day 8.5) cultured for 48 h using a whole embryo culture system. Embryos exposed to nicotine (1mM) revealed significantly severe morphological anomalies, increased levels of caspase-3 mRNA and lipid peroxidation, and decreased levels of cytoplasmic superoxide dismutase (SOD), mitochondrial manganese SOD, cytosolic glutathione peroxidase, phospholipid hydroperoxide glutathione peroxidase, hypoxia-inducible factor 1α, Bcl-x(L), and sirtuin1 (SIRT1) mRNAs and SOD activity compared to those in the normal control group. However, when resveratrol (1×10(-8) μM or 1×10(-7) μM) was added concurrently to the embryos exposed to nicotine, all the parameters in above improved conspicuously. These findings indicate that resveratrol has a noted protective effect against nicotine-induced teratogenesis in mouse embryos through its antioxidative and anti-apoptotic effects.

  7. Automated image-based phenotypic analysis in zebrafish embryos

    PubMed Central

    Vogt, Andreas; Cholewinski, Andrzej; Shen, Xiaoqiang; Nelson, Scott; Lazo, John S.; Tsang, Michael; Hukriede, Neil A.

    2009-01-01

    Presently, the zebrafish is the only vertebrate model compatible with contemporary paradigms of drug discovery. Zebrafish embryos are amenable to automation necessary for high-throughput chemical screens, and optical transparency makes them potentially suited for image-based screening. However, the lack of tools for automated analysis of complex images presents an obstacle to utilizing the zebrafish as a high-throughput screening model. We have developed an automated system for imaging and analyzing zebrafish embryos in multi-well plates regardless of embryo orientation and without user intervention. Images of fluorescent embryos were acquired on a high-content reader and analyzed using an artificial intelligence-based image analysis method termed Cognition Network Technology (CNT). CNT reliably detected transgenic fluorescent embryos (Tg(fli1:EGFP)y1) arrayed in 96-well plates and quantified intersegmental blood vessel development in embryos treated with small molecule inhibitors of anigiogenesis. The results demonstrate it is feasible to adapt image-based high-content screening methodology to measure complex whole organism phenotypes. PMID:19235725

  8. Metal mesh vitrification (MMV) method for cryopreservation of porcine embryos.

    PubMed

    Fujino, Y; Kojima, T; Nakamura, Y; Kobayashi, H; Kikuchi, K; Funahashi, H

    2008-09-15

    The objective was to develop a simpler, more reliable vitrification method for porcine embryos. Prepubertal donor gilts were induced to ovulate with eCG and hCG, and then inseminated artificially. Morulae and expanding blastocysts approximately 200 microm in diameter were collected 6 or 7d after hCG treatment. Embryos collected from donor gilts were maintained, so as to be individually recognizable, and handled in batches of four or five. The embryos together with a minimum volume (<2 microL) of vitrification solution were placed onto stainless steel metal meshes or plastic plates, and then plunged into liquid nitrogen-metal mesh vitrification (MMV) and plastic plate vitrification (PPV), respectively. The meshes or plates were stored in 1.8-mL cryotubes submerged in liquid nitrogen. Stored embryos were subsequently removed, cultured in medium for 24 h, and then assessed for viability. The survival rate (84.4%) of expanding blastocysts cooled by MMV was higher than that (53.1%) of embryos cooled by PPV (P<0.05). There was no significant difference in total cell number between MMV and PPV. The survival rate of morulae cooled by MMV was 55.0%. Transfer of 200 expanding blastocysts cooled by MMV to 10 synchronized recipient gilts resulted in 37 live piglets from 7 recipients. In conclusion, the MMV method was an effective vitrification procedure for cryopreservation of expanding porcine blastocysts. However, there was a batch effect on embryo survival after vitrification.

  9. Embryo production by ovum pick up from live donors.

    PubMed

    Galli, C; Crotti, G; Notari, C; Turini, P; Duchi, R; Lazzari, G

    2001-04-01

    Embryo production by in vitro techniques has increased steadily over the years. For cattle where this technology is more advanced and is applied more, the number of in vitro produced embryos transferred to final recipients was over 30,000 in 1998. An increasing proportion of in vitro produced embryos are coming from oocytes collected from live donors by ultrasound-guided follicular aspiration (ovum pick up, OPU). This procedure allows the repeated production of embryos from live donors of particular value and is a serious alternative to superovulation. Ovum pick up is a very flexible technique. It can be performed twice a week for many weeks without side effects on the donor's reproductive career. The donor can be in almost any physiological status and still be suitable for oocyte recovery. A scanner with a sectorial or convex probe and a vacuum pump are required. Collection is performed with minimal stress to the donor. An average of 8 to 10 oocytes are collected per OPU with an average production of 2 transferable embryos. The laboratory production of embryos from such oocytes does not differ from that of oocytes harvested at slaughter as the results after transfer to final recipients. For other species such as buffalo and horses OPU has been attempted similarly to cattle and data will be presented and reviewed. For small ruminants, laparotomy or laparoscopy seems the only reliable route so far to collect oocytes from live donors.

  10. Embryo transfer and luteal support in natural cycles.

    PubMed

    Vlaisavljevic, Veljko

    2007-06-01

    Embryo transfer policy and luteal supplementation was reviewed, comparing literature data and the results from the Maribor IVF Centre. A retrospective analysis of 1024 cycles in patients undergoing IVF, intracytoplasmic sperm injection (ICSI) or testicular sperm aspiration in unstimulated cycles was carried out using four different approaches for cycle monitoring. This showed that the most successful protocol for monitoring was administration of human chorionic gonadotrophin (HCG) when serum oestradiol was >0.49 nmol/l and follicle diameter was at least 15 mm. The implantation rate per transferred embryo was higher when a blastocyst was transferred (42.8%) rather than a day-2 embryo (23.5%) in the same monitoring protocol. Analysis of the influence of patient age on the success of oocyte retrieval, oocyte fertilization, embryo transfer rate and delivery rate demonstrates that patient age does not influence the rate of positive oocyte retrieval or fertilization rate as much as it influences pregnancy rate per embryo transfer. The delivery rate per cycle was dramatically influenced by age in patients over 38 years. There is no clear evidence in the literature as to whether luteal phase support is necessary in natural cycles for IVF/ICSI. Comparing the data, a higher pregnancy rate was observed if HCG was administered after embryo transfer.

  11. Research on embryos in Turkey with ethical and legal aspects

    PubMed Central

    Vatanoğlu-Lutz, Emine Elif

    2012-01-01

    Technically, the term embryo refers to the products of conception after implantation into the wall of the womb, usually nearly two weeks after fertilization, up until the eighth week. Embryos contain stem cells which, according to scientists, could be used to cure a wide range of conditions. Stem cells can be coaxed into growing cells of any other type, which makes them potentially very useful indeed. However, removing stem cells from an embryo will kill the embryo, which some people object to. From the mid 1970s, IVF was being developed and research was carried out on the spare embryos produced. This research helped to improve IVF techniques, as well as to better understand the earliest stages of human development. Research also shed light on a variety of inheritable disorders. In Turkish Law, assisted reproduction treatment (ART) services are regulated with the Regulation of Assisted Reproductive Treatment Centers Act (RAPTCA) The Regulation was issued in 1987, but it has been amended several times since. Also, article 90 of the Turkish Penal Code covers some aspects of research on embryos. At the same time, the Biomedicine Convention (Oviedo Convention), signed by Turkey and which entered into force in 2003, has binding regulations about this issue. Different legal regulations and some ethical guidelines are in conflict with each other, creating much confusion for the researchers. In this paper these conflicts are discussed, giving some practical proposals. PMID:24592037

  12. A maternal Ahr null genotype sensitizes embryos to chemical teratogenesis.

    PubMed

    Thomae, Tami L; Glover, Edward; Bradfield, Christopher A

    2004-07-16

    The aryl hydrocarbon receptor (encoded by the Ahr locus) is a ligand-activated transcription factor that mediates the toxicology and teratology of 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin). In an effort to understand the role of the maternal compartment in dioxin teratology, we designed a breeding strategy that allowed us to compare the teratogenic response in embryos from Ahr(-/-) (null) and Ahr(+/+) (wild-type) dams. Using this strategy, we demonstrate that embryos from the Ahr(-/-) dams are 5-fold more sensitive to dioxin-induced cleft palate and hydronephrosis as compared with embryos from an Ahr(+/+) dam. Moreover, this increased teratogenic sensitivity extends beyond dioxin, because embryos from Ahr(-/-) dams exhibited a 9-fold increase in their sensitivity to the fetotoxic effects of the glucocorticoid, dexamethasone. In searching for an explanation for this increased sensitivity, we found that more dioxin and dexamethasone reached the embryos from Ahr(-/-) dams as compared with embryos from Ahr(+/+) dams. We propose that increased deposition of teratogens/fetotoxicants to the embryonic compartment is the result of porto-systemic shunting and/or blocked P4501A induction in Ahr(-/-) dams. In addition to demonstrating the importance of maternal AHR in teratogenesis, these data may have implications that reach beyond the mechanism of action of dioxin. In this regard, the Ahr(-/-) mouse may provide a system that allows pharmacological agents and toxicants to be more easily studied in a model where first pass clearance is a significant obstacle.

  13. Embryo Aggregation Promotes Derivation Efficiency of Outgrowths from Porcine Blastocysts

    PubMed Central

    Lee, Sang-Goo; Park, Jin-Kyu; Choi, Kwang-Hwan; Son, Hye-Young; Lee, Chang-Kyu

    2015-01-01

    Porcine embryonic stem cells (pESCs) have become an advantageous experimental tool for developing therapeutic applications and producing transgenic animals. However, despite numerous reports of putative pESC lines, deriving validated pESC lines from embryos produced in vitro remains difficult. Here, we report that embryo aggregation was useful for deriving pESCs from in vitro-produced embryos. Blastocysts derived from embryo aggregation formed a larger number of colonies and maintained cell culture stability. Our derived cell lines demonstrated expression of pluripotent markers (alkaline phosphatase, Oct4, Sox2, and Nanog), an ability to form embryoid bodies, and the capacity to differentiate into the three germ layers. A cytogenetic analysis of these cells revealed that all lines derived from aggregated blastocysts had normal female and male karyotypes. These results demonstrate that embryo aggregation could be a useful technique to improve the efficiency of deriving ESCs from in vitro-fertilized pig embryos, studying early development, and deriving pluripotent ESCs in vitro in other mammals. PMID:26580280

  14. Brooding fathers, not siblings, take up nutrients from embryos

    PubMed Central

    Sagebakken, Gry; Ahnesjö, Ingrid; Mobley, Kenyon B.; Gonçalves, Inês Braga; Kvarnemo, Charlotta

    2010-01-01

    It is well known that many animals with placenta-like structures provide their embryos with nutrients and oxygen. However, we demonstrate here that nutrients can pass the other way, from embryos to the parent. The study was done on a pipefish, Syngnathus typhle, in which males brood fertilized eggs in a brood pouch for several weeks. Earlier research has found a reduction of embryo numbers during the brooding period, but the fate of the nutrients from these ‘reduced’ embryos has been unknown. In this study, we considered whether (i) the brooding male absorbs the nutrients, (ii) siblings absorb them, or (iii) a combination of both. Males were mated to two sets of females, one of which had radioactively labelled eggs (using 14C-labelled amino acids), such that approximately half the eggs in the brood pouch were labelled. This allowed us to trace nutrient uptake from these embryos. We detected that 14C-labelled amino acids were transferred to the male brood pouch, liver and muscle tissue. However, we did not detect any significant 14C-labelled amino-acid absorption by the non-labelled half-siblings in the brood pouch. Thus, we show, to our knowledge, for the first time, that males absorb nutrients derived from embryos through their paternal brood pouch. PMID:19939847

  15. Aberrant DNA methylation reprogramming in bovine SCNT preimplantation embryos

    PubMed Central

    Zhang, Sheng; Chen, Xin; Wang, Fang; An, Xinglan; Tang, Bo; Zhang, Xueming; Sun, Liguang; Li, Ziyi

    2016-01-01

    DNA methylation reprogramming plays important roles in mammalian embryogenesis. Mammalian somatic cell nuclear transfer (SCNT) embryos with reprogramming defects fail to develop. Thus, we compared DNA methylation reprogramming in preimplantation embryos from bovine SCNT and in vitro fertilization (IVF) and analyzed the influence of vitamin C (VC) on the reprogramming of DNA methylation. The results showed that global DNA methylation followed a typical pattern of demethylation and remethylation in IVF preimplantation embryos; however, the global genome remained hypermethylated in SCNT preimplantation embryos. Compared with the IVF group, locus DNA methylation reprogramming showed three patterns in the SCNT group. First, some pluripotency genes (POU5F1 and NANOG) and repeated elements (satellite I and α-satellite) showed insufficient demethylation and hypermethylation in the SCNT group. Second, a differentially methylated region (DMR) of an imprint control region (ICR) in H19 exhibited excessive demethylation and hypomethylation. Third, some pluripotency genes (CDX2 and SOX2) were hypomethylated in both the IVF and SCNT groups. Additionally, VC improved the DNA methylation reprogramming of satellite I, α-satellite and H19 but not that of POU5F1 and NANOG in SCNT preimplantation embryos. These results indicate that DNA methylation reprogramming was aberrant and that VC influenced DNA methylation reprogramming in SCNT embryos in a locus-specific manner. PMID:27456302

  16. Embryo transfer in competition horses: Managing mares and expectations

    PubMed Central

    Campbell, M L H

    2014-01-01

    Embryo transfer (ET) is an accepted and successful technique for obtaining foals from mares without interrupting their competition careers. Recent research, however, suggests that the potential of factors including heat, exercise, repeated embryo flushing and repeated manipulation of the reproductive cycle using exogenous hormones to have a negative impact on fertility may have been underestimated. This paper reviews the evidence base for involvement of these factors in repeated failures to recover embryos from nongeriatric competition mares without obvious clinical or pathological indications of reproductive abnormalities. It concludes that, for some mares at least, a cessation of exercise for the periovulatory period and the period between ovulation and embryo flushing, combined with careful management of flushing-induced endometritis, and minimal hormonal manipulation of the reproductive cycle, may be necessary to optimise embryo recovery rates. Mare owners may have been encouraged to request ET for their mares following high-profile examples in the media of elite mares that have produced foals by ET whilst competing. The veterinarian should educate mare owners about the multiple factors that may affect the chances of recovering an embryo from their mares, and should manage the expectations of mare owners so that they do not approach ET programmes in the expectation that there will be no disruption to their training and competition plans. PMID:25977596

  17. In vitro culture of coconut (Cocos nucifera L.) zygotic embryos.

    PubMed

    Engelmann, Florent; Malaurie, Bernard; N'Nan, Oulo

    2011-01-01

    Coconut is a very important crop for millions of people in tropical countries. With coconut, in vitro culture protocols have been developed with two main objectives, viz. the large scale production of particular types of coconuts and the international exchange and conservation of coconut germplasm. The methods described in this chapter have been developed in the framework of collaborative activities between research institutes in Côte d'Ivoire and France. Two coconut embryo in vitro collecting protocols have been established, one consisting of storing the disinfected embryos in a KCl solution until they are brought back to the laboratory, where they are re-disinfected and inoculated in vitro under sterile conditions, and the other including in vitro inoculation of the embryos in the field. For international germplasm exchange, zygotic embryos inoculated in vitro in plastic test tubes or endosperm cylinders containing embryos in plastic bags are used. For in vitro culture, embryos are inoculated on semi-solid medium supplemented with sucrose and activated charcoal and placed in the dark, and then transferred to light conditions with the same (solid or liquid) medium once the first true leaf is visible and the root system has started developing.

  18. Tribolium embryo morphogenesis: may the force be with you.

    PubMed

    Benton, Matthew A; Pavlopoulos, Anastasios

    2014-01-01

    Development of multicellular organisms depends on patterning and growth mechanisms encoded in the genome, but also on the physical properties and mechanical interactions of the constituent cells that interpret these genetic cues. This fundamental biological problem requires integrated studies at multiple levels of biological organization: from genes, to cell behaviors, to tissue morphogenesis. We have recently combined functional genetics with live imaging approaches in embryos of the insect Tribolium castaneum, in order to understand their remarkable transformation from a uniform single-layered blastoderm into a condensed multi-layered embryo covered by extensive extra-embryonic tissues. We first developed a quick and reliable methodology to fluorescently label various cell components in entire Tribolium embryos. Live imaging of labeled embryos at single cell resolution provided detailed descriptions of cell behaviors and tissue movements during normal embryogenesis. We then compared cell and tissue dynamics between wild-type and genetically perturbed embryos that exhibited altered relative proportions of constituent tissues. This systematic comparison led to a qualitative model of the molecular, cellular and tissue interactions that orchestrate the observed epithelial rearrangements. We expect this work to establish the Tribolium embryo as a powerful and attractive model system for biologists and biophysicists interested in the molecular, cellular and mechanical control of tissue morphogenesis.

  19. Fusion of blastomeres in mouse embryos under the action of femtosecond laser radiation. Efficiency of blastocyst formation and embryo development

    SciTech Connect

    Osychenko, A A; Zalesskii, A D; Krivokharchenko, A S; Zhakhbazyan, A K; Nadtochenko, V A; Ryabova, A V

    2015-05-31

    Using the method of femtosecond laser surgery we study the fusion of two-cell mouse embryos under the action of tightly focused femtosecond laser radiation with the fusion efficiency reaching 60%. The detailed statistical analysis of the efficiency of blastomere fusion and development of the embryo up to the blastocyst stage after exposure of the embryos from different mice to a femtosecond pulse is presented. It is shown that the efficiency of blastocyst formation essentially depends on the biological characteristics of the embryo, namely, the strain and age of the donor mouse. The possibility of obtaining hexaploid embryonal cells using the methods of femtosecond laser surgery is demonstrated. (extreme light fields and their applications)

  20. Implantation of fresh and thawed-warmed embryos in single embryo transfer cycles: interpreting the initial beta-HCG.

    PubMed

    Sites, Cynthia K; St Marie, Peter; Rahil, Tayyab

    2015-03-01

    Little is known about the effects of human embryo cryopreservation on developmental potential. Initial beta-HCG, indicating embryo implantation, was measured in 322 single embryo transfer cycles (246 fresh and 76 thawed-warmed). Median initial beta-HCG was higher for fresh compared with thawed-warmed transfers (126 versus 100 mIU/ml; P = 0.04). Blastocyst slow cooling resulted in a lower initial beta-HCG compared with vitrification (P = 0.01). Live birth rates were lower for blastocyst slow cooling (25%) compared with vitrification (71%) and fresh transfer (70%). We conclude that cryopreservation may impair an embryo's ability to produce beta-HCG, but that vitrification does not impair developmental potential.

  1. Effect of Embryo Density on In Vitro Development and Gene Expression in Bovine In Vitro-fertilized Embryos Cultured in a Microwell System

    PubMed Central

    SUGIMURA, Satoshi; AKAI, Tomonori; HASHIYADA, Yutaka; AIKAWA, Yoshio; OHTAKE, Masaki; MATSUDA, Hideo; KOBAYASHI, Shuji; KOBAYASHI, Eiji; KONISHI, Kazuyuki; IMAI, Kei

    2012-01-01

    Abstract To identify embryos individually during in vitro development, we previously developed the well-of-the-well (WOW) dish, which contains 25 microwells. Here we investigated the effect of embryo density (the number of embryos per volume of medium) on in vitro development and gene expression of bovine in vitro-fertilized embryos cultured in WOW dishes. Using both conventional droplet and WOW culture formats, 5, 15, and 25 bovine embryos were cultured in 125 µl medium for 168 h. The blastocysts at Day 7 were analyzed for number of cells and expression of ten genes (CDX2, IFN-tau, PLAC8, NANOG, OCT4, SOX2, AKR1B1, ATP5A1, GLUT1 and IGF2R). In droplet culture, the rates of formation of >4-cell cleavage embryos and blastocysts were significantly lower in embryos cultured at 5 embryos per droplet than in those cultured at 15 or 25 embryos per droplet, but not in WOW culture. In both droplet and WOW culture, developmental kinetics and blastocyst cell numbers did not differ among any groups. IFN-tau expression in embryos cultured at 25 embryos per droplet was significantly higher than in those cultured at 15 embryos per droplet and in artificial insemination (AI)-derived blastocysts. Moreover, IGF2R expression was significantly lower in the 25-embryo group than in the 5-embryo group and in AI-derived blastocysts. In WOW culture, these expressions were not affected by embryo density and were similar to those in AI-derived blastocysts. These results suggest that, as compared with conventional droplet culture, in vitro development and expression of IFN-tau and IGF2R in the microwell system may be insensitive to embryo density. PMID:23154384

  2. Effect of embryo density on in vitro development and gene expression in bovine in vitro-fertilized embryos cultured in a microwell system.

    PubMed

    Sugimura, Satoshi; Akai, Tomonori; Hashiyada, Yutaka; Aikawa, Yoshio; Ohtake, Masaki; Matsuda, Hideo; Kobayashi, Shuji; Kobayashi, Eiji; Konishi, Kazuyuki; Imai, Kei

    2013-01-01

    To identify embryos individually during in vitro development, we previously developed the well-of-the-well (WOW) dish, which contains 25 microwells. Here we investigated the effect of embryo density (the number of embryos per volume of medium) on in vitro development and gene expression of bovine in vitro-fertilized embryos cultured in WOW dishes. Using both conventional droplet and WOW culture formats, 5, 15, and 25 bovine embryos were cultured in 125 μl medium for 168 h. The blastocysts at Day 7 were analyzed for number of cells and expression of ten genes (CDX2, IFN-tau, PLAC8, NANOG, OCT4, SOX2, AKR1B1, ATP5A1, GLUT1 and IGF2R). In droplet culture, the rates of formation of >4-cell cleavage embryos and blastocysts were significantly lower in embryos cultured at 5 embryos per droplet than in those cultured at 15 or 25 embryos per droplet, but not in WOW culture. In both droplet and WOW culture, developmental kinetics and blastocyst cell numbers did not differ among any groups. IFN-tau expression in embryos cultured at 25 embryos per droplet was significantly higher than in those cultured at 15 embryos per droplet and in artificial insemination (AI)-derived blastocysts. Moreover, IGF2R expression was significantly lower in the 25-embryo group than in the 5-embryo group and in AI-derived blastocysts. In WOW culture, these expressions were not affected by embryo density and were similar to those in AI-derived blastocysts. These results suggest that, as compared with conventional droplet culture, in vitro development and expression of IFN-tau and IGF2R in the microwell system may be insensitive to embryo density.

  3. Supplementation of bovine embryo culture medium with L-arginine improves embryo quality via nitric oxide production.

    PubMed

    Santana, Priscila Di Paula Bessa; Silva, Thiago Velasco Guimarães; da Costa, Nathália Nogueira; da Silva, Bruno Barauna; Carter, Timothy Frederick; Cordeiro, Marcela da Silva; da Silva, Bruno José Martins; Santos, Simone do Socorro Damasceno; Herculano, Anderson Manoel; Adona, Paulo Roberto; Ohashi, Otávio Mitio; Miranda, Moysés dos Santos

    2014-10-01

    Nitric oxide (NO) is a cell-signaling molecule that regulates a variety of molecular pathways. We investigated the role of NO during preimplantation embryonic development by blocking its production with an inhibitor or supplementing in vitro bovine embryo cultures with its natural precursor, L-arginine, over different periods. Endpoints evaluated included blastocyst rates, development kinetics, and embryo quality. Supplementation with the NO synthase inhibitor N-Nitro-L-arginine-methyl ester (L-NAME) from Days 1 to 8 of culture decreased blastocyst (P < 0.05) and hatching (P < 0.05) rates. When added from Days 1 to 8, 50 mM L-arginine decreased blastocyst rates (P < 0.001); in contrast, when added from Days 5 to 8, 1 mM L-arginine improved embryo hatching rates (P < 0.05) and quality (P < 0.05) as well as increased POU5F1 gene expression (P < 0.05) as compared to the untreated control. Moreover, NO levels in the medium during this culture period positively correlated with the increased embryo hatching rates and quality (P < 0.05). These data suggest exerts its positive effects during the transition from morula to blastocyst stage, and that supplementing the embryo culture medium with L-arginine favors preimplantation development of bovine embryos.

  4. Artificial intelligence techniques for embryo and oocyte classification.

    PubMed

    Manna, Claudio; Nanni, Loris; Lumini, Alessandra; Pappalardo, Sebastiana

    2013-01-01

    One of the most relevant aspects in assisted reproduction technology is the possibility of characterizing and identifying the most viable oocytes or embryos. In most cases, embryologists select them by visual examination and their evaluation is totally subjective. Recently, due to the rapid growth in the capacity to extract texture descriptors from a given image, a growing interest has been shown in the use of artificial intelligence methods for embryo or oocyte scoring/selection in IVF programmes. This work concentrates the efforts on the possible prediction of the quality of embryos and oocytes in order to improve the performance of assisted reproduction technology, starting from their images. The artificial intelligence system proposed in this work is based on a set of Levenberg-Marquardt neural networks trained using textural descriptors (the local binary patterns). The proposed system was tested on two data sets of 269 oocytes and 269 corresponding embryos from 104 women and compared with other machine learning methods already proposed in the past for similar classification problems. Although the results are only preliminary, they show an interesting classification performance. This technique may be of particular interest in those countries where legislation restricts embryo selection. One of the most relevant aspects in assisted reproduction technology is the possibility of characterizing and identifying the most viable oocytes or embryos. In most cases, embryologists select them by visual examination and their evaluation is totally subjective. Recently, due to the rapid growth in our capacity to extract texture descriptors from a given image, a growing interest has been shown in the use of artificial intelligence methods for embryo or oocyte scoring/selection in IVF programmes. In this work, we concentrate our efforts on the possible prediction of the quality of embryos and oocytes in order to improve the performance of assisted reproduction technology

  5. Embryo implantation and GnRH antagonists: embryo implantation: the Rubicon for GnRH antagonists.

    PubMed

    Hernandez, E R

    2000-06-01

    When gonadotrophin-releasing hormone (GnRH) was discovered, the agonist and antagonist of GnRH were developed to control the release of FSH and LH by the gonadotrophs. More than 10 years of research were needed to develop a GnRH antagonist free of histamine release. Recent studies have shown that these GnRH antagonists are effective in preventing a rise in LH during ovarian stimulation in IVF. However, a decrease in ongoing pregnancies seems to suggest that implantation rates per transferred embryo are reduced in GnRH antagonist-stimulated cycles. In my opinion, these data highlight an area less well known to clinicians: the role of the GnRH antagonist at the cellular level in extrapituitary tissues. There are sufficient data in the literature suggesting that GnRH antagonist is an inhibitor of the cell cycle by decreasing the synthesis of growth factors. Given that, for folliculogenesis, blastomere formation and endometrium development, mitosis is everything; the interaction between the GnRH antagonist and the GnRH receptor (present in all these cells and tissues) may compromise the mitotic programme of these cells. This is the Rubicon for the GnRH antagonist: to demonstrate irrevocably that, at the minimal doses necessary to suppress LH release, it does not affect processes such as implantation, embryo development and folliculogenesis.

  6. Potential risk of equine herpes virus 1 (EHV-1) transmission by equine embryo transfer.

    PubMed

    Hebia, I; Fiéni, F; Duchamp, G; Destrumelle, S; Pellerin, J-L; Zientara, S; Vautherot, J-F; Bruyas, J-F

    2007-06-01

    The objective of this study was to determine whether the 10 wash cycles proposed by the International Embryo Transfer Society (IETS) for bovine embryos efficiently decontaminated equine embryos exposed to equine herpes virus 1 (EHV-1) in vitro. Donor mares and stallions were individually screened and shown to be negative for the virus by PCR detection of EHV-1 DNA in blood leukocytes, semen, and uterine lavages in which embryos were recovered. Twenty embryos were recovered and randomly assigned to one of two groups: 10 embryos were exposed for 24h to infectious EHV-1 at 10(6)TCID(50)/ml, and 10 embryos were used as negative controls. Exposed embryos were washed in accordance with IETS recommendations for ruminant and porcine embryos, before being incubated for 24 h with semiconfluent rabbit kidney (RK13) cells to detect any cytopathic effects (CPE), and finally tested for the presence of EHV-1 viral DNA by PCR. The embryo washing media were also assayed for the virus on RK 13 cells and by PCR. Control embryos were neither exposed to the virus nor washed. EHV-1 was not found in the control embryos, or in the last five washes of the exposed embryos. However, the virus was detected in 7/10 of the embryos exposed to EHV-1 for 24h, as well as in the first five washes of the embryos. The gradual disappearance of EHV-1 from the 10 successive wash solutions from the exposed embryos and the detection of viral DNA in 7/10 washed embryos by PCR, demonstrated that the washing procedure was unable to remove EHV-1 and suggested that EHV-1 could be attached to the acellular layer surrounding embryos (zona pellucida or capsule) or had penetrated the embryo.

  7. Forebrain neurogenesis: From embryo to adult

    PubMed Central

    Dennis, Daniel; Picketts, David; Slack, Ruth S.; Schuurmans, Carol

    2017-01-01

    A satellite symposium to the Canadian Developmental Biology Conference 2016 was held on March 16–17, 2016 in Banff, Alberta, Canada, entitled Forebrain Neurogenesis: From embryo to adult. The Forebrain Neurogenesis symposium was a focused, high-intensity meeting, bringing together the top Canadian and international researchers in the field. This symposium reported the latest breaking news, along with ‘state of the art’ techniques to answer fundamental questions in developmental neurobiology. Topics covered ranged from stem cell regulation to neurocircuitry development, culminating with a session focused on neuropsychiatric disorders. Understanding the underlying causes of neurodevelopmental disorders such as autism spectrum disorder (ASD) and attention deficit/hyperactivity disorder (ADHD) is of great interest as diagnoses of these conditions are climbing at alarming rates. For instance, in 2012, the Centers for Disease Control reported that the prevalence rate of ASD in the U.S. was 1 in 88; while more recent data indicate that the number is as high as 1 in 68 (Centers for Disease Control and Prevention MMWR Surveillance Summaries. Vol. 63. No. 2). Similarly, the incidence of ASD is on the rise in Canada, increasing from 1 in 150 in 2000 to 1 in 63 in 2012 in southeastern Ontario (Centers for Disease Control and Prevention). Currently very little is known regarding the deficits underlying these neurodevelopmental conditions. Moreover, the development of effective therapies is further limited by major gaps in our understanding of the fundamental processes that regulate forebrain development and adult neurogenesis. The Forebrain Neurogenesis satellite symposium was thus timely, and it played a key role in advancing research in this important field, while also fostering collaborations between international leaders, and inspiring young researchers.

  8. Live embryo imaging to follow cell cycle and chromosomes stability after nuclear transfer.

    PubMed

    Balbach, Sebastian T; Boiani, Michele

    2015-01-01

    Nuclear transfer (NT) into mouse oocytes yields a transcriptionally and functionally heterogeneous population of cloned embryos. Most studies of NT embryos consider only embryos at predefined key stages (e.g., morula or blastocyst), that is, after the bulk of reprogramming has taken place. These retrospective approaches are of limited use to elucidate mechanisms of reprogramming and to predict developmental success. Observing cloned embryo development using live embryo cinematography has the potential to reveal otherwise undetectable embryo features. However, light exposure necessary for live cell cinematography is highly toxic to cloned embryos. Here we describe a protocol for combined bright-field and fluorescence live-cell imaging of histone H2b-GFP expressing mouse embryos, to record cell divisions up to the blastocyst stage. This protocol, which can be adapted to observe other reporters such as Oct4-GFP or Nanog-GFP, allowed us to quantitatively analyze cleavage kinetics of cloned embryos.

  9. How couples who have undergone in vitro fertilization decide what to do with surplus frozen embryos.

    PubMed

    Nachtigall, Robert D; Mac Dougall, Kirstin; Harrington, Jennifer; Duff, Julia; Lee, Matthew; Becker, Gay

    2009-12-01

    In a qualitative interview study of 77 families with stored frozen embryos, we found that while embryo disposition decision making was influenced by individual life circumstances, embryo quantity/quality, personal values, embryo conceptualization, and clinic information, it was a stepwise process that could be represented as three sequential questions: (1) Will the embryos be used for additional attempts at conception? If not, (2) Will the embryos remain in storage? And if not, (3) Will the embryos be donated to other people or to science, or will they be destroyed? While almost two-thirds (63%) of participants kept their embryos in storage after 5 years, either passively through disagreement or indecision or actively to maintain embryo potential, avert feelings of loss, or as psychological or genetic "insurance," IVF clinic support and detailed information about options motivated families to make disposition decisions.

  10. Following the course of pre-implantation embryo patterning by non-linear microscopy.

    PubMed

    Kyvelidou, Christiana; Tserevelakis, George J; Filippidis, George; Ranella, Anthi; Kleovoulou, Anastasia; Fotakis, Costas; Athanassakis, Irene

    2011-12-01

    Embryo patterning is subject to intense investigation. So far only large, microscopically obvious structures like polar body, cleavage furrow, pro-nucleus shape can be evaluated in the intact embryo. Using non-linear microscopic techniques, the present work describes new methodologies to evaluate pre-implantation mouse embryo patterning. Third Harmonic Generation (THG) imaging, by detecting mitochondrial/lipid body structures, could provide valuable and complementary information as to the energetic status of pre-implantation embryos, time evolution of different developmental stages, embryo polarization prior to mitotic division and blastomere equivalence. Quantification of THG imaging detected highest signalling in the 2-cell stage embryos, while evaluating a 12-18% difference between blastomeres at the 8-cell stage embryos. Such a methodology provides novel, non-intrusive imaging assays to follow up intracellular structural patterning associated with the energetic status of a developing embryo, which could be successfully used for embryo selection during the in vitro fertilization process.

  11. Supplementation of insulin-transferrin-selenium to embryo culture medium improves the in vitro development of pig embryos.

    PubMed

    Das, Ziban Chandra; Gupta, Mukesh Kumar; Uhm, Sang Jun; Lee, Hoon Taek

    2014-08-01

    Insulin, transferrin and selenium (ITS) supplementation to oocyte maturation medium improves the post-fertilization embryonic development in pigs. ITS is also commonly used as a supplement for the in vitro culture (IVC) of embryos and stem cells in several mammalian species. However, its use during IVC of pig embryos has not been explored. This study investigated the effect of ITS supplementation to IVC medium on the in vitro development ability of pig embryos produced by parthenogenetic activation (PA), in vitro fertilization (IVF) or somatic cell nuclear transfer (SCNT). We observed that ITS had no significant effect on the rate of first cleavage (P > 0.05). However, the rate of blastocyst formation in ITS-treated PA (45.3 ± 1.9 versus 27.1 ± 2.3%), IVF (31.6 ± 0.6 versus 23.5 ± 0.6%) and SCNT (17.6 ± 2.3 versus 10.7 ± 1.4%) embryos was significantly higher (P < 0.05) than those of non-treated controls. Culture of PA embryos in the presence of ITS also enhanced the expansion and hatching ability (29.1 ± 3.0 versus 18.2 ± 3.8%; P < 0.05) of blastocysts and increased the total number of cells per blastocyst (53 ± 2.5 versus 40.9 ± 2.6; P < 0.05). Furthermore, the beneficial effect of ITS on PA embryos was associated with significantly reduced level of intracellular reactive oxygen species (ROS) (20.0 ± 2.6 versus 46.9 ± 3.0). However, in contrast to PA embryos, ITS had no significant effect on the blastocyst quality of IVF and SCNT embryos (P > 0.05). Taken together, these data suggest that supplementation of ITS to the IVC medium exerts a beneficial but differential effect on pig embryos that varies with the method of embryo production in vitro.

  12. Characterization of somatic embryo attached structures in Feijoa sellowiana Berg. (Myrtaceae).

    PubMed

    Correia, Sandra M; Canhoto, Jorge M

    2010-06-01

    The presence of an attached organ to somatic embryos of angiosperms connecting the embryo to the supporting tissue has been a subject of controversy. This study shows that 67% of the morphologically normal somatic embryos of Feijoa sellowiana possess this type of organ and that its formation was not affected by culture media composition. Histological and ultrastructural analysis indicated that the attached structures of somatic embryos displayed a great morphological diversity ranging from a few cells to massive and columnar structures. This contrast with the simple suspensors observed in zygotic embryos which were only formed by five cells. As well as the suspensor of zygotic embryos, somatic embryo attached structures undergo a process of degeneration in later stages of embryo development. Other characteristic shared by zygotic suspensors and somatic embryo attached structures was the presence of thick cell walls surrounding the cells. Elongated thin filaments were often associated with the structures attached to somatic embryos, whereas in other cases, tubular cells containing starch grains connected the embryo to the supporting tissue. These characteristics associated with the presence of plasmodesmata in the cells of the attached structures seem to indicate a role on embryo nutrition. However, cell proliferation in the attached structures resulting into new somatic embryos may also suggest a more complex relationship between the embryo and the structures connecting it to the supporting tissue.

  13. Preimplantation embryo metabolism and culture systems: experience from domestic animals and clinical implications.

    PubMed

    Absalón-Medina, V A; Butler, W R; Gilbert, R O

    2014-04-01

    Despite advantages of in vitro embryo production in many species, widespread use of this technology is limited by generally lower developmental competence of in vitro derived embryos compared to in vivo counterparts. Regardless, in vivo or in vitro gametes and embryos face and must adjust to multiple microenvironments especially at preimplantation stages. Moreover, the embryo has to be able to further adapt to environmental cues in utero to result in the birth of live and healthy offspring. Enormous strides have been made in understanding and meeting stage-specific requirements of preimplantation embryos, but interpretation of the data is made difficult due to the complexity of the wide array of culture systems and the remarkable plasticity of developing embryos that seem able to develop under a variety of conditions. Nevertheless, a primary objective remains meeting, as closely as possible, the preimplantation embryo requirements as provided in vivo. In general, oocytes and embryos develop more satisfactorily when cultured in groups. However, optimization of individual culture of oocytes and embryos is an important goal and area of intensive current research for both animal and human clinical application. Successful culture of individual embryos is of primary importance in order to avoid ovarian superstimulation and the associated physiological and psychological disadvantages for patients. This review emphasizes stage specific shifts in embryo metabolism and requirements and research to optimize in vitro embryo culture conditions and supplementation, with a view to optimizing embryo culture in general, and culture of single embryos in particular.

  14. Fresh embryo donation for human embryonic stem cell (hESC) research: the experiences and values of IVF couples asked to be embryo donors

    PubMed Central

    Haimes, E.; Taylor, K.

    2009-01-01

    BACKGROUND This article reports on an investigation of the views of IVF couples asked to donate fresh embryos for research and contributes to the debates on: the acceptability of human embryonic stem cell (hESC) research, the moral status of the human embryo and embryo donation for research. METHODS A hypothesis-generating design was followed. All IVF couples in one UK clinic who were asked to donate embryos in 1 year were contacted 6 weeks after their pregnancy result. Forty four in-depth interviews were conducted. RESULTS Interviewees were preoccupied with IVF treatment and the request to donate was a secondary consideration. They used a complex and dynamic system of embryo classification. Initially, all embryos were important but then their focus shifted to those that had most potential to produce a baby. At that point, ‘other’ embryos were less important though they later realise that they did not know what happened to them. Guessing that these embryos went to research, interviewees preferred not to contemplate what that might entail. The embryos that caused interviewees most concern were good quality embryos that might have produced a baby but went to research instead. ‘The’ embryo, the morally laden, but abstract, entity, did not play a central role in their decision-making. CONCLUSIONS This study, despite missing those who refuse to donate embryos, suggests that debates on embryo donation for hESC research should include the views of embryo donors and should consider the social, as well as the moral, status of the human embryo. PMID:19502616

  15. Phytohemagglutinin facilitates the aggregation of blastomere pairs from Day 5 donor embryos with Day 4 host embryos for chimeric bovine embryo multiplication.

    PubMed

    Simmet, Kilian; Reichenbach, Myriam; Reichenbach, Horst-Dieter; Wolf, Eckhard

    2015-12-01

    Multiplication of bovine embryos by the production of aggregation chimeras is based on the concept that few blastomeres of a donor embryo form the inner cell mass (ICM) and thus the embryo proper, whereas cells of a host embryo preferentially contribute to the trophectoderm (TE), the progenitor cells of the embryonic part of the placenta. We aggregated two fluorescent blastomeres from enhanced green fluorescent protein (eGFP) transgenic Day 5 morulae with two Day 4 embryos that did not complete their first cleavage until 27 hours after IVF and tested the effect of phytohemagglutinin-L (PHA) on chimeric embryo formation. The resulting blastocysts were characterized by differential staining of cell lineages using the TE-specific factor CDX2 and confocal laser scanning microscopy to facilitate the precise localization of eGFP-positive cells. The proportions of blastocyst development of sandwich aggregates with (n = 99) and without PHA (n = 46) were 85.9% and 54.3% (P < 0.05), respectively. Epifluorescence microscopy showed that the proportion of blastocysts with eGFP-positive cells in the ICM was higher in the PHA group than in the no-PHA group (40% vs. 16%; P < 0.05). Confocal laser scanning microscopy revealed that the total cell numbers of blastocysts from the PHA group of aggregation chimeras (n = 17; 207.8 ± 67.3 [mean ± standard deviation]) were higher (P < 0.05) than those of embryos without ZP and exposed to PHA (n = 30; 159.6 ± 42.2) and of handling control embryos (n = 19; 176.9 ± 53.3). The same was true for ICM cell counts (56.5 ± 22.0 vs. 37.7 ± 14.2 and 38.7 ± 12.4) and TE cell counts (151.2 ± 58.0 vs. 121.9 ± 37.4 and 138.3 ± 53.0), whereas the ICM/total cell number ratio was not different between the groups. Of the 17 chimeric blastocysts analyzed by confocal laser scanning microscopy, nine had eGFP-positive cells (three of them in the ICM, three in the TE, and three in both lineages). When integration in

  16. Creating and selling embryos for "donation": ethical challenges.

    PubMed

    Klitzman, Robert; Sauer, Mark V

    2015-02-01

    The commercial creation and sale of embryos has begun, which poses a series of ethical questions that have received little scholarly attention. Some of the concerns that arise are similar to those posed by the sale of gametes, while other issues differ markedly. Questions emerge, first, regarding the rights of the unborn children and their ability to know their biological parents. Companies that create human embryos de novo may wish to keep gamete providers anonymous. Many of these offspring thus will never learn that their parents are not their biologic parents. Yet, such disclosures, regarding not only one but both of these biologic parents, may be important for these individuals; and a lack of this knowledge may impede their physical and psychological health. Second, questions surface regarding the fees that providers should charge for embryos and whether these amounts should vary based on the traits of 1 or both of the gamete donors. Some prospective parents may seek specific traits in a baby (eg, height or eye/hair coloring), which prompts the creation of embryos from 2 gamete donors who possess these characteristics. Third, ownership of embryos created without an advanced directive by patients poses dilemmas (eg, disposition of any remaining embryos). Fourth, guidelines do not yet exist to limit the number of embryos sold from each pair of gamete donors. Hence, unbeknownst to each other, full siblings could potentially meet, get married, and procreate. This discussion has several critical implications for future practice and professional education and policy. Patients with diseases associated with genetic tests may well ask obstetricians, gynecologists, and other physicians about these techniques and practices. Clinicians can refer such patients to assisted reproductive technology specialists; however, familiarity with the basic aspects of the issues and complexities involved could aid these providers and their patients Several of these issues can be

  17. Creating and Selling Embryos for “Donation”: Ethical Challenges

    PubMed Central

    Klitzman, Robert; Sauer, Mark V.

    2015-01-01

    The commercial creation and sale of embryos has begun, posing a series of ethical questions that have received little scholarly attention. Some of the concerns that arise are similar to those posed by the sale of gametes, while other issues differ markedly. Questions emerge, firstly, regarding the rights of the unborn children – their ability to know their biological parents. Companies that create human embryos de novo may wish to keep gamete providers anonymous. Many of these offspring will thus never learn that their parents are not their biological parents. Yet, such disclosures – regarding not only one, but both of these biological parents – may be important for these individuals; and lack of this knowledge may impede their physical and psychological health. Secondly, questions surface regarding the fees that providers should charge for embryos, and whether these amounts should vary based on the traits of one or both of the gamete donors. Some prospective parents may seek specific traits in a baby (e.g., height or eye/hair coloring), prompting creation of embryos from two gamete donors who possess these characteristics. Thirdly, ownership of embryos created without an advanced directive by patients poses dilemmas – e.g., disposition of any remaining embryos. Fourthly, guidelines do not yet exist to limit the number of embryos sold from each pair of gamete donors. Hence, unbeknownst to each other, full siblings could potentially meet, get married and procreate. This discussion has several critical implications for future practice, and professional education and policy. Patients with diseases associated with genetic tests may well ask obstetricians, gynecologists and other physicians about these techniques and practices. Clinicians can refer such patients to Assisted Reproductive Technology specialists, but familiarity with the basic aspects of the issues and complexities involved could aid themselves and their patients Several of these issues can be

  18. Developmental imaging: the avian embryo hatches to the challenge.

    PubMed

    Kulesa, Paul M; McKinney, Mary C; McLennan, Rebecca

    2013-06-01

    The avian embryo provides a multifaceted model to study developmental mechanisms because of its accessibility to microsurgery, fluorescence cell labeling, in vivo imaging, and molecular manipulation. Early two-dimensional planar growth of the avian embryo mimics human development and provides unique access to complex cell migration patterns using light microscopy. Later developmental events continue to permit access to both light and other imaging modalities, making the avian embryo an excellent model for developmental imaging. For example, significant insights into cell and tissue behaviors within the primitive streak, craniofacial region, and cardiovascular and peripheral nervous systems have come from avian embryo studies. In this review, we provide an update to recent advances in embryo and tissue slice culture and imaging, fluorescence cell labeling, and gene profiling. We focus on how technical advances in the chick and quail provide a clearer understanding of how embryonic cell dynamics are beautifully choreographed in space and time to sculpt cells into functioning structures. We summarize how these technical advances help us to better understand basic developmental mechanisms that may lead to clinical research into human birth defects and tissue repair.

  19. Patients' views on the embryo storage time limits.

    PubMed

    Pereira, Margarida; Samorinha, Catarina; Alves, Elisabete; Machado, Helena; Amorim, Mariana; Silva, Susana

    2015-08-01

    The establishment of the length of embryo storage has been based on socio-political criteria. There are different regulations, guidelines and health care policies worldwide. This mixed-methods study aimed to assess the opinion of patients about the embryo storage time limit, and the perception of the criteria underlying the establishment of the storage period offered to them. Between August 2011 and December 2012, 534 IVF patients from Portugal participated in a quantitative questionnaire and 34 couples were interviewed. Overall, 38% of participants preferred the duration of 4-5 years, 38% extended it beyond 5 years and 23% indicated 3 years. Having experienced at least one previous cycle was directly associated with agreeing with a duration of storage longer than 5 years, for both women and men. Having children was inversely associated with longer duration of storage, among women. One-third of the 34 interviewed couples stated that their knowledge concerning embryo storage was insufficient. Nevertheless, all the interviewees reported at least one possible reason for the legal establishment of the storage period offered to them, highlighting financial costs and decreased embryo quality. There are misconceptions and gaps in awareness of cryopreservation, which may shape patients' opinions. Accurate information regarding policy on storage of embryos is needed.

  20. Planetary Embryo Bow Shocks as a Mechanism for Chondrule Formation

    NASA Astrophysics Data System (ADS)

    Mann, Christopher; Boley, Aaron C.; Morris, Melissa A.

    2015-01-01

    We investigate the plausibility of a planetary embryo bow shock as a mechanism for chondrule formation in the early solar system. A Mars-size planetary embryo traveling on a moderately excited orbit through the dusty early environment of the solar system will experience supersonic velocities relative to the circularly orbiting gas and dust. The resulting bow shock can thermally process solids that pass through it, with a wide range of possible conditions depending on impact radius. Volatile outgassing by the embryo along with some gas capture from the surrounding nebula can produce temporary atmospheres. We use radiation hydrodynamics simulations with direct particle integration to model the consequences of solids that encounter a bow shock produced by a 3000 km embryo with relative speeds to the gas of 5, 6, and 7 km/s. The embryos are envisaged to be surrounded by low- and high-mass atmospheres (0.75 and 6.25 Martian-mass atmospheres, respectively), and we explore different opacities for the gas. We find that a high-mass atmosphere and low dust opacity can produce peak temperatures and cooling rates that are most consistent with constraints set by chondrule furnace studies for plausible shock speeds.

  1. Chlormequat chloride retards rat embryo growth in vitro.

    PubMed

    Xiagedeer, Bayindala; Wu, Shuang; Liu, Yingjuan; Hao, Weidong

    2016-08-01

    Chlormequat chloride is the most widely used plant growth regulator in agriculture to promote sturdier growth of grain crops by avoidance of lodging. Therefore, human exposure to chlormequat chloride is very common, but its developmental toxicity has not been studied. Thus, we investigated the developmental toxicity of chlormequat chloride by applying rat whole embryo culture (WEC) model, limb bud micromass culture and 3T3 fibroblast cytotoxicity test. Chlormequat chloride at 150μg/ml (0.93mM) retarded the rat embryo growth without causing significant morphological malformations and at 500μg/ml (3.1mM) caused both retardation and morphological malformation of the embryos. However, the proliferation and differentiation of limb bud cells were not affected by chlormequat chloride at as high as up to 1000μg/ml (6.2mM) applied. This concentration of chlormequat chloride did not affect the cell viability as examined by 3T3 fibroblast cytotoxicity test either, suggesting that cellular toxicity may not play a role in chlormequat induced inhibition of rat embryo growth. Collectively, our results demonstrated that chlormequat chloride may affect embryo growth and development without inhibiting cell viability.

  2. Energetics of lizard embryos are not canalized by thermal acclimation.

    PubMed

    Angilletta, Michael J; Lee, Vivian; Silva, Albert C

    2006-01-01

    In some species of ectotherms, temperature has little or no effect on the amount of energy expended during embryonic development. This phenomenon can result from either of two mechanisms: (1) a shorter incubation period at higher temperatures, which offsets the expected increase in metabolic rate, or (2) a compensatory decrease in the rate at which embryos expend energy for maintenance. To distinguish the relative importance of these two mechanisms, we quantified the acute and chronic effects of temperature on embryonic metabolism in the eastern fence lizard (Sceloporus undulatus). First, we measured metabolic rates of individual embryos at 27 degrees, 31 degrees, and 34 degrees C. Second, we examined the capacity for thermal acclimation by measuring the metabolic rates of embryos at 30 degrees C, after a period of incubation at either 28 degrees or 32 degrees C. As with adult reptiles, the metabolic rates of embryos increased with an acute increase in temperature; the Q(10) of metabolic rate from 27 degrees to 34 degrees C was 2.1 (+/-0.2). No evidence of thermal acclimation was observed either early or late in development. In S. undulatus, a shorter incubation period at higher temperatures appears to play the primary role in canalizing the energy budget of an embryo, but a reduction in the cost of growth could play a secondary role.

  3. Heat shock protein expression enhances heat tolerance of reptile embryos.

    PubMed

    Gao, Jing; Zhang, Wen; Dang, Wei; Mou, Yi; Gao, Yuan; Sun, Bao-Jun; Du, Wei-Guo

    2014-09-22

    The role of heat shock proteins (HSPs) in heat tolerance has been demonstrated in cultured cells and animal tissues, but rarely in whole organisms because of methodological difficulties associated with gene manipulation. By comparing HSP70 expression patterns among representative species of reptiles and birds, and by determining the effect of HSP70 overexpression on embryonic development and hatchling traits, we have identified the role of HSP70 in the heat tolerance of amniote embryos. Consistent with their thermal environment, and high incubation temperatures and heat tolerance, the embryos of birds have higher onset and maximum temperatures for induced HSP70 than do reptiles, and turtles have higher onset and maximum temperatures than do lizards. Interestingly, the trade-off between benefits and costs of HSP70 overexpression occurred between life-history stages: when turtle embryos developed at extreme high temperatures, HSP70 overexpression generated benefits by enhancing embryo heat tolerance and hatching success, but subsequently imposed costs by decreasing heat tolerance of surviving hatchlings. Taken together, the correlative and causal links between HSP70 and heat tolerance provide, to our knowledge, the first unequivocal evidence that HSP70 promotes thermal tolerance of embryos in oviparous amniotes.

  4. Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound

    PubMed Central

    Denbeigh, Janet M.; Nixon, Brian A.; Puri, Mira C.; Foster, F. Stuart

    2015-01-01

    Ultrasound contrast-enhanced imaging can convey essential quantitative information regarding tissue vascularity and perfusion and, in targeted applications, facilitate the detection and measure of vascular biomarkers at the molecular level. Within the mouse embryo, this noninvasive technique may be used to uncover basic mechanisms underlying vascular development in the early mouse circulatory system and in genetic models of cardiovascular disease. The mouse embryo also presents as an excellent model for studying the adhesion of microbubbles to angiogenic targets (including vascular endothelial growth factor receptor 2 (VEGFR2) or αvβ3) and for assessing the quantitative nature of molecular ultrasound. We therefore developed a method to introduce ultrasound contrast agents into the vasculature of living, isolated embryos. This allows freedom in terms of injection control and positioning, reproducibility of the imaging plane without obstruction and motion, and simplified image analysis and quantification. Late gestational stage (embryonic day (E)16.6 and E17.5) murine embryos were isolated from the uterus, gently exteriorized from the yolk sac and microbubble contrast agents were injected into veins accessible on the chorionic surface of the placental disc. Nonlinear contrast ultrasound imaging was then employed to collect a number of basic perfusion parameters (peak enhancement, wash-in rate and time to peak) and quantify targeted microbubble binding in an endoglin mouse model. We show the successful circulation of microbubbles within living embryos and the utility of this approach in characterizing embryonic vasculature and microbubble behavior. PMID:25867243

  5. Telomere lengths in human oocytes, cleavage stage embryos and blastocysts

    PubMed Central

    Turner, S.; Wong, H.P.; Rai, J.; Hartshorne, G.M.

    2010-01-01

    Telomeres are repeated sequences that protect the ends of chromosomes and harbour DNA repair proteins. Telomeres shorten during each cell division in the absence of telomerase. When telomere length becomes critically short, cell senescence occurs. Telomere length therefore reflects both cellular ageing and capacity for division. We have measured telomere length in human germinal vesicle (GV) oocytes and preimplantation embryos, by quantitative fluorescence in situ hybridization (Q-FISH), providing baseline data towards our hypothesis that telomere length is a marker of embryo quality. The numbers of fluorescent foci suggest that extensive clustering of telomeres occurs in mature GV stage oocytes, and in preimplantation embryos. When calculating average telomere length by assuming that each signal presents one telomere, the calculated telomere length decreased from the oocyte to the cleavage stages, and increased between the cleavage stages and the blastocyst (11.12 versus 8.43 versus 12.22 kb, respectively, P < 0.001). Other methods of calculation, based upon expected maximum and minimum numbers of telomeres, confirm that telomere length in blastocysts is significantly longer than cleavage stages. Individual blastomeres within an embryo showed substantial variation in calculated average telomere length. This study implies that telomere length changes according to the stage of preimplantation embryo development. PMID:20573647

  6. Hormetic effect induced by depleted uranium in zebrafish embryos.

    PubMed

    Ng, C Y P; Cheng, S H; Yu, K N

    2016-06-01

    The present work studied the hormetic effect induced by uranium (U) in embryos of zebrafish (Danio rerio) using apoptosis as the biological endpoint. Hormetic effect is characterized by biphasic dose-response relationships showing a low-dose stimulation and a high-dose inhibition. Embryos were dechorionated at 4h post fertilization (hpf), and were then exposed to 10 or 100μg/l depleted uranium (DU) in uranyl acetate solutions from 5 to 6 hpf. For exposures to 10μg/l DU, the amounts of apoptotic signals in the embryos were significantly increased at 20 hpf but were significantly decreased at 24 hpf, which demonstrated the presence of U-induced hormesis. For exposures to 100μg/l DU, the amounts of apoptotic signals in the embryos were significantly increased at 20, 24 and 30 hpf. Hormetic effect was not shown but its occurrence between 30 and 48 hpf could not be ruled out. In conclusion, hormetic effect could be induced in zebrafish embryos in a concentration- and time-dependent manner.

  7. Zebrafish (Danio rerio) embryos as a model for testing proteratogens.

    PubMed

    Weigt, Stefan; Huebler, Nicole; Strecker, Ruben; Braunbeck, Thomas; Broschard, Thomas H

    2011-03-15

    Zebrafish embryos have been shown to be a useful model for the detection of direct acting teratogens. This communication presents a protocol for a 3-day in vitro zebrafish embryo teratogenicity assay and describes results obtained for 10 proteratogens: 2-acetylaminofluorene, benzo[a]pyrene, aflatoxin B(1), carbamazepine, phenytoin, trimethadione, cyclophosphamide, ifosfamide, tegafur and thio-TEPA. The selection of the test substances accounts for differences in structure, origin, metabolism and water solubility. Apart from 2-acetylaminofluorene, which mainly produces lethal effects, all proteratogens tested were teratogenic in zebrafish embryos exposed for 3 days. The test substances and/or the substance class produced characteristic patterns of fingerprint endpoints. Several substances produced effects that could be identified already at 1 dpf (days post fertilization), whereas the effects of others could only be identified unambiguously after hatching at ≥ 3 dpf. The LC₅₀ and EC₅₀ values were used to calculate the teratogenicity index (TI) for the different substances, and the EC₂₀ values were related to human plasma concentrations. Results lead to the conclusion that zebrafish embryos are able to activate proteratogenic substances without addition of an exogenous metabolic activation system. Moreover, the teratogenic effects were observed at concentrations relevant to human exposure data. Along with other findings, our results indicate that zebrafish embryos are a useful alternative method for traditional teratogenicity testing with mammalian species.

  8. Biolistic techniques for transfection of mosquito embryos (Anopheles gambiae).

    PubMed

    Mialhe, E; Miller, L H

    1994-05-01

    To compensate for the extremely low rates of transformation by DNA microinjection into mosquito embryos of Anopheles gambiae, biolistic techniques were evaluated for introduction of DNA into large numbers of mosquito embryos. Biolistic experiments were first performed with a commercially available instrument intended for this purpose, according to the recommended procedure. The amount of DNA delivered was measured by the expression of luciferase under the control of the Drosophila heat shock protein (hsp) 70 promoter. Despite attempts to optimize biolistic parameters, the level of luciferase activity was low and highly variable. Two other methods of biolistic delivery of DNA-coated particles in aqueous suspension were then evaluated. One method used the gas explosion of the commercially available instrument (mentioned above) to drive an aqueous suspension of DNA-coated particles at high pressure. This method reproducibly increased the level of expression about 100-fold without greatly reducing embryo viability. Another method, which was recently described for plant transfection, uses lower pressure to deliver the aqueous suspension of DNA-coated particles. The level of expression of luciferase and the survival of embryos were equivalent to that obtained with the instrument modified for aqueous delivery of particles. Thus, both aqueous methods offer the advantages of reproducibly delivering more DNA to the embryos. Moreover, these methods could be suitable for delivering DNA mixed with proteins, such as restriction endonucleases and integrases, that may be destroyed by ethanol precipitation used in the standard PDS-1000/He method.

  9. Direct gene disruption by TALENs in medaka embryos.

    PubMed

    Wang, Tiansu; Hong, Yunhan

    2014-06-10

    Targeted gene disruption (GD) is powerful for generating genetic alterations in animal genomes. Engineered endonucleases such as zinc finger nucleases and transcription activator-like effector nucleases (TALENs) allow for GD directly in animal embryos to achieve germline transmission. Here we report procedures and parameters of TALEN-mediated GD in the fish medaka by using a germ cell-specific gene dnd as a model. Embryos at the 1-cell stage were microinjected with synthetic TALEN mRNAs and examined for the survival rate and GD efficiency. Medaka embryos can tolerate a high dosage of TALEN-mRNA injection and exhibit a steadily increasing GD efficiency with increasing mRNA dosages before peaking at 100 ng/μl. This dosage produced ~24% efficiency for somatic GD. Some of the animals from manipulated embryos developed into fertile female and male. Most importantly, four fish (3 males and 1 female) examined by progeny-test were able to produce GD-bearing male and female gametes for germline transmission to F1 generation at ~10% efficiency. Therefore, TALEN is proficient for somatic and germline GD in medaka embryos, and disruption of one dnd copy does not compromise somatic development and gamete production.

  10. Can artificial techniques supply morally neutral human embryos for research?

    PubMed

    Cheshire, William P; Jones, Nancy L

    2005-01-01

    Amidst controversy surrounding research on human embryos, biotechnology has conceived a substitute in the artificial human embryo. We examine the claim that novel embryos constructed artificially should be exempt from ethical restraints appropriate for research on embryos that come into being through natural processes. Morally relevant differences in intrinsic value depend on the sense in which the entity may be artificial, whether in regard to constituent matter, genetic or cellular form, generative means, or intended purpose. Considering each of these Aristotelian categories from a physicalist viewpoint, technology can achieve only limited degrees of artificiality because redesigned embryos still retain most of their natural features and relationships. From an essentialist viewpoint, the very limits of technology preclude the capability of manipulating the fundamental nature or essence of the individual who, even at the embryonic stage of life, cannot be made to be artificial through and through. A human may possess artificially contributed attributes but cannot be an artificial being. Classification of novel human organisms as artificial, therefore, is insufficient grounds by which to relinquish the principle that human moral status should be recognized for all living beings of human origin. In uncertain cases, at least the possibility of special human moral status should be considered present in organisms that are derived asexually, are developmentally defective, or are otherwise technologically altered.

  11. Contrast imaging in mouse embryos using high-frequency ultrasound.

    PubMed

    Denbeigh, Janet M; Nixon, Brian A; Puri, Mira C; Foster, F Stuart

    2015-03-04

    Ultrasound contrast-enhanced imaging can convey essential quantitative information regarding tissue vascularity and perfusion and, in targeted applications, facilitate the detection and measure of vascular biomarkers at the molecular level. Within the mouse embryo, this noninvasive technique may be used to uncover basic mechanisms underlying vascular development in the early mouse circulatory system and in genetic models of cardiovascular disease. The mouse embryo also presents as an excellent model for studying the adhesion of microbubbles to angiogenic targets (including vascular endothelial growth factor receptor 2 (VEGFR2) or αvβ3) and for assessing the quantitative nature of molecular ultrasound. We therefore developed a method to introduce ultrasound contrast agents into the vasculature of living, isolated embryos. This allows freedom in terms of injection control and positioning, reproducibility of the imaging plane without obstruction and motion, and simplified image analysis and quantification. Late gestational stage (embryonic day (E)16.6 and E17.5) murine embryos were isolated from the uterus, gently exteriorized from the yolk sac and microbubble contrast agents were injected into veins accessible on the chorionic surface of the placental disc. Nonlinear contrast ultrasound imaging was then employed to collect a number of basic perfusion parameters (peak enhancement, wash-in rate and time to peak) and quantify targeted microbubble binding in an endoglin mouse model. We show the successful circulation of microbubbles within living embryos and the utility of this approach in characterizing embryonic vasculature and microbubble behavior.

  12. Plant regeneration from somatic embryos ofTaxus brevifolia.

    PubMed

    Chee, P P

    1996-12-01

    Taxusbrevifolia is the source of paclitaxel (Taxol®), an anticancer drug. A method for regeneration ofTaxus brevifolia from immature zygotic embryos via somatic embryogenesis is described. Embryogenic callus tissues were obtained by culturing immature zygotic embryos on Lloyd and McCown medium (MCM) supplemented with 160 μM 2,4-dichlorophenoxyacetic acid (2,4-D) + 5 μM benzylaminopurine (BA) + 5 μM naphthaleneacetic acid (NAA) for 4 weeks. Putative embryoids were obtained following transfer of cultures to MCM medium supplemented with 4 μM BA + 5 μM kinetin + 1 μM NAA for 6 to 8 weeks. Conversion of embryos was obtained on MCM medium supplemented with 40 μM abscisic acid (ABA) + 1% activated charcoal. Development of bipolar structures with recognizable shoot and root apices was observed in somatic embryos. Five percent of somatic embryos were regenerated into plantlets on half-strength growth regulator-free MCM medium.

  13. Current status and future direction of cryopreservation of camelid embryos.

    PubMed

    Herrid, M; Vajta, G; Skidmore, J A

    2017-02-01

    Over the past 3 decades, and similar to the horse industry, fresh embryo transfer has been widely practiced on large commercial scales in different camelid species, especially the dromedary camel and alpaca. However, the inability to cryopreserve embryos significantly reduces its broader application, and as such limits the capacity to utilize elite genetic resources internationally. In addition, cryopreservation of the semen of camelids is also difficult, suggesting an extreme sensitivity of the germplasm to cooling and freezing. As a result, genetic resources of camelids must continue to be maintained as living collections of animals. Due to concerns over disease outbreaks such as that of the highly pathogenic Middle East Respiratory Syndrome in the Middle East and Asia, there is an urgent need to establish an effective gene banking system for camelid species, especially the camel. The current review compares and summarizes recent progress in the field of camelid embryo cryopreservation, identifying four possible reasons for the slow development of an effective protocol and describing eight future directions to improve the current protocols. At the same time, the results of a recent dromedary camel embryo transfer study which produced a high morphologic integrity and survival rate of Open Pulled Straw-vitrified embryos are also discussed.

  14. Modeling embryo transfer into a closed uterine cavity.

    PubMed

    Yaniv, Sarit; Jaffa, Ariel J; Elad, David

    2012-11-01

    Embryo transfer (ET) is the last manual intervention after extracorporeal fertilization. After the ET procedure is completed, the embryos are conveyed in the uterus for another two to four days due to spontaneous uterine peristalsis until the window time for implantation. The role of intrauterine fluid flow patterns in transporting the embryos to their implantation site during and after ET was simulated by injection of a liquid bolus into a two-dimensional liquid-filled channel with a closed fundal end via a liquid-filled catheter inserted in the channel. Numerical experiments revealed that the intrauterine fluid field and the embryos transport pattern were strongly affected by the closed fundal end. The embryos re-circulated in small loops around the vicinity where they were deposited from the catheter. The transport pattern was controlled by the uterine peristalsis factors, such as amplitude and frequency of the uterine walls motility, as well as the synchronization between the onset of catheter discharge and uterine peristalsis. The outcome of ET was also dependent on operating parameters such as placement of the catheter tip within the uterine cavity and the delivery speed of the catheter load. In conclusion, this modeling study highlighted important parameters that should be considered during ET procedures in order to increase the potential for pregnancy success.

  15. Respiration and Mitochondrial Biogenesis in Germinating Embryos of Maize 1

    PubMed Central

    Ehrenshaft, Marilyn; Brambl, Robert

    1990-01-01

    Function of the cyanide-sensitive mitochondrial electron transport system was required for germination of the Zea mays embryo. Respiration of the standard electron transport system (rather than the alternate oxidase) began immediately upon initiation of imbibition. This respiration depended upon cytochrome c oxidase and ATPase that were conserved in an active form in the quiescent embryo rather than upon newly synthesized or assembled enzyme complexes. Immunoprecipitation of radiolabeled subunits of these enzymes showed that the initiation of mitochondrial biogenetic activities, including de novo synthesis of nuclear- and mitochondrial-encoded enzyme subunit peptides, was strongly induced after 6 hours of embryo germination. Undetectable or very low levels of transcripts for subunits 1 and 2 of the F1-ATPase and subunit 2 of cytochrome c oxidase were present in the quiescent embryo; these transcripts accumulated rapidly between 6 and 12 hours of germination and their translation products were rapidly synthesized between 6 and 24 hours. An exception was the gene for subunit 9 of the ATPase; transcripts of this mitochondrial gene were abundant in the dry embryo and rapidly accumulated further upon initiation of imbibition; they were translated actively during the first 6 hours. We isolated and sequenced a near full-length cDNA for subunit 2 (beta) of the F1-ATPase, and we compared the deduced protein sequence with related sequences of other organisms. Images Figure 2 Figure 3 Figure 5 PMID:16667450

  16. Use of "excess" human embryos for stem cell research: protecting women's rights and health.

    PubMed

    Cohen, C B

    2000-01-01

    Proposed National Institutes of Health guidelines for stem cell research are too narrowly drawn and do not adequately protect the freedom of choice and health of women who donate embryos. They need to be expanded to cover not only the point of embryo donation, but also that of embryo creation. Guidelines are provided to ensure that donors undergoing hyperstimulation and egg retrieval gave voluntary informed consent to the production of embryos that might later prove in excess. A standard for determining when embryos have been overproduced is presented to address the possibility that additional embryos will be created for stem cell research in violation of the guidelines and at risk to women's health.

  17. Dual Positive Regulation of Embryo Implantation by Endocrine and Immune Systems--Step-by-Step Maternal Recognition of the Developing Embryo.

    PubMed

    Fujiwara, Hiroshi; Araki, Yoshihiko; Imakawa, Kazuhiko; Saito, Shigeru; Daikoku, Takiko; Shigeta, Minoru; Kanzaki, Hideharu; Mori, Takahide

    2016-03-01

    In humans, HCG secreted from the implanting embryo stimulates progesterone production of the corpus luteum to maintain embryo implantation. Along with this endocrine system, current evidence suggests that the maternal immune system positively contributes to the embryo implantation. In mice, immune cells that have been sensitized with seminal fluid and then the developing embryo induce endometrial differentiation and promote embryo implantation. After hatching, HCG activates regulatory T and B cells through LH/HCG receptors and then stimulates uterine NK cells and monocytes through sugar chain receptors, to promote and maintain pregnancy. In accordance with the above, the intrauterine administration of HCG-treated PBMC was demonstrated to improve implantation rates in women with repeated implantation failures. These findings suggest that the maternal immune system undergoes functional changes by recognizing the developing embryos in a stepwise manner even from a pre-fertilization stage and facilitates embryo implantation in cooperation with the endocrine system.

  18. Beneficial effect of two culture systems with small groups of embryos on the development and quality of in vitro-produced bovine embryos.

    PubMed

    Cebrian-Serrano, A; Salvador, I; Silvestre, M A

    2014-02-01

    Currently, in vitro-produced embryos derived by ovum pick up (OPU) and in vitro fertilization (IVF) technologies represent approximately one-third of the embryos worldwide in cattle. Nevertheless, the culture of small groups of embryos from an individual egg donor is an issue that OPU-IVF laboratories have to face. In this work, we tested whether the development and quality of the preimplantation embryos in vitro cultured in low numbers (five embryos) could be improved by the addition of epidermal growth factor, insulin, transferrin and selenium (EGF-ITS) or by the WOW system. With this aim, immature oocytes recovered from slaughtered heifers were in vitro matured and in vitro fertilized. Presumptive zygotes were then randomly cultured in four culture conditions: one large group (LG) (50 embryos/500 μl medium) and three smaller groups [five embryos/50 μl medium without (control) or with EGF-ITS (EGF-ITS) and five embryos per microwell in the WOW system (WOW)]. Embryos cultured in LG showed a greater ability to develop to blastocyst stage than embryos cultured in smaller groups, while the blastocyst rate of WOW group was significantly higher than in control. The number of cells/blastocyst in LG was higher than control or WOW, whereas the apoptosis rate per blastocyst was lower. On the other hand, the addition of EGF-ITS significantly improved both parameters compared to the control and resulted in similar embryo quality to LG. In conclusion, the WOW system improved embryo development, while the addition of EGF-ITS improved the embryo quality when smaller groups of embryos were cultured.

  19. Whole mount nuclear fluorescent imaging: convenient documentation of embryo morphology.

    PubMed

    Sandell, Lisa L; Kurosaka, Hiroshi; Trainor, Paul A

    2012-11-01

    Here, we describe a relatively inexpensive and easy method to produce high quality images that reveal fine topological details of vertebrate embryonic structures. The method relies on nuclear staining of whole mount embryos in combination with confocal microscopy or conventional wide field fluorescent microscopy. In cases where confocal microscopy is used in combination with whole mount nuclear staining, the resulting embryo images can rival the clarity and resolution of images produced by scanning electron microscopy (SEM). The fluorescent nuclear staining may be performed with a variety of cell permeable nuclear dyes, enabling the technique to be performed with multiple standard microscope/illumination or confocal/laser systems. The method may be used to document morphology of embryos of a variety of organisms, as well as individual organs and tissues. Nuclear stain imaging imposes minimal impact on embryonic specimens, enabling imaged specimens to be utilized for additional assays.

  20. The role of auxin signaling in early embryo pattern formation.

    PubMed

    Smit, Margot E; Weijers, Dolf

    2015-12-01

    Pattern formation of the early Arabidopsis embryo generates precursors to all major cell types, and is profoundly controlled by the signaling molecule auxin. Here we discuss recent milestones in our understanding of auxin-dependent embryo patterning. Auxin biosynthesis, transport and response mechanisms interact to generate local auxin accumulation in the early embryo. New auxin-dependent reporters help identifying these sites, while atomic structures of transcriptional response mediators help explain the diverse outputs of auxin signaling. Key auxin outputs are control of cell identity and cell division orientation, and progress has been made towards understanding the cellular basis of each. Importantly, a number of studies have combined computational modeling and experiments to analyze the developmental role, genetic circuitry and molecular mechanisms of auxin-dependent cell division control.

  1. Expression of SRY transcripts in preimplantation human embryos

    SciTech Connect

    Fiddler, M.; Abdel-Rahman, B.; Rappolee, D.A.

    1995-01-02

    We have examined the expression of SRY mRNA in individual in vitro fertilized preimplantation human embryos; because of ethical constraints, these studies were confined to embryos with one and three pronuclei. Using a sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) assay, we observed SRY mRNA at the one-cell through the blastula stages but not in spermatoza. These results indicate that the de novo transcription of this sex-specific gene occurs at a developmental time considerably earlier than that of gonadal differentiation. Our results also indicate that in vitro fertilized embryos with one pronucleus are likely to be diploid. 39 refs., 1 fig., 2 tabs.

  2. Cadmium induces an apoptotic response in sea urchin embryos

    PubMed Central

    Agnello, Maria; Filosto, Simone; Scudiero, Rosaria; Rinaldi, Anna M.; Roccheri, Maria C.

    2007-01-01

    Cadmium is a heavy metal toxic for living organisms even at low concentrations. It does not have any biological role, and since it is a permanent metal ion, it is accumulated by many organisms. In the present paper we have studied the apoptotic effects of continuous exposure to subacute/sublethal cadmium concentrations on a model system: Paracentrotus lividus embryos. We demonstrated, by atomic absorption spectrometry, that the intracellular amount of metal increased during exposure time. We found, using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, that long treatments with cadmium triggered a severe DNA fragmentation. We demonstrated, by immunocytochemistry on whole-mount embryos, that treatment with cadmium causes activation of caspase-3 and cleavage of death substrates α-fodrin and lamin A. Incubating the embryos since fertilization with Z-DEVD FMK, a caspase-3 inhibitor, we found, by immunocytochemistry, that cleavage by caspase-3 and cleavage of death substrates were inactivated. PMID:17441506

  3. Rayleigh instability of the inverted one-cell amphibian embryo

    NASA Astrophysics Data System (ADS)

    Nouri, Comron; Luppes, Roel; Veldman, Arthur E. P.; Tuszynski, Jack A.; Gordon, Richard

    2008-03-01

    The one-cell amphibian embryo is modeled as a rigid spherical shell containing equal volumes of two immiscible fluids with different densities and viscosities and a surface tension between them. The fluids represent denser yolk in the bottom hemisphere and clearer cytoplasm and the germinal vesicle in the top hemisphere. The unstable equilibrium configuration of the inverted system (the heavier fluid on top) depends on the value of the contact angle. The theoretically calculated normal modes of perturbation and the instability of each mode are in agreement with the results from ComFlo computational fluid dynamic simulations of the same system. The two dominant types of modes of perturbation give rise to axisymmetric and asymmetric sloshing of the cytoplasm of the inverted embryos, respectively. This work quantifies our hypothesis that the axisymmetric mode corresponds to failure of development, and the asymmetric sloshing mode corresponds to development proceeding normally, but with reversed pigmentation, for inverted embryos.

  4. Determinant molecular markers for peri-gastrulating bovine embryo development.

    PubMed

    Hue, Isabelle

    2016-01-01

    Peri-gastrulation defines the time frame between blastocyst formation and implantation that also corresponds in cattle to elongation, pregnancy recognition and uterine secretion. Optimally, this developmental window prepares the conceptus for implantation, placenta formation and fetal development. However, this is a highly sensitive period, as evidenced by the incidence of embryo loss or early post-implantation mortality after AI, embryo transfer or somatic cell nuclear transfer. Elongation markers have often been used within this time frame to assess developmental defects or delays, originating either from the embryo, the uterus or the dam. Comparatively, gastrulation markers have not received great attention, although elongation and gastrulation are linked by reciprocal interactions at the molecular and cellular levels. To make this clearer, this peri-gastrulating period is described herein with a focus on its main developmental landmarks, and the resilience of the landmarks in the face of biotechnologies is questioned.

  5. Cumulus and granulosa cell markers of oocyte and embryo quality

    PubMed Central

    Uyar, Asli; Torrealday, Saioa; Seli, Emre

    2013-01-01

    Lack of an objective, accurate, and noninvasive embryo assessment strategy remains one of the major challenges encountered in in vitro fertilization. Cumulus and mural granulosa cells reflect the characteristics of the oocyte, providing a noninvasive means to assess oocyte quality. Specifically, transcriptomic profiling of follicular cells may help identify biomarkers of oocyte and embryo competence. Current transcriptomics technologies include quantitative reverse transcriptase–polymerase chain reaction (qRT-PCR) for analysis of individual genes and microarrays and high-throughput deep sequencing for whole genome expression profiling. Recently, using qRT-PCR and microarray technologies, a multitude of studies correlated changes in cumulus or granulosa cell gene expression with clinically relevant outcome parameters, including in vitro embryo development and pregnancy. While the initial findings are promising, a clinical benefit from the use of identified biomarker genes remains to be demonstrated in randomized controlled trials. PMID:23498999

  6. Characterization of starch-debranching enzymes in pea embryos

    PubMed

    Zhu; Hylton; Rossner; Smith

    1998-10-01

    Two distinct types of debranching enzymes have been identified in developing pea (Pisum sativum L.) embryos using native gel analysis and tests of substrate preference on purified or partially purified activities. An isoamylase-like activity capable of hydrolyzing amylopectin and glycogen but not pullulan is present throughout development and is largely or entirely confined to the plastid. Activities capable of hydrolyzing pullulan are present both inside and outside of the plastid, and extraplastidial activity increases relative to the plastidial activity during development. Both types of debranching enzyme are also present in germinating embryos. We argue that debranching enzymes are likely to have a role in starch metabolism in the plastid of the developing embryo and in starch degradation during germination.

  7. Mitotic wavefronts mediated by mechanical signaling in early Drosophila embryos

    NASA Astrophysics Data System (ADS)

    Kang, Louis; Idema, Timon; Liu, Andrea; Lubensky, Tom

    2013-03-01

    Mitosis in the early Drosophila embryo demonstrates spatial and temporal correlations in the form of wavefronts that travel across the embryo in each cell cycle. This coordinated phenomenon requires a signaling mechanism, which we suggest is mechanical in origin. We have constructed a theoretical model that supports nonlinear wavefront propagation in a mechanically-excitable medium. Previously, we have shown that this model captures quantitatively the wavefront speed as it varies with cell cycle number, for reasonable values of the elastic moduli and damping coefficient of the medium. Now we show that our model also captures the displacements of cell nuclei in the embryo in response to the traveling wavefront. This new result further supports that mechanical signaling may play an important role in mediating mitotic wavefronts.

  8. Wavefronts and mechanical signaling in early Drosophila embryos

    NASA Astrophysics Data System (ADS)

    Idema, Timon; Dubuis, Julien; Manning, Lisa; Nelson, Philip; Liu, Andrea

    2012-02-01

    Mitosis in the early syncytial Drosophila embryo has a high degree of spatial and temporal correlations, visible as mitotic wavefronts that travel across the embryo. This mitosis wavefront is preceded by another wavefront which corresponds to chromosome condensation. The two wavefronts are separated by a time interval that is independent of cell cycle and propagate at the same speed for a given embryo in a given cycle. We study the wavefronts in the context of excitable medium theory, using two different models, one with biochemical signaling and one with mechanical signaling. We find that the dependence of wavefront speed on cell cycle number is most naturally explained via a mechanical signaling, and that the entire process suggests a scenario in which biochemical and mechanical signaling are coupled.

  9. Ex vivo recovery of preimplantatory embryos in bitches.

    PubMed

    Commin, L; Buff, S; Rosset, E; Joly, T; Guerin, P; Neto, V

    2012-12-01

    The present study was conducted to investigate the timing of preimplantatory development in the dog and to evaluate the efficiency of flushing oviducts and uterine horns to collect embryos. Among the embryonic structures collected between day 8 and day 12 after ovulation, 43 % were at the 1-16 cells stage, 23% were at the morula stage and 34% at the blastocyst stage. Our collection method yielded to a recovery rate of 61.3 %, and 7.1 ± 0.7 embryos were harvested per bitch. In addition, the ovulation rate reached 11.6 ± 0.8 per bitch. The first morulae were observed from day 9 post-ovulation, while the first blastocyst appeared from day 10. Two-thirds of the collected morulae-blastocysts were obtained between the 11th and the 12th day after ovulation. To the moment, we suggest this is the best period to harvest canine embryo for cryopreservation.

  10. Characterization of Starch-Debranching Enzymes in Pea Embryos1

    PubMed Central

    Zhu, Zhi-Ping; Hylton, Christopher M.; Rössner, Ute; Smith, Alison M.

    1998-01-01

    Two distinct types of debranching enzymes have been identified in developing pea (Pisum sativum L.) embryos using native gel analysis and tests of substrate preference on purified or partially purified activities. An isoamylase-like activity capable of hydrolyzing amylopectin and glycogen but not pullulan is present throughout development and is largely or entirely confined to the plastid. Activities capable of hydrolyzing pullulan are present both inside and outside of the plastid, and extraplastidial activity increases relative to the plastidial activity during development. Both types of debranching enzyme are also present in germinating embryos. We argue that debranching enzymes are likely to have a role in starch metabolism in the plastid of the developing embryo and in starch degradation during germination. PMID:9765544

  11. Temperature and photoperiod responses of soybean embryos cultured in vitro

    NASA Technical Reports Server (NTRS)

    Raper, C. D. Jr; Patterson, R. P.; Raper CD, J. r. (Principal Investigator)

    1986-01-01

    Temperature and photoperiod each have direct effects on growth rate of excised embryos of soybean (Glycine max (L.) Merrill). To determine if the effects of photoperiod are altered by temperature, embryos of 'Ransom II' were cultured in vitro at 18, 24, and 30 degrees C under photoperiod durations of 12 and 18 h at an irradiance of 9 W m-2 (700 to 850 nm) and a photosynthetic photon flux density of 58 micromoles m-2 s-1 (400 to 700 nm). Accumulation rates of fresh and dry weight were greater under 18-h than 12-h photoperiods over the entire range of temperature. Water content of the culture embryos was not affected by photoperiod but was greater at 18 and 30 than 24 degrees C. The accumulation rate of dry weight increased from 18 to 26 but declined at 30 degrees C.

  12. Embryo research--why the Cardinal is wrong.

    PubMed Central

    Walton

    1990-01-01

    Reasons are given for suggesting that individuation of the human embryo does not begin until the primitive streak forms at about the fourteenth day after conception; this view, though contested by many, is held by very many committed Christians of all denominations. In the conceptus or pre-embryo, after the formation of a blastocyst at about four-five days after fertilisation, biopsy of a single cell from the outer layer of cells (which later can form the membranes and placenta) can be used to determine the sex of the conceptus and will ultimately be used to detect the presence of an abnormal gene such as that for Duchenne-type muscular dystrophy, without detriment to development of the basal cell mass from which the embryo forms. The potential benefits in the prevention of inherited disease are profound. PMID:2287013

  13. Embryo research--why the Cardinal is wrong.

    PubMed

    Walton

    1990-12-01

    Reasons are given for suggesting that individuation of the human embryo does not begin until the primitive streak forms at about the fourteenth day after conception; this view, though contested by many, is held by very many committed Christians of all denominations. In the conceptus or pre-embryo, after the formation of a blastocyst at about four-five days after fertilisation, biopsy of a single cell from the outer layer of cells (which later can form the membranes and placenta) can be used to determine the sex of the conceptus and will ultimately be used to detect the presence of an abnormal gene such as that for Duchenne-type muscular dystrophy, without detriment to development of the basal cell mass from which the embryo forms. The potential benefits in the prevention of inherited disease are profound.

  14. Developmental effects of exposing Drosophila embryos to ether vapour.

    PubMed

    Bownes, M; Seiler, M

    1977-01-01

    Drosophila embryos at precise developmental stages were exposed to ether vapour. The defects in the resulting embryos and adults were observed. Ether disrupted embroygenesis in specific ways, causing defects primarily at the anterior of the embryo and disorganizing the arrangement of the segments. Adults showed deficiencies and duplications of many imaginal disc and histoblast derivatives. Phenocopies of the bithorax mutation which transforms metathorax to mesothorax were observed. They were first induced at the syncytial blastoderm stage, had their peak of production at the cellular blastoderm, and were no longer observed after the anterior and posterior midgut were partially invaginated. It was observed that not only are the halter/wing transformations confined to the anterior compartment, but also leg 3 to leg 2 transformations only occurred in the anterior leg compartment.

  15. Embryo-scale tissue mechanics during Drosophila gastrulation movements

    PubMed Central

    Rauzi, Matteo; Krzic, Uros; Saunders, Timothy E.; Krajnc, Matej; Ziherl, Primož; Hufnagel, Lars; Leptin, Maria

    2015-01-01

    Morphogenesis of an organism requires the development of its parts to be coordinated in time and space. While past studies concentrated on defined cell populations, a synthetic view of the coordination of these events in a whole organism is needed for a full understanding. Drosophila gastrulation begins with the embryo forming a ventral furrow, which is eventually internalized. It is not understood how the rest of the embryo participates in this process. Here we use multiview selective plane illumination microscopy coupled with infrared laser manipulation and mutant analysis to dissect embryo-scale cell interactions during early gastrulation. Lateral cells have a denser medial–apical actomyosin network and shift ventrally as a compact cohort, whereas dorsal cells become stretched. We show that the behaviour of these cells affects furrow internalization. A computational model predicts different mechanical properties associated with tissue behaviour: lateral cells are stiff, whereas dorsal cells are soft. Experimental analysis confirms these properties in vivo. PMID:26497898

  16. Phenotype classification of zebrafish embryos by supervised learning.

    PubMed

    Jeanray, Nathalie; Marée, Raphaël; Pruvot, Benoist; Stern, Olivier; Geurts, Pierre; Wehenkel, Louis; Muller, Marc

    2015-01-01

    Zebrafish is increasingly used to assess biological properties of chemical substances and thus is becoming a specific tool for toxicological and pharmacological studies. The effects of chemical substances on embryo survival and development are generally evaluated manually through microscopic observation by an expert and documented by several typical photographs. Here, we present a methodology to automatically classify brightfield images of wildtype zebrafish embryos according to their defects by using an image analysis approach based on supervised machine learning. We show that, compared to manual classification, automatic classification results in 90 to 100% agreement with consensus voting of biological experts in nine out of eleven considered defects in 3 days old zebrafish larvae. Automation of the analysis and classification of zebrafish embryo pictures reduces the workload and time required for the biological expert and increases the reproducibility and objectivity of this classification.

  17. Effect of the afterloaded external guidance embryo transfer technique on pregnancy rates in single embryo transfer cycles

    PubMed Central

    Yılmaz, Nafiye; Oruç, Ayla Sargın; Zeyrek, Tugba; Görkem, Ümit; İnal, Hasan Ali; Engin-Üstün, Yaprak; Gülerman, Cavidan

    2013-01-01

    Objective To investigate effect of the afterloaded external guidance embryo transfer technique on pregnancy rates in single embryo transfer intracytoplasmic sperm injection (ICSI) cycles. Material and Methods This retrospective study was performed at the Dr. Zekai Tahir Burak Women’s Health Research and Education Hospital. Three hundred and thirteen women who underwent ICSI were included in the study. Subjects were categorized according to the embryo transfer technique; Group 1 (n: 232): easy transfer with a soft catheter, Group 2 (n: 45): after external guidance transfer, and Group 3 (n: 36): difficult transfer with a stylet. Basal parameters, clinical and laboratory IVF outcomes and pregnancy rates were studied. Results Infertility etiology, basal follicle stimulating hormone (FSH) levels, antral follicle count, duration of stimulation, total dose of gonadotropin, peak estradiol levels, endometrial thickness, oocyte number, 2 PN, and fertilization rate were similar between the three groups (p>0.05). Despite the decreased pregnancy rate in Group 3, there were no differences in clinical pregnancy rates among the groups (p=0.204). Conclusion Embryo transfer is one of the critical steps in assisted reproduction procedures. Using the afterloaded external guidance embryo transfer technique did not improve pregnancy rates. PMID:24592095

  18. Cotyledonary somatic embryos of Pinus pinaster Ait. most closely resemble fresh, maturing cotyledonary zygotic embryos: biological, carbohydrate and proteomic analyses.

    PubMed

    Morel, Alexandre; Trontin, Jean-François; Corbineau, Françoise; Lomenech, Anne-Marie; Beaufour, Martine; Reymond, Isabelle; Le Metté, Claire; Ader, Kevin; Harvengt, Luc; Cadene, Martine; Label, Philippe; Teyssier, Caroline; Lelu-Walter, Marie-Anne

    2014-11-01

    Cotyledonary somatic embryos (SEs) of maritime pine are routinely matured for 12 weeks before being germinated and converted to plantlets. Although regeneration success is highly dependent on SEs quality, the date of harvesting is currently determined mainly on the basis of morphological features. This empirical method does not provide any accurate information about embryo quality with respect to storage compounds (proteins, carbohydrates). We first analyzed SEs matured for 10, 12 and 14 weeks by carrying out biological (dry weight, water content) and biochemical measurements (total protein and carbohydrate contents). No difference could be found between collection dates, suggesting that harvesting SEs after 12 weeks is appropriate. Cotyledonary SEs were then compared to various stages, from fresh to fully desiccated, in the development of cotyledonary zygotic embryos (ZEs). We identified profiles that were similar using hierarchical ascendant cluster analysis (HCA). Fresh and dehydrated ZEs could be distinguished, and SEs clustered with fresh ZEs. Both types of embryo exhibited similar carbohydrate and protein contents and signatures. This high level of similarity (94.5 %) was further supported by proteome profiling. Highly expressed proteins included storage, stress-related, late embryogenesis abundant and energy metabolism proteins. By comparing overexpressed proteins in developing and cotyledonary SEs or ZEs, some (23 proteins) could be identified as candidate biomarkers for the late, cotyledonary stage. This is the first report of useful generic protein markers for monitoring embryo development in maritime pine. Our results also suggest that improvements of SEs quality may be achieved if the current maturation conditions are refined.

  19. Equine cloning: in vitro and in vivo development of aggregated embryos.

    PubMed

    Gambini, Andrés; Jarazo, Javier; Olivera, Ramiro; Salamone, Daniel F

    2012-07-01

    The production of cloned equine embryos remains highly inefficient. Embryo aggregation has not yet been tested in the equine, and it might represent an interesting strategy to improve embryo development. This study evaluated the effect of cloned embryo aggregation on in vitro and in vivo equine embryo development. Zona-free reconstructed embryos were individually cultured in microwells (nonaggregated group) or as 2- or 3-embryo aggregates (aggregated groups). For in vitro development, they were cultured until blastocyst stage and then either fixed for Oct-4 immunocytochemical staining or maintained in in vitro culture where blastocyst expansion was measured daily until Day 17 or the day on which they collapsed. For in vivo assays, Day 7-8 blastocysts were transferred to synchronized mares and resultant vesicles, and cloned embryos were measured by ultrasonography. Embryo aggregation improved blastocyst rates on a per well basis, and aggregation did not imply additional oocytes to obtain blastocysts. Embryo aggregation improved embryo quality, nevertheless it did not affect Day 8 and Day 16 blastocyst Oct-4 expression patterns. Equine cloned blastocysts expanded and increased their cell numbers when they were maintained in in vitro culture, describing a particular pattern of embryo growth that was unexpectedly independent of embryo aggregation, as all embryos reached similar size after Day 7. Early pregnancy rates were higher using blastocysts derived from aggregated embryos, and advanced pregnancies as live healthy foals also resulted from aggregated embryos. These results indicate that the strategy of aggregating embryos can improve their development, supporting the establishment of equine cloned pregnancies.

  20. Identification of bovine embryos cultured in groups by attachment of barcodes to the zona pellucida.

    PubMed

    Novo, Sergi; Morató, Roser; Penon, Oriol; Duran, Sara; Barrios, Leonardo; Nogués, Carme; Plaza, José Antonio; Pérez-García, Luisa; Mogas, Teresa; Ibáñez, Elena

    2014-06-01

    The low number of oocytes collected from unstimulated donors by ovum pick-up means that embryos produced from each individual female have to be cultured individually or in very small groups. However, it has been demonstrated that single-embryo culture is less efficient than embryo culture in groups. To overcome this limitation, we developed a direct embryo-tagging system, which allows the collective culture of embryos from different origins whilst preserving their pedigree. Presumptive bovine zygotes were tagged with eight wheat-germ agglutinin biofunctionalised polysilicon barcodes attached to the outer surface of the zona pellucida (ZP). Four different barcodes were used to encode groups of 20-25 embryos, which were then cultured in the same drop. Cleavage, Day-7 and Day-8 blastocysts and barcode retention rates were assessed. In addition, Day-7 blastocysts were vitrified and warmed. Barcode attachment to the ZP of bovine embryos affected neither in vitro embryo development nor post-warming survival of the tagged embryos. All the embryos maintained barcodes attached until Day 8 of culture (3.63±0.37 barcodes per embryo) and could be identified. In conclusion, identification of embryos by barcodes attached to the ZP is feasible and will allow the culture of embryos from different donors in the same drop.

  1. Automated Zebrafish Chorion Removal and Single Embryo Placement: Optimizing Throughput of Zebrafish Developmental Toxicity Screens

    PubMed Central

    Mandrell, David; Truong, Lisa; Jephson, Caleb; Sarker, Mushfiqur R.; Moore, Aaron; Lang, Christopher; Simonich, Michael T.; Tanguay, Robert L.

    2012-01-01

    The potential of the developing zebrafish model for toxicology and drug discovery is limited by inefficient approaches to manipulating and chemically exposing zebrafish embryos—namely, manual placement of embryos into 96- or 384-well plates and exposure of embryos while still in the chorion, a barrier of poorly characterized permeability enclosing the developing embryo. We report the automated dechorionation of 1600 embryos at once at 4 h postfertilization (hpf) and placement of the dechorionated embryos into 96-well plates for exposure by 6 hpf. The process removed ≥95% of the embryos from their chorions with 2% embryo mortality by 24 hpf, and 2% of the embryos malformed at 120 hpf. The robotic embryo placement allocated 6-hpf embryos to 94.7% ± 4.2% of the wells in multiple 96-well trials. The rate of embryo mortality was 2.8% (43 of 1536) from robotic handling, the rate of missed wells was 1.2% (18 of 1536), and the frequency of multipicks was <0.1%. Embryo malformations observed at 24 hpf occurred nearly twice as frequently from robotic handling (16 of 864; 1.9%) as from manual pipetting (9 of 864; 1%). There was no statistical difference between the success of performing the embryo placement robotically or manually. PMID:22357610

  2. Acorns containing deeper plumule survive better: how white oaks counter embryo excision by rodents.

    PubMed

    Zhang, Mingming; Dong, Zhong; Yi, Xianfeng; Bartlow, Andrew W

    2014-01-01

    Several squirrel species excise the embryo of acorns of most white oak species to arrest germination for long-term storage. However, it is not clear how these acorns counter embryo excision and survive in the arms race of coevolution. In this study, we simulated the embryo excision behavior of squirrels by removing 4 mm of cotyledon from the apical end of white oak acorns differing in embryo depths to investigate the effects of embryo excision on acorn germination and seedling performance of white oak species. The embryo depth in the cotyledons was significantly different among white oak acorns, with Quercus mongolica containing the embryo most deeply in the acorns. We found that artificial embryo excision significantly decreased acorn germination rates of Quercus variabilis, Quercus acutissima, Quercus aliena, Quercus aliena var. acutiserrata, Quercus serrata. var. brevipetiolata but not Q. mongolica. Artificial embryo excision exerted significant negative impacts on seedling performance of all oak species except Quercus aliena. Our study demonstrates the role of embryo depth of acorns in countering embryo excision by squirrels and may explain the fact that squirrels do not perform embryo excision in acorns of Q. mongolica with deeper embryos. This apparent adaptation of acorns sheds light on the coevolutionary dynamics between oaks and their seed predators.

  3. Mitochondrial and DNA damage in bovine somatic cell nuclear transfer embryos.

    PubMed

    Hwang, In-Sun; Bae, Hyo-Kyung; Cheong, Hee-Tae

    2013-01-01

    The generation of reactive oxygen species (ROS) and subsequent mitochondrial and DNA damage in bovine somatic cell nuclear transfer (SCNT) embryos were examined. Bovine enucleated oocytes were electrofused with donor cells and then activated by a combination of Ca-ionophore and 6-dimethylaminopurine culture. The H2O2 and ˙OH radical levels, mitochondrial morphology and membrane potential (ΔΨ), and DNA fragmentation of SCNT and in vitro fertilized (IVF) embryos at the zygote stage were analyzed. The H2O2 (35.6 ± 1.1 pixels/embryo) and ˙OH radical levels (44.6 ± 1.2 pixels/embryo) of SCNT embryos were significantly higher than those of IVF embryos (19.2 ± 1.5 and 23.8 ± 1.8 pixels/embryo, respectively, p < 0.05). The mitochondria morphology of SCNT embryos was diffused within the cytoplasm. The ΔΨ of SCNT embryos was significantly lower (p < 0.05) than that of IVF embryos (0.95 ± 0.04 vs. 1.21 ± 0.06, red/green). Moreover, the comet tail length of SCNT embryos was longer than that of IVF embryos (515.5 ± 26.4 μm vs. 425.6 ± 25.0 μm, p < 0.05). These results indicate that mitochondrial and DNA damage increased in bovine SCNT embryos, which may have been induced by increased ROS levels.

  4. Effect of donor cell type on nuclear remodelling in rabbit somatic cell nuclear transfer embryos.

    PubMed

    Tian, J; Song, J; Li, H; Yang, D; Li, X; Ouyang, H; Lai, L

    2012-08-01

    Cloned rabbits have been produced for many years by somatic cell nuclear transfer (SCNT). The efficiency of cloning by SCNT, however, has remained extremely low. Most cloned embryos degenerate in utero, and the few that develop to term show a high incidence of post-natal death and abnormalities. The cell type used for donor nuclei is an important factor in nuclear transfer (NT). As reported previously, NT embryos reconstructed with fresh cumulus cells (CC-embryos) have better developmental potential than those reconstructed with foetal fibroblasts (FF-embryos) in vivo and in vitro. The reason for this disparity in developmental capacity is still unknown. In this study, we compared active demethylation levels and morphological changes between the nuclei of CC-embryos and FF-embryos shortly after activation. Anti-5-methylcytosine immunofluorescence of in vivo-fertilized and cloned rabbit embryos revealed that there was no detectable active demethylation in rabbit zygotes or NT-embryos derived from either fibroblasts or CC. In the process of nuclear remodelling, however, the proportion of nuclei with abnormal appearance in FF-embryos was significantly higher than that in CC-embryos during the first cell cycle. Our study demonstrates that the nuclear remodelling abnormality of cloned rabbit embryos may be one important factor for the disparity in developmental success between CC-embryos and FF-embryos.

  5. Comparison of gene expression in fresh and frozen-thawed human preimplantation embryos.

    PubMed

    Shaw, Lisa; Sneddon, Sharon F; Brison, Daniel R; Kimber, Susan J

    2012-11-01

    Identification and characterisation of differentially regulated genes in preimplantation human embryonic development are required to improve embryo quality and pregnancy rates in IVF. In this study, we examined expression of a number of genes known to be critical for early development and compared expression profiles in individual preimplantation human embryos to establish any differences in gene expression in fresh compared to frozen-thawed embryos used routinely in IVF. We analysed expression of 19 genes by cDNA amplification followed by quantitative real-time PCR in a panel of 44 fresh and frozen-thawed human preimplantation embryos. Fresh embryos were obtained from surplus early cleavage stage embryos and frozen-thawed embryos from cryopreserved 2PN embryos. Our aim was to determine differences in gene expression between fresh and frozen-thawed human embryos, but we also identified differences in developmental expression patterns for particular genes. We show that overall gene expression among embryos of the same stage is highly variable and our results indicate that expression levels between groups did differ and differences in expression of individual genes was detected. Our results show that gene expression from frozen-thawed embryos is more consistent when compared with fresh, suggesting that cryopreserved embryos may represent a reliable source for studying the molecular events underpinning early human embryo development.

  6. CHONDRULE FORMATION IN BOW SHOCKS AROUND ECCENTRIC PLANETARY EMBRYOS

    SciTech Connect

    Morris, Melissa A.; Desch, Steven J.; Athanassiadou, Themis; Boley, Aaron C.

    2012-06-10

    Recent isotopic studies of Martian meteorites by Dauphas and Pourmand have established that large ({approx}3000 km radius) planetary embryos existed in the solar nebula at the same time that chondrules-millimeter-sized igneous inclusions found in meteorites-were forming. We model the formation of chondrules by passage through bow shocks around such a planetary embryo on an eccentric orbit. We numerically model the hydrodynamics of the flow and find that such large bodies retain an atmosphere with Kelvin-Helmholtz instabilities allowing mixing of this atmosphere with the gas and particles flowing past the embryo. We calculate the trajectories of chondrules flowing past the body and find that they are not accreted by the protoplanet, but may instead flow through volatiles outgassed from the planet's magma ocean. In contrast, chondrules are accreted onto smaller planetesimals. We calculate the thermal histories of chondrules passing through the bow shock. We find that peak temperatures and cooling rates are consistent with the formation of the dominant, porphyritic texture of most chondrules, assuming a modest enhancement above the likely solar nebula average value of chondrule densities (by a factor of 10), attributable to settling of chondrule precursors to the midplane of the disk or turbulent concentration. We calculate the rate at which a planetary embryo's eccentricity is damped and conclude that a single planetary embryo scattered into an eccentric orbit can, over {approx}10{sup 5} years, produce {approx}10{sup 24} g of chondrules. In principle, a small number (1-10) of eccentric planetary embryos can melt the observed mass of chondrules in a manner consistent with all known constraints.

  7. The Roles of Glutathione Peroxidases during Embryo Development.

    PubMed

    Ufer, Christoph; Wang, Chi Chiu

    2011-01-01

    Embryo development relies on the complex interplay of the basic cellular processes including proliferation, differentiation, and apoptotic cell death. Precise regulation of these events is the basis for the establishment of embryonic structures and the organ development. Beginning with fertilization of the oocyte until delivery the developing embryo encounters changing environmental conditions such as varying levels of oxygen, which can give rise to reactive oxygen species (ROS). These challenges are met by the embryo with metabolic adaptations and by an array of anti-oxidative mechanisms. ROS can be deleterious by modifying biological molecules including lipids, proteins, and nucleic acids and may induce abnormal development or even embryonic lethality. On the other hand ROS are vital players of various signaling cascades that affect the balance between cell growth, differentiation, and death. An imbalance or dysregulation of these biological processes may generate cells with abnormal growth and is therefore potentially teratogenic and tumorigenic. Thus, a precise balance between processes generating ROS and those decomposing ROS is critical for normal embryo development. One tier of the cellular protective system against ROS constitutes the family of selenium-dependent glutathione peroxidases (GPx). These enzymes reduce hydroperoxides to the corresponding alcohols at the expense of reduced glutathione. Of special interest within this protein family is the moonlighting enzyme glutathione peroxidase 4 (Gpx4). This enzyme is a scavenger of lipophilic hydroperoxides on one hand, but on the other hand can be transformed into an enzymatically inactive cellular structural component. GPx4 deficiency - in contrast to all other GPx family members - leads to abnormal embryo development and finally produces a lethal phenotype in mice. This review is aimed at summarizing the current knowledge on GPx isoforms during embryo development and tumor development with an emphasis on

  8. Bessel beam fluorescence lifetime tomography of live embryos (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Xu, Dongli; Peng, Leilei

    2016-03-01

    Optical tomography allows isotropic 3D imaging of embryos. Scanning-laser optical tomography (SLOT) has superior light collecting efficiency than wide-field optical tomography, making it ideal for fluorescence imaging of live embryos. We previously reported an imaging system that combines SLOT with a novel Fourier-multiplexed fluorescence lifetime imaging (FmFLIM) technique named FmFLIM-SLOT. FmFLIM-SLOT performs multiplexed FLIM-FRET readout of multiple FRET sensors in live embryos. Here we report a recent effort on improving the spatial resolution of the FmFLIM-SLOT system in order to image complex biochemical processes in live embryos at the cellular level. Optical tomography has to compromise between resolution and the depth of view. In SLOT, the commonly-used focused Gaussian beam diverges quickly from the focal plane, making it impossible to achieve high resolution imaging in a large volume specimen. We thus introduce Bessel beam laser-scanning tomography, which illuminates the sample with a spatial-light-modulator-generated Bessel beam that has an extended focal depth. The Bessel beam is scanned across the whole specimen. Fluorescence projection images are acquired at equal angular intervals as the sample rotates. Reconstruction artifacts due to annular-rings of the Bessel beam are removed by a modified 3D filtered back projection algorithm. Furthermore, in combination of Fourier-multiplexing fluorescence lifetime imaging (FmFLIM) method, the Bessel FmFLIM-SLOT system is capable of perform 3D lifetime imaging of live embryos at cellular resolution. The system is applied to in-vivo imaging of transgenic Zebrafish embryos. Results prove that Bessel FmFLIM-SLOT is a promising imaging method in development biology research.

  9. A computational model for BMP movement in sea urchin embryos.

    PubMed

    van Heijster, Peter; Hardway, Heather; Kaper, Tasso J; Bradham, Cynthia A

    2014-12-21

    Bone morphogen proteins (BMPs) are distributed along a dorsal-ventral (DV) gradient in many developing embryos. The spatial distribution of this signaling ligand is critical for correct DV axis specification. In various species, BMP expression is spatially localized, and BMP gradient formation relies on BMP transport, which in turn requires interactions with the extracellular proteins Short gastrulation/Chordin (Chd) and Twisted gastrulation (Tsg). These binding interactions promote BMP movement and concomitantly inhibit BMP signaling. The protease Tolloid (Tld) cleaves Chd, which releases BMP from the complex and permits it to bind the BMP receptor and signal. In sea urchin embryos, BMP is produced in the ventral ectoderm, but signals in the dorsal ectoderm. The transport of BMP from the ventral ectoderm to the dorsal ectoderm in sea urchin embryos is not understood. Therefore, using information from a series of experiments, we adapt the mathematical model of Mizutani et al. (2005) and embed it as the reaction part of a one-dimensional reaction-diffusion model. We use it to study aspects of this transport process in sea urchin embryos. We demonstrate that the receptor-bound BMP concentration exhibits dorsally centered peaks of the same type as those observed experimentally when the ternary transport complex (Chd-Tsg-BMP) forms relatively quickly and BMP receptor binding is relatively slow. Similarly, dorsally centered peaks are created when the diffusivities of BMP, Chd, and Chd-Tsg are relatively low and that of Chd-Tsg-BMP is relatively high, and the model dynamics also suggest that Tld is a principal regulator of the system. At the end of this paper, we briefly compare the observed dynamics in the sea urchin model to a version that applies to the fly embryo, and we find that the same conditions can account for BMP transport in the two types of embryos only if Tld levels are reduced in sea urchin compared to fly.

  10. Cloning of bovine embryos by multiple nuclear transfer.

    PubMed

    Takano, H; Kozai, C; Shimizu, S; Kato, Y; Tsunoda, Y

    1997-05-01

    The in vitro development of multiple generation bovine nuclear transferred embryos to blastocysts and their survival ability after freezing and thawing were examined. Parent donor embryos which had 20 to 50 cells were recovered from superovulated cows. Follicular oocytes matured in vitro were used as recipient oocytes. The recipient oocytes enucleated at 22 to 24 h after the onset of maturation were preactivated at 33 h. Enucleated oocytes with a donor blastomere were fused 9 h after activation by an electric stimulus and the fused oocytes were cultured in vitro (first generation). Reconstituted oocytes that had developed to the 8- to 16-cell stage 3 to 4 d after fusion were used as donor embryos for the next generation. Recloning procedures were performed twice (second and third generations). The proportion of recipient oocytes successfully fused with a blastomere increased with the cycle of nuclear transfer. Eighty to 86% of fused oocytes developed to the 2-cell stage and there was no significant difference with the generation. The proportion of reconstituted embryos receiving blastomeres derived from first generation embryos had higher developmental ability in vitro, than those derived from other generations (43 vs 31% for 8 to 16-cell stage, 37 vs 20 and 21% for blastocyst stage). The number of cloned blastocysts increased with repeated nuclear transfer (once: 6.2 +/- 4.3, twice: 19.8 +/- 9.2 and three times: 30.0 +/- 14.7) but varied greatly with each parent donor embryo. The in vitro viability of cloned blastocysts after freezing and thawing (59%) was low but not significantly different from that obtained for in vitro fertilized blastocysts (72%). After transfer of either fresh or frozen-thawed cloned blastocysts to 21 recipients, 10 of them were pregnant on Day 60. Four and 3 offspring were produced from 20 fresh and 14 frozen-thawed blastocysts,respectively.

  11. The Hamster as a Model for Embryo Implantation

    PubMed Central

    Reese, Jeff; Wang, Hehai; Ding, Tianbing; Paria, B. C.

    2008-01-01

    Defects in preimplantation embryonic development, uterine receptivity, and implantation are the leading cause of infertility, pregnancy problems and birth defects. Significant progress has been made in our basic understanding of these processes using the mouse model, where implantation is ovarian estrogen-dependent in the presence of progesterone. However, an animal model where implantation is progesterone-dependent must also be studied to gain a full understanding of the embryo and uterine events that are required for implantation. In this regard, the hamster is a useful model and this review summarizes the information currently available regarding mechanisms involved in synchronous preimplantation embryo and uterine development for implantation in this species. PMID:18178492

  12. The Constitution of the Human Embryo as Substantial Change

    PubMed Central

    Alvargonzález, David

    2016-01-01

    This paper analyzes the transformation from the human zygote to the implanted embryo under the prism of substantial change. After a brief introduction, it vindicates the Aristotelian ideas of substance and accident, and those of substantial and accidental change. It then claims that the transformation from the multicelled zygote to the implanted embryo amounts to a substantial change. Pushing further, it contends that this substantial change cannot be explained following patterns of genetic reductionism, emergence, and self-organization, and proposes Gustavo Bueno’s idea of anamorphosis as a means to encapsulate criticism against such positions. PMID:26850033

  13. Regeneration of cilia in heavily irradiated sea urchin embryos

    SciTech Connect

    Rustad, R.C.

    1981-12-01

    Cilia were removed from blastulae, gastrulae, and plutei of the sea urchins Arbacia punctulata and Lytechinus variegatus by shaking the embryos in hypertonic media. Exposure to 50 krad (and in some experiments 100 krad) of ..gamma.. radiation either before or after deciliation had no effect on the time of appearance of regenerating cilia. There were no visually obvious differences in the rate of growth of the cilia in control and irradiated embryos. The cilia commenced beating at the same time, but the initial beating sometimes seemed less vigorous following irradiation. The data support the hypothesis that radiation has no major effect on the assembly from mature basal bodies of the microtubules of cilia.

  14. Superovulation and embryo transfer in wood bison (Bison bison athabascae).

    PubMed

    Toosi, Behzad M; Tribulo, Andres; Lessard, Carl; Mastromonaco, Gabriela F; McCorkell, Robert B; Adams, Gregg P

    2013-09-15

    Two experiments were done to develop an effective superovulatory treatment protocol in wood bison for the purpose of embryo collection and transfer. In experiment 1, donor bison were assigned randomly to four treatment groups (N = 5 per group) to examine the effects of method of synchronization (follicular ablation vs. estradiol-progesterone treatment) and ovarian follicular superstimulation (single slow-release vs. split dose of FSH). Recipient bison were synchronized with donor bison by either follicular ablation (N = 8) or estradiol-progesterone treatment (N = 9). In experiment 2, bison were assigned randomly to four treatment groups (N = 5 per group) to examine the ovarian response to two versus four doses of FSH, and the effect of progesterone (ovarian superstimulation with or without an intravaginal progesterone-releasing device). Donor bison were inseminated with fresh chilled wood bison semen 12 and 24 hours after treatment with GnRH (experiment 1) or LH (experiment 2). The ovarian response was assessed using ultrasonography. In experiment 1, the number of large follicles (≥ 7 mm) increased in response to both FSH treatments, but the diameter of the largest follicle detected 4 and 5 days after the start of ovarian superstimulation was greater in bison treated with a single dose of FSH than in those treated with two doses (P < 0.05). A total of 10 ova and/or embryos were collected. One blastocyst was transferred to each of five recipient bison resulting in the birth of two live wood bison calves. In experiment 2, two doses of FSH resulted in a greater number of large follicles (≥ 9 mm) on Days 4, 5, and 6 (P < 0.05) after beginning of superstimulation (Day 0), and more ovulations than four doses of FSH (11.2 ± 2.4 vs. 6.4 ± 0.8; P < 0.05). Embryo collection was performed on only five donors, and a total of 19 ova and/or embryos were recovered. In summary, fewer FSH treatments were as good or better than multiple treatments, consistent with the notion

  15. Functional studies of regulatory genes in the sea urchin embryo.

    PubMed

    Cavalieri, Vincenzo; Di Bernardo, Maria; Spinelli, Giovanni

    2009-01-01

    Sea urchin embryos are characterized by an extremely simple mode of development, rapid cleavage, high transparency, and well-defined cell lineage. Although they are not suitable for genetic studies, other approaches are successfully used to unravel mechanisms and molecules involved in cell fate specification and morphogenesis. Microinjection is the elective method to study gene function in sea urchin embryos. It is used to deliver precise amounts of DNA, RNA, oligonucleotides, peptides, or antibodies into the eggs or even into blastomeres. Here we describe microinjection as it is currently applied in our laboratory and show how it has been used in gene perturbation analyses and dissection of cis-regulatory DNA elements.

  16. [Embryo initiation from Pinus sibirica megagametophytes in in vitro culture].

    PubMed

    Tret'iakova, I N; Voroshilova, E V

    2014-01-01

    Megagametophytes of Siberian pine were cultured on an in vitro culture medium 1/2 LV supplemented with growth regulators 2,4-dichlorophenoxyacetic acid (2,4-D) and benzylaminopurine (6-BAP) to form embryos. The competency of somatic cell of explants to embryogenesis manifested itself in an organized growth and polarity. A coenocyte consisting of long vacuolated cells was formed in the megagametophyte culture. Then, the migration of the nuclei to one of the poles of the cell, their division, and formation of embryoids was observed. The megagametophyte culture of the Siberian pine differed from the zygotic embryo culture by the absence of asymmetric division in the vacuolated cell.

  17. Toxicity of selenium to developing Xenopus laevis embryos.

    PubMed

    Browne, C L; Dumont, J N

    1979-07-01

    Se in the form of sodium selenite is toxic to Xenopus laevis embryos and tadpoles continuously exposed to concentrations above 1 ppm. Concentrations of 2 ppm and above result in severe developmental abnormalities and increased mortality. Uptake and loss of radioactive Se from water are rapid, but depuration is not complete indicating that some Se can remain bound by the organism. The facts that Se is toxic at low levels to Xenopus embryos and tadpoles, can cause developmental abnormalities, and accumulates in tissues suggest that increased release of Se compounds into the environment poses a potential threat to aquatic organisms.

  18. Recent microfluidic devices for studying gamete and embryo biomechanics.

    PubMed

    Lai, David; Takayama, Shuichi; Smith, Gary D

    2015-06-25

    The technical challenges of biomechanic research such as single cell analysis at a high monetary cost, labor, and time for just a small number of measurements is a good match to the strengths of microfluidic devices. New scientific discoveries in the fertilization and embryo development process, of which biomechanics is a major subset of interest, is crucial to fuel the continual improvement of clinical practice in assisted reproduction. The following review will highlight some recent microfluidic devices tailored for gamete and embryo biomechanics where biomimicry arises as a major theme of microfluidic device design and function, and the application of fundamental biomechanic principles are used to improve outcomes of cryopreservation.

  19. Day 2 versus day 3 embryo transfer in poor responders: a prospective randomized trial.

    PubMed

    Shahine, Lora K; Milki, Amin A; Westphal, Lynn M; Baker, Valerie L; Behr, Barry; Lathi, Ruth B

    2011-01-01

    Day 2 embryo transfer has been suggested as a method to improve pregnancy rates in poor responders compared with day 3 transfer. Our prospective randomized controlled trial does not show a difference in outcomes based on day of embryo transfer.

  20. OpenSource lab-on-a-chip physiometer for accelerated zebrafish embryo biotests.

    PubMed

    Akagi, Jin; Hall, Chris J; Crosier, Kathryn E; Cooper, Jonathan M; Crosier, Philip S; Wlodkowic, Donald

    2014-01-02

    Zebrafish (Danio rerio) embryo assays have recently come into the spotlight as convenient experimental models in both biomedicine and ecotoxicology. As a small aquatic model organism, zebrafish embryo assays allow for rapid physiological, embryo-, and genotoxic tests of drugs and environmental toxins that can be simply dissolved in water. This protocol describes prototyping and application of an innovative, miniaturized, and polymeric chip-based device capable of immobilizing a large number of living fish embryos for real-time and/or time-lapse microscopic examination. The device provides a physical address designation to each embryo during analysis, continuous perfusion of medium, and post-analysis specimen recovery. Miniaturized embryo array is a new concept of immobilization and real-time drug perfusion of multiple individual and developing zebrafish embryos inside the mesofluidic device. The OpenSource device presented in this protocol is particularly suitable to perform accelerated fish embryo biotests in ecotoxicology and phenotype-based pharmaceutical screening.

  1. [Sensitivity of fragmented and stratified sea urchin embryos to cytotoxic neuropharmacological preparations].

    PubMed

    Buznikov, G A; Manukhin, B N; Rakic, L; Kudriashova, N I; Mndzhoian, O L

    1979-01-01

    The red half embryos and related quarter embryos (yolk and pigment) of Arbacia lixula, obtained by means of centrifugation of the eggs in sucrose gradient, retain the normal level of sensitivity and supersensitivity to cytotoxic neuropharmaca, antagonists of biogenic monoamines. The white half embryos and clear quarter embryos practically lack supersensitivity whereas the granular quarter embryos restore it to the initial level. The non-pigmented blastomers of stratified embryos are characterized by somewhat weakened supersensitivity. A suggestion is put forward that the supersensitive embryos of A. lixula possess a sensibilizing factor which couples the supersensitivity receptors with the processes of cell division and moves together with the yolk granules upon centrifugation. This factor is not observed in the Strongylocentrotus granularis embryos lacking evident supersensitivity.

  2. [Controversy in ART: should we cryopreserve oocytes or embryos? Do prefer oocytes].

    PubMed

    Boyer, P

    2014-09-01

    Since the beginning of IVF, cryopreservation concern spermatozoa or embryos due to the poor efficiency of oocyte freezing. To date, oocyte vitrification allows changing our practice privileging female gamete vitrification instead of human embryo freezing.

  3. Identification of the glycerol kinase gene and its role in diapause embryo restart and early embryo development of Artemia sinica.

    PubMed

    Cheng, Cheng; Yao, Feng; Chu, Bing; Li, Xuejie; Liu, Yan; Wu, Yang; Mei, Yanli; Wang, Peisheng; Hou, Lin; Zou, Xiangyang

    2014-03-01

    Glycerol kinase (GK) catalyzes the rate-limiting step in glycerol utilization by transferring a phosphate from ATP to glycerol, yielding glycerol 3-phosphate, which is an important intermediate for both energy metabolism and glycerolipid production. Artemia sinica has an unusual diapause process under stress conditions of high salinity, low temperature and lack of food. In the process, diapause embryos of A. sinica (brine shrimp) accumulate high concentrations of glycerol as a cryoprotectant to prevent low temperature damage to embryos. Upon embryo restart, glycerol is converted into glucose and other carbohydrates. Therefore, GK plays an important role in the diapause embryo restart process. However, the role of GK in diapause termination of embryo development in A. sinica remains unknown. In the present study, a 2096 bp full-length cDNA of gk from A. sinica (As-gk) was obtained, encoding putative 551 amino acids, 60.6 kDa protein. As a crucial enzyme in glycerol uptake and metabolism, GK has been conserved structurally and functionally during evolution. The expression pattern of As-gk was investigated by quantitative real-time PCR and Western blotting. Expression locations of As-gk were analyzed using in situ hybridization. As-gk was widely distributed in the early embryo and several main parts of Artemia after differentiation. The expression of As-GK was also induced by stresses such as cold exposure and high salinity. This initial research into the expression pattern and stress response of GK in Artemia provides a sound basis for further understanding of the function and regulation of genes in early embryonic development in A. sinica and the stress response.

  4. 75 FR 69717 - In the Matter of: Edentify, Inc., Embryo Development Corp., Enclaves Group, Inc., Energytec, Inc...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-11-15

    ..., Inc., Embryo Development Corp., Enclaves Group, Inc., Energytec, Inc., Enesco Group, Inc... securities of Embryo Development Corp. because it has not filed any periodic reports since the period...

  5. Use of DNA strand damage (Comet assay) and embryo hatching effects to assess contaminant exposure in blue crab (Callinectes sapidus) embryos

    SciTech Connect

    Lee, R.F.; Steinert, S.A.; Nakayama, K.; Oshima, Y.

    1999-07-01

    After fertilization, blue crab eggs are embedded in a sponge which is attached to the female abdomen during embryo development. Embryos after 9 stages in the egg sac hatch into a swimming zoea stage (stage 10). The authors have developed a bioassay where embryo development is monitored in culture plates with and without toxicants in the water. Toxicant effects are based on determining the percentage of embryos which hatch to zoea. Hatching EC{sub 50} (toxicant concentration at which 50% of the embryos fail to hatch) for a number of pesticides, organometallics and metals were determined. The test takes from 2 to 6 days depending on the embryo stage selected for the study. In addition to embryo development effects the prevalence of DNA single-strand breaks in individual embryo cells were determined using the single cell gel electrophoresis method (Comet assay). A good correlation between DNA strand breakage and embryo defects was found after exposure to genotoxic contaminants. Thus, the bioassay linking DNA damage to embryo hatching effects is rapid, sensitive and mechanistically relevant.

  6. HUWE1 plays important role in mouse preimplantation embryo development and the dysregulation is associated with poor embryo development in humans

    PubMed Central

    Chen, L. J.; Xu, W. M.; Yang, M.; Wang, K.; Chen, Y.; Huang, X. J.; Ma, Q. H.

    2016-01-01

    HUWE1 is a HECT domain containing ubiquitin ligase implicated in neurogenesis, spermatogenesis and cancer development. The purpose of the current study is to investigate the role of HUWE1 in early embryo development. Here we demonstrate that Huwe1 is expressed in both nucleus and cytoplasm of preimplantation mouse embryos as well as gametes. Hypoxia (5% O2) treatment could significantly increase Huwe1 expression during mouse embryo development process. HUWE1 knockdown inhibited normal embryonic development and reduced blastocyst formation, and increased apoptotic cell numbers were observed in the embryos of HUWE1 knockdown group. Human embryo staining result showed that reduced HUWE1 staining was observed in the poor-quality embryos. Furthermore, Western blot result showed that significantly reduced expression of HUWE1 was observed in the villi of miscarriage embryos compared with the normal control, indicating that reduced expression of HUWE1 is related to poor embryo development. Oxidative reagent, H2O2 inhibited HUWE1 expression in human sperm, indicating that HUWE1 expression in sperm is regulated by oxidative stress. In conclusion, these results suggest that HUWE1 protein could contribute to preimplantation embryo development and dysregulated expression of HUWE1 could be related to poor embryo development and miscarriage in IVF clinic. PMID:27901130

  7. 10 CFR 835.206 - Limits for the embryo/fetus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 4 2014-01-01 2014-01-01 false Limits for the embryo/fetus. 835.206 Section 835.206... Exposure § 835.206 Limits for the embryo/fetus. (a) The equivalent dose limit for the embryo/fetus from the... provided in § 835.206(a) shall be avoided. (c) If the equivalent dose to the embryo/fetus is determined...

  8. 10 CFR 835.206 - Limits for the embryo/fetus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 4 2012-01-01 2012-01-01 false Limits for the embryo/fetus. 835.206 Section 835.206... Exposure § 835.206 Limits for the embryo/fetus. (a) The equivalent dose limit for the embryo/fetus from the... provided in § 835.206(a) shall be avoided. (c) If the equivalent dose to the embryo/fetus is determined...

  9. 10 CFR 835.206 - Limits for the embryo/fetus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 4 2011-01-01 2011-01-01 false Limits for the embryo/fetus. 835.206 Section 835.206... Exposure § 835.206 Limits for the embryo/fetus. (a) The equivalent dose limit for the embryo/fetus from the... provided in § 835.206(a) shall be avoided. (c) If the equivalent dose to the embryo/fetus is determined...

  10. 10 CFR 835.206 - Limits for the embryo/fetus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 4 2010-01-01 2010-01-01 false Limits for the embryo/fetus. 835.206 Section 835.206... Exposure § 835.206 Limits for the embryo/fetus. (a) The equivalent dose limit for the embryo/fetus from the... provided in § 835.206(a) shall be avoided. (c) If the equivalent dose to the embryo/fetus is determined...

  11. 10 CFR 835.206 - Limits for the embryo/fetus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 4 2013-01-01 2013-01-01 false Limits for the embryo/fetus. 835.206 Section 835.206... Exposure § 835.206 Limits for the embryo/fetus. (a) The equivalent dose limit for the embryo/fetus from the... provided in § 835.206(a) shall be avoided. (c) If the equivalent dose to the embryo/fetus is determined...

  12. Development of rat tetraploid and chimeric embryos aggregated with diploid cells.

    PubMed

    Shinozawa, T; Sugawara, A; Matsumoto, A; Han, Y-J; Tomioka, I; Inai, K; Sasada, H; Kobayashi, E; Matsumoto, H; Sato, E

    2006-11-01

    In the present study, we examined the preimplantation and postimplantation development of rat tetraploid embryos produced by electrofusion of 2-cell-stage embryos. Developmental rate of tetraploid embryos to morula or blastocyst stage was 93% (56/60) and similar to that found in diploid embryos (95%, 55/58). After embryo transfer, rat tetraploid embryos showed implantation and survived until day 8 of pregnancy, however the conceptuses were aberrant on day 9. In mouse, tetraploid embryos have the ability to support the development of blastomeres that cannot develop independently. As shown in the present study, a pair of diploid blastomeres from the rat 8-cell-stage embryo degenerated immediately after implantation. Therefore, we examined whether rat tetraploid embryos have the ability to support the development of 2/8 blastomeres. We produced chimeric rat embryos in which a pair of diploid blastomeres from an 8-cell-stage green fluorescent protein negative (GFP-) embryo was aggregated with three tetraploid blastomeres from 4-cell GFP-positive (GFP+) embryos. The developmental rate of rat 2n(GFP-) <--> 4n(GFP+) embryos to the morula or blastocyst stages was 93% (109/117) and was similar to that found for 2n(GFP-) <--> 2n(GFP+) embryos (100%, 51/51). After embryo transfer, 2n(GFP-) <--> 4n(GFP+) conceptuses were examined on day 14 of pregnancy, the developmental rate to fetus was quite low (4%, 4/109) and they were all aberrant and smaller than 2n(GFP-) <--> 2n(GFP+) conceptuses, whereas immunohistochemical analysis showed no staining for GFP in fetuses. Our results suggest that rat tetraploid embryos are able to prolong the development of diploid blastomeres that cannot develop independently, although postimplantation development was incomplete.

  13. A three-dimensional culture system using alginate hydrogel prolongs hatched cattle embryo development in vitro.

    PubMed

    Zhao, Shuan; Liu, Zhen-Xing; Gao, Hui; Wu, Yi; Fang, Yuan; Wu, Shuai-Shuai; Li, Ming-Jie; Bai, Jia-Hua; Liu, Yan; Evans, Alexander; Zeng, Shen-Ming

    2015-07-15

    No successful method exists to maintain the three-dimensional architecture of hatched embryos in vitro. Alginate, a linear polysaccharide derived from brown algae, has characteristics that make it an ideal material as a three-dimensional (3D) extracellular matrix for in vitro cell, tissue, or embryo culture. In this study, alginate hydrogel was used for IVC of posthatched bovine embryos to observe their development under the 3D system. In vitro-fertilized and parthenogenetically activated posthatched bovine blastocysts were cultured in an alginate encapsulation culture system (AECS), an alginate overlay culture system (AOCS), or control culture system. After 18 days of culture, the survival rate of embryos cultured in AECS was higher than that in the control group (P < 0.05), and the embryos were expanded and elongated in AECS with the maximal length of 1.125 mm. When the AECS shrinking embryos were taken out of the alginate beads on Day 18 and cultured in the normal culture system, 9.09% of them attached to the bottoms of the plastic wells and grew rapidly, with the largest area of an attached embryo being 66.00 mm(2) on Day 32. The embryos cultured in AOCS developed monovesicular or multivesicular morphologies. Total cell number of the embryos cultured in AECS on Day 19 was significantly higher than that of embryos on Day 8. Additionally, AECS and AOCS supported differentiation of the embryonic cells. Binuclear cells were visible in Day-26 adherent embryos, and the messenger RNA expression patterns of Cdx2 and Oct4 in AOCS-cultured embryos were similar to those in vivo embryos, whereas IFNT and ISG15 messenger RNA were still expressed in Day-26 and Day-32 prolong-cultured embryos. In conclusion, AECS and AOCS did support cell proliferation, elongation, and differentiation of hatched bovine embryos during prolonged IVC. The culture system will be useful to further investigate the molecular mechanisms controlling ruminant embryo elongation and implantation.

  14. Developmental kinetics of in vitro-produced bovine embryos: An aid for making decisions.

    PubMed

    Carrocera, S; Caamaño, J N; Trigal, B; Martín, D; Díez, C

    2016-03-15

    Embryo developmental kinetics and embryo survival after cryopreservation have been correlated with embryo quality and viability. The main objectives of this work were to analyze developmental ability and quality of in vitro-produced bovine embryos in relation to their kinetics and to establish a criterion of quality to predict further viability. Embryos were classified and grouped by their specific stage of development (2, 3-4, or ≥ 5 cells) at 44 hours post insemination (hpi) and cultured separately up to Day 8. On Days 7 and 8, good quality expanded blastocysts were vitrified or frozen. Cryopreserved surviving hatched embryos were stained for cell counts. Embryos at a more advanced stage (3-4 cells, and ≥5 cells) developed to morulae (P < 0.001) and blastocysts (P < 0.01) at higher rates than those embryos that had cleaved once by 44 hpi. Vitrification improved the hatching rates of blastocysts at 48 hours (P < 0.001) when compared with slow-rate freezing within each group of embryos (3-4 cells and ≥5 cells). After vitrification/warming, blastocysts coming from 3- to 4-cell embryos had higher hatching rates at 48 hours than those that came from ≥5-cell embryos. With regard to differential cell counts, no effect of the initial developmental stage was observed after warming/thawing. However, trophectoderm and total cells were higher in vitrified/warmed than in the frozen/thawed embryos (P < 0.001). These data show that selecting IVF embryos at 44 hpi, after the evaluation of their in vitro embryo development, could be used as noninvasive markers of embryo developmental competence and may help to select IVF embryos that would be more suitable for cryopreservation.

  15. 10 CFR 20.1208 - Dose equivalent to an embryo/fetus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 1 2012-01-01 2012-01-01 false Dose equivalent to an embryo/fetus. 20.1208 Section 20... Limits § 20.1208 Dose equivalent to an embryo/fetus. (a) The licensee shall ensure that the dose equivalent to the embryo/fetus during the entire pregnancy, due to the occupational exposure of a...

  16. 9 CFR 113.37 - Detection of pathogens by the chicken embryo inoculation test.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... embryo inoculation test. 113.37 Section 113.37 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.37 Detection of pathogens by the chicken embryo...-serum mixture shall be inoculated into each of at least 20 fully susceptible chicken embryos. (1)...

  17. 9 CFR 113.37 - Detection of pathogens by the chicken embryo inoculation test.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... embryo inoculation test. 113.37 Section 113.37 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.37 Detection of pathogens by the chicken embryo...-serum mixture shall be inoculated into each of at least 20 fully susceptible chicken embryos. (1)...

  18. 10 CFR 20.1208 - Dose equivalent to an embryo/fetus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 1 2011-01-01 2011-01-01 false Dose equivalent to an embryo/fetus. 20.1208 Section 20... Limits § 20.1208 Dose equivalent to an embryo/fetus. (a) The licensee shall ensure that the dose equivalent to the embryo/fetus during the entire pregnancy, due to the occupational exposure of a...

  19. 10 CFR 20.1208 - Dose equivalent to an embryo/fetus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 1 2010-01-01 2010-01-01 false Dose equivalent to an embryo/fetus. 20.1208 Section 20... Limits § 20.1208 Dose equivalent to an embryo/fetus. (a) The licensee shall ensure that the dose equivalent to the embryo/fetus during the entire pregnancy, due to the occupational exposure of a...

  20. 9 CFR 113.37 - Detection of pathogens by the chicken embryo inoculation test.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... embryo inoculation test. 113.37 Section 113.37 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.37 Detection of pathogens by the chicken embryo...-serum mixture shall be inoculated into each of at least 20 fully susceptible chicken embryos. (1)...

  1. 9 CFR 113.37 - Detection of pathogens by the chicken embryo inoculation test.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... embryo inoculation test. 113.37 Section 113.37 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.37 Detection of pathogens by the chicken embryo...-serum mixture shall be inoculated into each of at least 20 fully susceptible chicken embryos. (1)...

  2. 9 CFR 113.37 - Detection of pathogens by the chicken embryo inoculation test.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... embryo inoculation test. 113.37 Section 113.37 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.37 Detection of pathogens by the chicken embryo...-serum mixture shall be inoculated into each of at least 20 fully susceptible chicken embryos. (1)...

  3. 10 CFR 20.1208 - Dose equivalent to an embryo/fetus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 1 2013-01-01 2013-01-01 false Dose equivalent to an embryo/fetus. 20.1208 Section 20... Limits § 20.1208 Dose equivalent to an embryo/fetus. (a) The licensee shall ensure that the dose equivalent to the embryo/fetus during the entire pregnancy, due to the occupational exposure of a...

  4. 10 CFR 20.1208 - Dose equivalent to an embryo/fetus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 1 2014-01-01 2014-01-01 false Dose equivalent to an embryo/fetus. 20.1208 Section 20... Limits § 20.1208 Dose equivalent to an embryo/fetus. (a) The licensee shall ensure that the dose equivalent to the embryo/fetus during the entire pregnancy, due to the occupational exposure of a...

  5. Why we should not select the faster embryo: lessons from mice and cattle.

    PubMed

    Gutierrez-Adan, Alfonso; White, Carlee R; Van Soom, Ann; Mann, Mellissa R W

    2015-06-01

    Many studies have shown that in vitro culture can negatively impact preimplantation development. This necessitates some selection criteria for identifying the best-suited embryos for transfer. That said, embryo selection after in vitro culture remains a subjective process in most mammalian species, including cows, mice and humans. General consensus in the field is that embryos that develop in a timely manner have the highest developmental competence and viability after transfer. Herein lies the key question: what is a timely manner? With emerging data in bovine and mouse supporting increased developmental competency in embryos with moderate rates of development, it is time to question whether the fastest developing embryos are the best embryos for transfer in the human clinic. This is especially relevant to epigenetic gene regulation, including genomic imprinting, where faster developing embryos exhibit loss of imprinted methylation, as well as to sex selection bias, where faster developmental rates of male embryos may lead to biased embryo transfer and, in turn, biased sex ratios. In this review, we explore evidence surrounding the question of developmental timing as it relates to bovine embryo quality, mouse embryo quality and genomic imprint maintenance, and embryo sex.

  6. Evaluation of a quali embryo model for the detection of botulism toxin type A activity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Japanese quail embryo (Coturnix japonica) was evaluated for use as a bioassay to detect biologically active botulinum toxin serotype A (BoNT/A). Day 15 of incubation embryos were injected with decreasing dosages of BoNT/A from 250 to 0.5 ng of toxin. At 1 day post-injection, embryos receiving ...

  7. Pregnancy rate following transfer of in vitro produced lamb derived embryos in two embryonic stages.

    PubMed

    Shirazi, A; Shams-Esfandabadi, N; Ahmadi, E; Jadidi, M; Heidari, B

    2008-03-15

    Ovine embryos were produced by maturation, fertilization and in vitro culture (IVM/IVF/IVC) of oocytes collected from slaughtered prepubertal ewes. At 24 h post IVM, oocytes were fertilized with fresh semen collected from Lori-Bakhtiari breed at a concentration of 1.0 x l0(6) sperm mL(-1). The presumptive ova/embryos were transferred into the embryo culture medium at 22-24 h post IVF. Following 4 to 7 day in culture, embryos (at morula and blastocyst stage, respectively) were transferred surgically to the uterine horn of synchronized recipients. Pregnancy was diagnosed at day 30 by hormonal assay and at days 55 and 140 of gestation by ultrasonography and pregnancies were allowed to go to term. A total of nine ewes received 27 embryos (3 embryos/ewe). Five ewes received 15 embryos at morula stage and four ewes received 12 embryos at blastocyst stage. From those received morula stage embryos one was pregnant on day 30 (20%), though no pregnancy was diagnosed on each of days 55 and 140. While from those received blastocyst stage embryos, three ewes were pregnant on day 30 (75%) and two ewes (50%) remained pregnant on each of days 55 and 140. In conclusion, day 4 IVM-IVF morula stage embryos had a lower survival rate than did day 7 IVM-IVF blastocysts embryos, following transfer to the synchronized recipient ewes.

  8. Avian egg odour encodes information on embryo sex, fertility and development.

    PubMed

    Webster, Ben; Hayes, William; Pike, Thomas W

    2015-01-01

    Avian chemical communication is a rapidly emerging field, but has been hampered by a critical lack of information on volatile chemicals that communicate ecologically relevant information (semiochemicals). A possible, but as yet unexplored, function of olfaction and chemical communication in birds is in parent-embryo and embryo-embryo communication. Communication between parents and developing embryos may act to mediate parental behaviour, while communication between embryos can control the synchronicity of hatching. Embryonic vocalisations and vibrations have been implicated as a means of communication during the later stages of development but in the early stages, before embryos are capable of independent movement and vocalisation, this is not possible. Here we show that volatiles emitted from developing eggs of Japanese quail (Coturnix japonica) convey information on egg fertility, along with the sex and developmental status of the embryo. Specifically, egg volatiles changed over the course of incubation, differed between fertile and infertile eggs, and were predictive of embryo sex as early as day 1 of incubation. Egg odours therefore have the potential to facilitate parent-embryo and embryo-embryo interactions by allowing the assessment of key measures of embryonic development long before this is possible through other modalities. It also opens up the intriguing possibility that parents may be able to glean further relevant information from egg volatiles, such as the health, viability and heritage of embryos. By determining information conveyed by egg-derived volatiles, we hope to stimulate further investigation into the ecological role of egg odours.

  9. Transcriptomic and proteomic analysis reveals mechanisms of embryo abortion during chrysanthemum cross breeding

    PubMed Central

    Zhang, Fengjiao; Wang, Zhiquan; Dong, Wen; Sun, Chunqing; Wang, Haibin; Song, Aiping; He, Lizhong; Fang, Weimin; Chen, Fadi; Teng, Nianjun

    2014-01-01

    Embryo abortion is the main cause of failure in chrysanthemum cross breeding, and the genes and proteins associated with embryo abortion are poorly understood. Here, we applied RNA sequencing and isobaric tags for relative and absolute quantitation (iTRAQ) to analyse transcriptomic and proteomic profiles of normal and abortive embryos. More than 68,000 annotated unigenes and 700 proteins were obtained from normal and abortive embryos. Functional analysis showed that 140 differentially expressed genes (DEGs) and 41 differentially expressed proteins (DEPs) were involved in embryo abortion. Most DEGs and DEPs associated with cell death, protein degradation, reactive oxygen species scavenging, and stress-response transcriptional factors were significantly up-regulated in abortive embryos relative to normal embryos. In contrast, most genes and proteins related to cell division and expansion, the cytoskeleton, protein synthesis and energy metabolism were significantly down-regulated in abortive embryos. Furthermore, abortive embryos had the highest activity of three executioner caspase-like enzymes. These results indicate that embryo abortion may be related to programmed cell death and the senescence- or death-associated genes or proteins contribute to embryo abortion. This adds to our understanding of embryo abortion and will aid in the cross breeding of chrysanthemum and other crops in the future. PMID:25288482

  10. Robust embryo identification using first polar body single nucleotide polymorphism microarray-based DNA fingerprinting.

    PubMed

    Treff, Nathan R; Su, Jing; Kasabwala, Natasha; Tao, Xin; Miller, Kathleen A; Scott, Richard T

    2010-05-01

    This study sought to validate a novel, minimally invasive system for embryo tracking by single nucleotide polymorphism microarray-based DNA fingerprinting of the first polar body. First polar body-based assignments of which embryos implanted and were delivered after multiple ET were 100% consistent with previously validated embryo DNA fingerprinting-based assignments.

  11. Evaluation of a quail embryo model for the detection of botulinum toxin type A activity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The quail embryo was evaluated for use as a bioassay to detect biologically active botulinum toxin serotype A (BoNT/A). Day 15 of incubation embryos were injected with decreasing dosages of BoNT/A from 250 to 0.5 ng of toxin. At 1 day post-injection, embryos receiving 20 ng of BoNT or higher had m...

  12. A Marble Embryo: Meanings of a Portrait from 1900

    PubMed Central

    Hopwood, Nick

    2012-01-01

    Portraits of scientists use attributes of discovery to construct identities; portraits that include esoteric accessories may fashion identities for these too. A striking example is a marble bust of the anatomist Wilhelm His by the Leipzig sculptor Carl Seffner. Made in 1900, it depicts the founder of modern human embryology looking down at a model embryo in his right hand. This essay reconstructs the design and viewing of this remarkable portrait in order to shed light on private and public relations between scientists, research objects and audiences. The bust came out of a collaboration to model the face of the composer Johann Sebastian Bach and embodies a shared commitment to anatomical exactitude in three dimensions. His’s research agendas and public character explain the contemplative pose and unprecedented embryo model, which he had laboriously constructed from material a midwife supplied. The early contexts of display in the His home and art exhibitions suggest interpretive resources for viewers and hence likely meanings. Seffner’s work remains exceptional, but has affinities to portraits of human embryologists and embryos produced since 1960. Embryo images have acquired such controversial prominence that the model may engage us more strongly now than it did exhibition visitors around 1900. PMID:22606754

  13. Moist multi-scale models for the hurricane embryo

    SciTech Connect

    Majda, Andrew J.; Xing, Yulong; Mohammadian, Majid

    2010-01-01

    Determining the finite-amplitude preconditioned states in the hurricane embryo, which lead to tropical cyclogenesis, is a central issue in contemporary meteorology. In the embryo there is competition between different preconditioning mechanisms involving hydrodynamics and moist thermodynamics, which can lead to cyclogenesis. Here systematic asymptotic methods from applied mathematics are utilized to develop new simplified moist multi-scale models starting from the moist anelastic equations. Three interesting multi-scale models emerge in the analysis. The balanced mesoscale vortex (BMV) dynamics and the microscale balanced hot tower (BHT) dynamics involve simplified balanced equations without gravity waves for vertical vorticity amplification due to moist heat sources and incorporate nonlinear advective fluxes across scales. The BMV model is the central one for tropical cyclogenesis in the embryo. The moist mesoscale wave (MMW) dynamics involves simplified equations for mesoscale moisture fluctuations, as well as linear hydrostatic waves driven by heat sources from moisture and eddy flux divergences. A simplified cloud physics model for deep convection is introduced here and used to study moist axisymmetric plumes in the BHT model. A simple application in periodic geometry involving the effects of mesoscale vertical shear and moist microscale hot towers on vortex amplification is developed here to illustrate features of the coupled multi-scale models. These results illustrate the use of these models in isolating key mechanisms in the embryo in a simplified content.

  14. Ex Ovo Model for Directly Visualizing Chick Embryo Development

    ERIC Educational Resources Information Center

    Dorrell, Michael I.; Marcacci, Michael; Bravo, Stephen; Kurz, Troy; Tremblay, Jacob; Rusing, Jack C.

    2012-01-01

    We describe a technique for removing and growing chick embryos in culture that utilizes relatively inexpensive materials and requires little space. It can be readily performed in class by university, high school, or junior high students, and teachers of any grade level should be able to set it up for their students. Students will be able to…

  15. Intersensory Redundancy Educates Selective Attention in Bobwhite Quail Embryos

    ERIC Educational Resources Information Center

    Lickliter, Robert; Bahrick, Lorraine E.; Markham, Rebecca G.

    2006-01-01

    We assessed whether exposure to amodal properties in bimodal stimulation (e.g. rhythm, rate, duration) could educate attention to amodal properties in subsequent unimodal stimulation during prenatal development. Bobwhite quail embryos were exposed to an individual bobwhite maternal call under several experimental and control conditions during the…

  16. Intersensory Redundancy Enhances Memory in Bobwhite Quail Embryos

    ERIC Educational Resources Information Center

    Lickliter, Robert; Bahrick, Lorraine E.; Honeycutt, Hunter

    2004-01-01

    Information presented concurrently and redundantly to 2 or more senses (intersensory redundancy) has been shown to recruit attention and promote perceptual learning of amodal stimulus properties in animal embryos and human infants. This study examined whether the facilitative effect of intersensory redundancy also extends to the domain of memory.…

  17. Culturing Chick Embryos--A Simplification of New's Method.

    ERIC Educational Resources Information Center

    Downie, J. R.

    1979-01-01

    Describes a simplified version of New's method for culturing early chick embryos. The technique allows continuous observation of the critical first three days of development and the conditions for setting up successful cultures are also presented to help both teachers and students. (HM)

  18. Genetics of Lipid-Storage Management in Caenorhabditis elegans Embryos

    PubMed Central

    Schmökel, Verena; Memar, Nadin; Wiekenberg, Anne; Trotzmüller, Martin; Schnabel, Ralf; Döring, Frank

    2016-01-01

    Lipids play a pivotal role in embryogenesis as structural components of cellular membranes, as a source of energy, and as signaling molecules. On the basis of a collection of temperature-sensitive embryonic lethal mutants, a systematic database search, and a subsequent microscopic analysis of >300 interference RNA (RNAi)–treated/mutant worms, we identified a couple of evolutionary conserved genes associated with lipid storage in Caenorhabditis elegans embryos. The genes include cpl-1 (cathepsin L–like cysteine protease), ccz-1 (guanine nucleotide exchange factor subunit), and asm-3 (acid sphingomyelinase), which is closely related to the human Niemann-Pick disease–causing gene SMPD1. The respective mutant embryos accumulate enlarged droplets of neutral lipids (cpl-1) and yolk-containing lipid droplets (ccz-1) or have larger genuine lipid droplets (asm-3). The asm-3 mutant embryos additionally showed an enhanced resistance against C band ultraviolet (UV-C) light. Herein we propose that cpl-1, ccz-1, and asm-3 are genes required for the processing of lipid-containing droplets in C. elegans embryos. Owing to the high levels of conservation, the identified genes are also useful in studies of embryonic lipid storage in other organisms. PMID:26773047

  19. Antioxidant Rescue of Selenomethionine-Induced Teratogenesis in Zebrafish Embryos.

    PubMed

    Arnold, M C; Forte, J E; Osterberg, J S; Di Giulio, R T

    2016-02-01

    Selenium (Se) is an essential micronutrient that can be found at toxic concentrations in surface waters contaminated by runoff from agriculture and coal mining. Zebrafish (Danio rerio) embryos were exposed to aqueous Se in the form of selenate, selenite, and l-selenomethionine (SeMet) in an attempt to determine if oxidative stress plays a role in selenium embryo toxicity. Selenate and selenite exposure did not induce embryo deformities (lordosis and craniofacial malformation). l-selenomethionine, however, induced significantly higher deformity rates at 100 µg/L compared with controls. SeMet exposure induced a dose-dependent increase in the catalytic subunit of glutamate-cysteine ligase (gclc) and reached an 11.7-fold increase at 100 µg/L. SeMet exposure also reduced concentrations of TGSH, RGSH, and the TGSH:GSSG ratio. Pretreatment with 100 µM N-acetylcysteine significantly reduced deformities in the zebrafish embryos secondarily treated with 400 µg/L SeMet from approximately 50–10 % as well as rescued all three of the significant glutathione level differences seen with SeMet alone. Selenite exposure induced a 6.6-fold increase in expression of the glutathione-S-transferase pi class 2 (gstp2) gene, which is involved in xenobiotic transformation and possibly oxidative stress. These results suggest that aqueous exposure to SeMet can induce significant embryonic teratogenesis in zebrafish that are at least partially attributed to oxidative stress.

  20. Transcriptome dynamics in early embryos of the ascidian, Ciona intestinalis.

    PubMed

    Matsuoka, Terumi; Ikeda, Tatsuro; Fujimaki, Kotaro; Satou, Yutaka

    2013-12-15

    Maternally provided mRNAs and proteins direct early development and activate the zygotic genome. Using microarrays, we examined the dynamics of transcriptomes during the early development of a basal chordate, Ciona intestinalis. Microarray analysis of unfertilized eggs, as well as 8-, and 16- and 32-cell embryos revealed that nearly half of the genes encoded in the genome were expressed maternally, and that approximately only one-fourth of these genes were expressed at similar levels among eggs obtained from different individuals. Genes encoding proteins involved in protein phosphorylation were enriched in this latter group. More than 90% of maternal RNAs were not reduced before the 16-cell stage when the zygotic developmental program begins. Additionally we obtained gene expression profiles of individual blastomeres from the 8- and 16-cell embryos. On the basis of these profiles, we concluded that the posterior-most localization, which has been reported for over 20 different transcripts, is the only major localization pattern of maternal transcripts. Our data also showed that maternal factors establish only nine distinct patterns of zygotic gene expression at the 16-cell stage. Therefore, one of the main developmental functions of maternally supplied information is to establish these nine distinct expression patterns in the 16-cell embryo. The dynamics of transcriptomes in early-stage embryos provides a foundation for studying how maternal information starts the zygotic program.

  1. Planetary Embryo Bow Shocks as a Mechanism for Chondrule Formation

    NASA Astrophysics Data System (ADS)

    Mann, Christopher R.; Boley, Aaron C.; Morris, Melissa A.

    2016-02-01

    We use radiation hydrodynamics with direct particle integration to explore the feasibility of chondrule formation in planetary embryo bow shocks. The calculations presented here are used to explore the consequences of a Mars-size planetary embryo traveling on a moderately excited orbit through the dusty, early environment of the solar system. The embryo’s eccentric orbit produces a range of supersonic relative velocities between the embryo and the circularly orbiting gas and dust, prompting the formation of bow shocks. Temporary atmospheres around these embryos, which can be created via volatile outgassing and gas capture from the surrounding nebula, can non-trivially affect thermal profiles of solids entering the shock. We explore the thermal environment of solids that traverse the bow shock at different impact radii, the effects that planetoid atmospheres have on shock morphologies, and the stripping efficiency of planetoidal atmospheres in the presence of high relative winds. Simulations are run using adiabatic and radiative conditions, with multiple treatments for the local opacities. Shock speeds of 5, 6, and 7 km s-1 are explored. We find that a high-mass atmosphere and inefficient radiative conditions can produce peak temperatures and cooling rates that are consistent with the constraints set by chondrule furnace studies. For most conditions, the derived cooling rates are potentially too high to be consistent with chondrule formation.

  2. Sperm is epigenetically programmed to regulate gene transcription in embryos

    PubMed Central

    Teperek, Marta; Simeone, Angela; Gaggioli, Vincent; Miyamoto, Kei; Allen, George E.; Erkek, Serap; Kwon, Taejoon; Marcotte, Edward M.; Zegerman, Philip; Bradshaw, Charles R.; Peters, Antoine H.F.M.; Gurdon, John B.; Jullien, Jerome

    2016-01-01

    For a long time, it has been assumed that the only role of sperm at fertilization is to introduce the male genome into the egg. Recently, ideas have emerged that the epigenetic state of the sperm nucleus could influence transcription in the embryo. However, conflicting reports have challenged the existence of epigenetic marks on sperm genes, and there are no functional tests supporting the role of sperm epigenetic marking on embryonic gene expression. Here, we show that sperm is epigenetically programmed to regulate embryonic gene expression. By comparing the development of sperm- and spermatid-derived frog embryos, we show that the programming of sperm for successful development relates to its ability to regulate transcription of a set of developmentally important genes. During spermatid maturation into sperm, these genes lose H3K4me2/3 and retain H3K27me3 marks. Experimental removal of these epigenetic marks at fertilization de-regulates gene expression in the resulting embryos in a paternal chromatin-dependent manner. This demonstrates that epigenetic instructions delivered by the sperm at fertilization are required for correct regulation of gene expression in the future embryos. The epigenetic mechanisms of developmental programming revealed here are likely to relate to the mechanisms involved in transgenerational transmission of acquired traits. Understanding how parental experience can influence development of the progeny has broad potential for improving human health. PMID:27034506

  3. CULTIVATION OF CHICKEN POX VIRUS IN DEVELOPING CHICK EMBRYOS

    DTIC Science & Technology

    The virus of chicken pox adapts readily and multiplies in the chorio- allantoic membranes of a chick embryo. A virus which has undergone several...passages on chorioallantoic membrane causes macroscopic changes in it. The chicken pox virus possesses a hemagglutinating capacity.

  4. PLANETARY EMBRYO BOW SHOCKS AS A MECHANISM FOR CHONDRULE FORMATION

    SciTech Connect

    Mann, Christopher R.; Boley, Aaron C.; Morris, Melissa A.

    2016-02-20

    We use radiation hydrodynamics with direct particle integration to explore the feasibility of chondrule formation in planetary embryo bow shocks. The calculations presented here are used to explore the consequences of a Mars-size planetary embryo traveling on a moderately excited orbit through the dusty, early environment of the solar system. The embryo’s eccentric orbit produces a range of supersonic relative velocities between the embryo and the circularly orbiting gas and dust, prompting the formation of bow shocks. Temporary atmospheres around these embryos, which can be created via volatile outgassing and gas capture from the surrounding nebula, can non-trivially affect thermal profiles of solids entering the shock. We explore the thermal environment of solids that traverse the bow shock at different impact radii, the effects that planetoid atmospheres have on shock morphologies, and the stripping efficiency of planetoidal atmospheres in the presence of high relative winds. Simulations are run using adiabatic and radiative conditions, with multiple treatments for the local opacities. Shock speeds of 5, 6, and 7 km s{sup −1} are explored. We find that a high-mass atmosphere and inefficient radiative conditions can produce peak temperatures and cooling rates that are consistent with the constraints set by chondrule furnace studies. For most conditions, the derived cooling rates are potentially too high to be consistent with chondrule formation.

  5. Chromosome remodeling and differentiation of tetraploid embryos during preimplantation development.

    PubMed

    Park, Mi-Ryung; Lee, Ah-Reum; Bui, Hong-Thuy; Park, Chankyu; Park, Keun-Kyu; Cho, Ssang-Goo; Song, Hyuk; Kim, Jae-Hwan; Nguyen, Van Thuan; Kim, Jin-Hoi

    2011-07-01

    Although it is known that the tetraploid embryo contributes only to the placenta, the question of why tetraploid embryos differentiate into placenta remains unclear. To study the effect of electrofusion on the development of mouse tetraploid oocytes, mouse two-cell embryos were fused and cultured in vitro in Chatot-Ziomek-Bavister medium. After electrofusion, two chromosome sets from the tetraploid blastomere were individually duplicated before nuclear fusion. At 8-10 hr after electrofusion, each chromosome set was condensing and the nuclear membrane was breaking down. Around 12-14 hr after electrofusion, the two chromosome sets had combined together and had reached the second mitotic metaphase, at this point with 8n sets of chromosomes. Interestingly, we discovered that expression of OCT4, an inner cell mass cells biomarker, is lost by the tetraploid expanded blastocysts, but that CDX2, a trophectoderm cells biomarker, is strongly expressed at this stage. This observation provides evidence clarifying why tetraploid embryos contribute only to trophectoderm.

  6. Cardiac hypertrophy in chick embryos induced by hypothermia

    SciTech Connect

    Boehm, C.; Johnson, T.R.; Caston, J.D.; Przybylski, R.J.

    1987-01-01

    A decrease in incubation temperature from 38 to 32/sup 0/C elicits a decrease in chicken embryo size and weight with concomitant heart enlargement if done after day 10 of incubation. When assayed at day 18 of incubation with the hypothermia started on day 11 or 14, evidence is presented that the heart enlargement is an hypertrophy with no detectable hyperplasia. Supporting data are presented for various physical parameters showing increases in heart wet and dry weight, volume, area, wall thickness, and cell size. There was little difference in DNA content and nuclear (/sup 3/H)thymidine labeling index between hearts of control and hypothermic embryos. Hearts of hypothermic embryos showed a slight increase in water content and considerable increases in RNA, protein, and glycogen content per unit DNA. The average size of polysomes isolated from hypothermic hearts was larger than that of polysomes isolated from controls. Microscopic studies showed no obvious increase in amount of capillary beds, connective tissue, and myocardial cells. Annulate lamellae were found only in myocardial cells of hypothermic embryos in sparse amounts and low frequency but always associated with large deposits of glycogen.

  7. SYNTHESIS AND STORAGE OF MICROTUBULE PROTEINS BY SEA URCHIN EMBRYOS

    PubMed Central

    Raff, Rudolf A.; Greenhouse, Gerald; Gross, Kenneth W.; Gross, Paul R.

    1971-01-01

    Studies employing colchicine binding, precipitation with vinblastine sulfate, and acrylamide gel electrophoresis confirm earlier proposals that Arbacia punctulata and Lytechinus pictus eggs and embryos contain a store of microtubule proteins. Treatment of 150,000 g supernatants from sea urchin homogenates with vinblastine sulfate precipitates about 5% of the total soluble protein, and 75% of the colchicine-binding activity. Electrophoretic examination of the precipitate reveals two very prominent bands. These have migration rates identical to those of the A and B microtubule proteins of cilia. These proteins can be made radioactive at the 16 cell stage and at hatching by pulse labeling with tritiated amino acids. By labeling for 1 hr with leucine-3H in early cleavage, then culturing embryos in the presence of unlabeled leucine, removal of newly synthesized microtubule proteins from the soluble pool can be demonstrated. Incorporation of labeled amino acids into microtubule proteins is not affected by culturing embryos continuously in 20 µg/ml of actinomycin D. Microtubule proteins appear, therefore, to be synthesized on "maternal" messenger RNA. This provides the first protein encoded by stored or "masked" mRNA in sea urchin embryos to be identified. PMID:5165266

  8. Gamete and embryo-fetal origins of adult diseases.

    PubMed

    Monti, Manuela

    2016-08-10

    By putting together the most advanced evidences supporting the 'gamete and embryo-fetal origins of adult diseases' the two editors, Prof. He-Feng Huang and Prof. Jian-Zhong Sheng (Hangzhou, People's Republic of China) did a great workk.....

  9. Characteristics of sugar uptake by immature maize embryos

    SciTech Connect

    Griffith, S.M.; Jones, R.J.; Brenner, M.L.

    1986-04-01

    Characteristics of sugar uptake by immature maize embryos were determined in vitro utilizing a /sup 14/C-sugar solution incubation method. Hexose uptake rates were greater than those for sucrose, however, all showed biphasic kinetics. Glucose and fructose saturable components were evidence at <50 mM and sucrose at <5 mM. Chemical inhibitors (CCCP, DNP, NaCN, and PCMBS) and low temperature reduced sugar uptake. Sucrose influx was pH dependent while glucose was not. Embryos maintained a high sucrose to hexose ratio throughout development. At 25 days after pollination sucrose levels exceeded 200 mM while hexose levels remained below 5 mM. Glucose was rapidly converted to sucrose upon transport into the embryo. These circumstantial data indicate that sugar uptake by immature maize embryos is metabolically dependent and carrier mediated. Furthermore, sucrose transport appears to occur against its concentration gradient involving a H+/sucrose cotransport mechanism, while glucose influx is driven by its concentration gradient and subsequent metabolism.

  10. Immunocytochemistry of the amphibian embryo--from overview to ultrastructure.

    PubMed

    Kurth, Thomas

    2003-06-01

    Amphibian embryos are standard research objects to study pattern formation and morphogenesis. Due to their external development and robust nature, experimental manipulations such as microinjections or transplantations can be easily performed. However, most immunocytochemical approaches addressing the specific localization of proteins are hampered by the fragility of the large and yolky embryonic cells which render high resolution staining difficult. Immunocytochemical data are therefore often restricted to either overall patterns in whole embryo preparations or to immunofluorescent localization with limited resolution on sections. High resolution or ultrastructural protein localization data are rare and can be achieved only with time consuming procedures. Here, a comparative study of immunocytochemical methods suitable for light and electron microscopy using different kinds of plastic resins is presented. Three main approaches are described: preembedding staining of whole embryos, postembedding staining of ultrathin sections and preembedding staining of vibratome sections. All the procedures are designed to study protein expression in early amphibian embryos en gros as well as en detail and the described techniques are suitable to combine two or three levels of resolution on the very same biological specimen. Examples are presented and advantages and disadvantages of the different protocols are discussed.

  11. The VIRTUAL EMBRYO. A Computational Framework for Developmental Toxicity

    EPA Science Inventory

    EPA’s ‘Virtual Embryo Project’ (v-Embryo™) is focused on the predictive toxicology of children’s health and developmental defects following prenatal exposure to environmental chemicals. The research is motivated by scientific principles in systems biology as a framework for the g...

  12. Thermal effects in laser-assisted embryo hatching

    NASA Astrophysics Data System (ADS)

    Douglas-Hamilton, Diarmaid H.; Conia, Jerome D.

    2000-08-01

    Diode lasers [(lambda) equals 1480 nm] are used with in-vitro fertilization [IVF] as a promoter of embryo hatching. A focused laser beam is applied in vitro to form a channel in the zona pellucida (shell) of the pre-embryo. After transfer into the uterus, the embryo hatches: it extrudes itself through the channel and implants into the uterine wall. Laser-assisted hatching can result in improving implantation and pregnancy success rates. We present examples of zone pellucida ablation using animal models. In using the laser it is vital not to damage pre-embryo cells, e.g. by overheating. In order to define safe regimes we have derived some thermal side-effects of zona pellucida removal. The temperature profile in the beam and vicinity is predicted as function of laser pulse duration and power. In a crossed-beam experiment a HeNe laser probe detects the temperature-induced change in refractive index. We find that the diode laser beam produces superheated water approaching 200 C on the beam axis. Thermal histories during and following the laser pulse are given for regions in the neighborhood of the beam. We conclude that an optimum regime exists with pulse duration

  13. TRANSCRIPTIONAL RESPONSES OF MOUSE EMBRYO CULTURES EXPOSED TO BROMOCHLOROACETIC ACID

    EPA Science Inventory

    Transcriptional responses of mouse embryo cultures exposed to bromochloroacetic acid

    Edward D. Karoly?*, Judith E. Schmid* and E. Sidney Hunter III*
    ?Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina and *Reproductive Tox...

  14. MicroRNA processing machinery in the developing chick embryo.

    PubMed

    Carraco, Gil; Gonçalves, Ana N; Serra, Carlos; Andrade, Raquel P

    2014-11-01

    Gene expression regulation during embryo development is under strict regulation to ensure proper gene expression in both time and space. The involvement of microRNAs (miRNA) in early vertebrate development is documented and inactivation of different proteins involved in miRNA synthesis results in severe malformations or even arrests vertebrate embryo development. However, there is very limited information on when and in what tissues the genes encoding these proteins are expressed. Herein, we report a detailed characterization of the expression patterns of DROSHA, DGCR8, XPO5 and DICER1 in the developing chick embryo, from HH1 (when the egg is laid) to HH25 (5-days incubation), using whole mount in situ hybridization and cross-section analysis. We found that these genes are co-expressed in multiple tissues, mostly after stage HH4. Before early gastrulation DICER1 expression was never detected, suggesting the operation of a Dicer-independent pathway for miRNA synthesis. Our results support an important role for miRNAs in vertebrate embryo development and provide the necessary framework to unveil additional roles for these RNA processing proteins in development.

  15. Lipid transport to avian oocytes and to the developing embryo.

    PubMed

    Schneider, Wolfgang J

    2016-05-01

    Studies of receptor-mediated lipoprotein metabolic pathways in avian species have revealed that physiological intricacies of specific cell types are highly analogous to those in mammals. A prime example for the power of comparative studies across different animal kingdoms, elucidated in the chicken, is that the expression of different lipoprotein receptors in somatic cells and oocytes are the key to oocyte growth. In avian species, yolk precursor transport from the hen's liver to rapidly growing oocytes and the subsequent transfer of yolk nutrients via the yolk sac to the developing embryo are highly efficient processes. Oocytes grow from a diameter of 5 mm to 2.5-3 cm in only 7 days, and the yolk sac transfers nutrients from the yolk stored in the mature oocyte to the embryo within just 2 weeks. The underlying key transport mechanism is receptor-mediated endocytosis of macromolecules, i.e., of hepatically synthesized yolk precursors for oocyte growth, and of mature yolk components for embryo nutrition, respectively. Recently, the receptors involved, as well as the role of lipoprotein synthesis in the yolk sac have been identified. As outlined here, lipoprotein degradation/resynthesis cycles and the expression of lipoprotein receptors are not only coordinated with the establishment of the follicular architecture embedding the oocyte, but also with the generation of the yolk sac vasculature essential for nutrient transfer to the embryo.

  16. Spatial distribution of the Sm antigen in Drosophila early embryos.

    PubMed

    Ségalat, L; Lepesant, J A

    1992-01-01

    Anti-Sm antibodies recognize the major small nuclear RNA-protein particles (snRNPs) involved in pre-mRNA processing. The spatial distribution of the snRNPs has been investigated in Drosophila embryos up to the cellularization stage (cycle 14), using the Y12 anti-Sm antibody. Our results show that: 1) all or most of the Sm antigen is localized in the cytoplasm of the syncytial blastoderm until the 12th cycle of division, in both the nuclear and cytoplasmic compartments at cycle 13, and then in the nuclei at cycle 14 and later. This relocalization takes place when zygotic transcriptional activation occurs; 2) at the subcellular level, the Sm antigen localizes in a speckled pattern and in foci-like structures within the nucleus of Drosophila blastoderm embryos; 3) strikingly, some nuclei of embryos at the 14th cycle appear to contain more snRNPs than others. The position of these nuclei differs from one embryo to another, and their distribution does not resemble any known developmental pattern of Drosophila embryogenesis. We propose that random differences in snRNP concentration may serve as an epigenetic signal for stochastic events occurring during development.

  17. Twin Xenopus laevis embryos appearing from flattened eggs.

    PubMed

    Sato, Eiji

    2014-01-01

    Remarkable progress has recently been made in molecular biology of double axis formation in Xenopus laevis. Leaving aside, for the time being, the problem of the gene expressions regulating Xenopus laevis development, here I show that pulse treatment could induce formation of a secondary axis in a fertilized Xenopus laevis egg. At 3 min after insemination, metal oxides were added to Xenopus fertilized eggs, and then twin embryos appeared. Zirconium oxide (ZrO2) was the most effective metal oxide for producing twin embryos. ZrO2 was added to the fertilized eggs, and 30 sec later, the eggs were dejellied with cysteine solution and washed within 7 min after insemination. The fertilized eggs began flattening at around 15 min after insemination. When the degree of flattening (the vertical length of the egg divided by the horizontal length) of the eggs at the 16- and 32-cell stages became less than 0.4 degrees, production of twin embryos occurred. Many flattened eggs at less than 0.4 degrees formed twin embryos. The third cleavage of eggs treated with metal oxides was meridional, while the normal third cleavage was horizontal.

  18. Desiccation tolerance during different desiccation strategies in A. angustifolia embryos

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Brazilian pine (Araucaria angustifolia) is native to the Atlantic Rainforest of Brazil and is an endangered species. The mature seeds are recalcitrant and have large embryos (about 2.5 cm in length) that contain more than 1 g H2O.g dry mass (dm)-1. Successful cryopreservation requires reduction of ...

  19. METHOXYCHLOR ACCELERATES EMBRYO TRANSPORT THROUGH THE RAT REPRODUCTIVE TRACT

    EPA Science Inventory

    The estrogenic pesticide methoxychlor (MXC) is known to reduce implantation, and, in our previous work, this reduction has been attributed to a direct effect on uterine function. The present study was designed to investigate the effect of MXC on embryo transport rate, another phe...

  20. Perflurooctanoic Acid Induces Developmental Cardiotoxicity in Chicken Embryos and Hatchlings

    EPA Science Inventory

    Perfluorooctanoic acid (PFOA) is a widespread environmental contaminant that is detectable in serum of the general U.S. population. PFOA is a known developmental toxicant that induces mortality in mammalian embryos and is thought to induce toxicity via interaction with the peroxi...

  1. Pathology of the human embryo and previable fetus

    SciTech Connect

    Kalousek, D.K. ); Fitch, N.; Paradice, B.

    1990-01-01

    Topics covered in this book include a general review of normal embryonic and fetal development; abortion and the basic approach to the examination of aborted embryos and fetuses; and pathologic findings detected on examination of products of conception. The authors illustrate specific morphologic lesions and the variable expression of genetic syndromes in the embryonic and fetal periods.

  2. Proteomic responses of sea urchin embryos to stressful ultraviolet radiation.

    PubMed

    Adams, N L; Campanale, J P; Foltz, K R

    2012-11-01

    Solar ultraviolet radiation (UVR, 290-400 nm) penetrates into seawater and can harm shallow-dwelling and planktonic marine organisms. Studies dating back to the 1930s revealed that echinoids, especially sea urchin embryos, are powerful models for deciphering the effects of UVR on embryonic development and how embryos defend themselves against UV-induced damage. In addition to providing a large number of synchronously developing embryos amenable to cellular, biochemical, molecular, and single-cell analyses, the purple sea urchin, Strongylocentrotus purpuratus, also offers an annotated genome. Together, these aspects allow for the in-depth study of molecular and biochemical signatures of UVR stress. Here, we review the effects of UVR on embryonic development, focusing on the early-cleavage stages, and begin to integrate data regarding single-protein responses with comprehensive proteomic assessments. Proteomic studies reveal changes in levels of post-translational modifications to proteins that respond to UVR, and identify proteins that can then be interrogated as putative targets or components of stress-response pathways. These responsive proteins are distributed among systems upon which targeted studies can now begin to be mapped. Post-transcriptional and translational controls may provide early embryos with a rapid, fine-tuned response to stress during early stages, especially during pre-blastula stages that rely primarily on maternally derived defenses rather than on responses through zygotic gene transcription.

  3. Synchrotron X-ray tomographic microscopy of fossil embryos.

    PubMed

    Donoghue, Philip C J; Bengtson, Stefan; Dong, Xi-ping; Gostling, Neil J; Huldtgren, Therese; Cunningham, John A; Yin, Chongyu; Yue, Zhao; Peng, Fan; Stampanoni, Marco

    2006-08-10

    Fossilized embryos from the late Neoproterozoic and earliest Phanerozoic have caused much excitement because they preserve the earliest stages of embryology of animals that represent the initial diversification of metazoans. However, the potential of this material has not been fully realized because of reliance on traditional, non-destructive methods that allow analysis of exposed surfaces only, and destructive methods that preserve only a single two-dimensional view of the interior of the specimen. Here, we have applied synchrotron-radiation X-ray tomographic microscopy (SRXTM), obtaining complete three-dimensional recordings at submicrometre resolution. The embryos are preserved by early diagenetic impregnation and encrustation with calcium phosphate, and differences in X-ray attenuation provide information about the distribution of these two diagenetic phases. Three-dimensional visualization of blastomere arrangement and diagenetic cement in cleavage embryos resolves outstanding questions about their nature, including the identity of the columnar blastomeres. The anterior and posterior anatomy of embryos of the bilaterian worm-like Markuelia confirms its position as a scalidophoran, providing new insights into body-plan assembly among constituent phyla. The structure of the developing germ band in another bilaterian, Pseudooides, indicates a unique mode of germ-band development. SRXTM provides a method of non-invasive analysis that rivals the resolution achieved even by destructive methods, probing the very limits of fossilization and providing insight into embryology during the emergence of metazoan phyla.

  4. Dignity, marriage and embryo adoption: a look at Dignitas Personae.

    PubMed

    Murphy, Timothy F

    2011-12-01

    The Catholic Church's 2008 Dignitas Personae discusses the moral implications of respecting the dignity of all human beings, regardless of the stage of development. In that text, the Vatican's Congregation for the Doctrine of the Faith argues that respect for this dignity is incompatible with the conception of embryos outside marriage as well as assisted reproduction treatments and certain kinds of human embryonic research. Not only that, but the Congregation also rejects efforts at embryo adoption. As a matter of secular moral philosophy, this view of dignity is disputable and this article shows how an alternate view of dignity--one that depends on interests as against status--serves as a better foundation for decisions about ways in which to help people have children. This view of dignity is entirely compatible with a wide array of assisted reproduction treatments and research and is compatible with the conception of embryos for single parents or opposite-sex couples looking to have children. Using its notion of human dignity, the Congregation makes a case against embryo adoption, but that case is unconvincing given the permissible exercise of individual conscience and the presumptive importance of rescuing human lives where they can be rescued.

  5. AGL15, a MADS domain protein expressed in developing embryos.

    PubMed Central

    Heck, G R; Perry, S E; Nichols, K W; Fernandez, D E

    1995-01-01

    To extend our knowledge of genes expressed during early embryogenesis, the differential display technique was used to identify and isolate mRNA sequences that accumulate preferentially in young Brassica napus embryos. One of these genes encodes a new member of the MADS domain family of regulatory proteins; it has been designated AGL15 (for AGAMOUS-like). AGL15 shows a novel pattern of expression that is distinct from those of previously characterized family members. RNA gel blot analyses and in situ hybridization techniques were used to demonstrate that AGL15 mRNA accumulated primarily in the embryo and was present in all embryonic tissues, beginning at least as early as late globular stage in B. napus. Genomic and cDNA clones corresponding to two AGL15 genes from B. napus and the homologous single-copy gene from Arabidopsis, which is located on chromosome 5, were isolated and analyzed. Antibodies prepared against overexpressed Brassica AGL15 lacking the conserved MADS domain were used to probe immunoblots, and AGL15-related proteins were found in embryos of a variety of angiosperms, including plants as distantly related as maize. Based on these data, we suggest that AGL15 is likely to be an important component of the regulatory circuitry directing seed-specific processes in the developing embryo. PMID:7549483

  6. Antioxidant rescue of selenomethionine-induced teratogenesis in zebrafish embryos

    PubMed Central

    Arnold, M.C.; Forte, J.E.; Osterberg, J.S.; Di Giulio, R.T.

    2015-01-01

    Selenium (Se) is an essential micronutrient that can be found at toxic concentrations in surface waters contaminated by runoff from agriculture and coal mining. Zebrafish (Danio rerio) embryos were exposed to aqueous Se in the form of selenate, selenite, and L-selenomethionine (SeMet) in an attempt to determine if oxidative stress plays a role in selenium embryo toxicity. Selenate and selenite exposure did not induce embryo deformities (lordosis and craniofacial malformation). L-selenomethionine, however, induced significantly higher deformity rates at 100 μg/L compared to controls. SeMet exposure induced a dose-dependent increase in the catalytic subunit of glutamate-cysteine ligase (gclc) and reached an 11.7-fold increase at 100 μg/L. SeMet exposure also reduced concentrations of TGSH, RGSH, and the TGSH:GSSG ratio. Pretreatment with 100 μM N-acetylcysteine (NAC) significantly reduced deformities in the zebrafish embryos secondarily treated with 400 μg/L SeMet from approximately 50% to 10% as well as rescued all three of the significant glutathione level differences seen with SeMet alone. Selenite exposure induced a 6.6-fold increase in expression of the glutathione-S-transferase pi class 2 (gstp2) gene, which is involved in xenobiotic transformation and possibly oxidative stress. These results suggest that aqueous exposure to SeMet can induce significant embryonic teratogenesis in zebrafish that are at least partially attributed to oxidative stress. PMID:26498942

  7. Juvenile hormone and allatostatins in the German cockroach embryo.

    PubMed

    Maestro, José L; Pascual, Núria; Treiblmayr, Karl; Lozano, Jesús; Bellés, Xavier

    2010-09-01

    Levels of juvenile hormone III (JH), FGLamide allatostatin peptides (ASTs), ASTs precursor (preproAST) mRNA and methyl farnesoate epoxidase (CYP15A1) mRNA were measured in embryos of the cockroach Blattella germanica. JH starts to rise just after dorsal closure, reaches maximal levels between 60% and 80% of embryogenesis, and decrease subsequently to undetectable levels. ASTs show low levels during the first two thirds of embryogenesis, increase thereafter and maintain high levels until hatching. PreproAST mRNA shows quite high levels during the two days following oviposition, thus behaving as a maternal transcript, the levels then become very low until mid embryogenesis, and increase afterwards, peaking towards the end of embryo development. CYP15A1 transcripts were detected around 25% embryogenesis and the levels tended to increase through embryogenesis, although differences amongst the days studied were not statistically significant. The opposite patterns of JH and AST towards the end of embryo development, along with the detection of AST immunoreactivity in corpora allata from late embryos, suggest that JH decline is caused by the increase of AST. Moreover, the uncorrelated patterns of JH concentration and CYP15A1 mRNA levels suggest that CYP15A1 expression does not modulate JH production.

  8. Zygotic genome activation in isogenic and hybrid plant embryos.

    PubMed

    Del Toro-De León, Gerardo; Lepe-Soltero, Daniel; Gillmor, C Stewart

    2016-02-01

    Zygotic genome activation (ZGA) is the onset of large-scale transcription that occurs after fertilization. In animal embryos, ZGA occurs after a period of transcriptional quiescence that varies between species. In plants, the timing of ZGA may also vary between species, and may or may not occur in a parent-of-origin dependent manner: some studies have shown a maternal bias in mRNA transcripts and gene activity in early embryogenesis, while other experiments have found the contribution of maternal and paternal genomes to be equal. In order to differentiate between maternal and paternal mRNAs, RNA sequencing studies of ZGA in plants have used embryos hybrid for polymorphic accessions. A recent genetic assay in Arabidopsis demonstrated significant variation in paternal allele activity between some hybrid combinations and isogenic embryos, as well as between different hybrid combinations, suggesting a possible source for conflicting results obtained by various experiments on paternal genome activation. We review recent literature on paternal genome activation studies in the zygote in both isogenic and hybrid embryos, and discuss possible explanations for the effects of hybridization on gene expression in early embryogenesis in plants.

  9. Structured illumination fluorescence correlation spectroscopy for velocimetry in Zebrafish embryos

    NASA Astrophysics Data System (ADS)

    Pozzi, Paolo; Rossetti, Leone; Sironi, Laura; Freddi, Stefano; D'Alfonso, Laura; Caccia, Michele; Bouzin, Margaux; Collini, Maddalena; Chirico, Giuseppe

    2013-02-01

    The vascular system of Zebrafish embryos is studied by means of Fluorescence Correlation and Image Correlation Spectroscopy. The long term project addresses biologically relevant issues concerning vasculogenesis and cardiogenesis and in particular mechanical interaction between blood flow and endothelial cells. To this purpose we use Zebrafish as a model system since the transparency of its embryos facilitates morphological observation of internal organs in-vivo. The correlation analysis provides quantitative characterization of fluxes in blood vessels in vivo. We have pursued and compared two complementary routes. In a first one we developed a two-spots two-photon setup in which the spots are spaced at adjustable micron-size distances (1-40 μm) along a vessel and the endogenous (autofluorescence) or exogenous (dsRed transgenic erythrocytes) signal is captured with an EM-CCD and cross-correlated. In this way we are able to follow the morphology of the Zebrafish embryo, simultaneously measure the heart pulsation, the velocity of red cells and of small plasma proteins. These data are compared to those obtained by image correlations on Zebrafish vessels. The two methods allows to characterize the motion of plasma fluids and erythrocytes in healthy Zebrafish embryos to be compared in the future to pathogenic ones.

  10. Transcriptomic Analysis of the Porcine Endometrium during Embryo Implantation

    PubMed Central

    Lin, Haichao; Wang, Huaizhong; Wang, Yanping; Liu, Chang; Wang, Cheng; Guo, Jianfeng

    2015-01-01

    In pigs, successful embryo implantation is an important guarantee for producing litter size, and early embryonic loss occurring on day 12–30 of gestation critically affects the potential litter size. The implantation process is regulated by the expression of numerous genes, so comprehensive analysis of the endometrium is necessary. In this study, RNA sequencing (RNA-Seq) technology is used to analyze endometrial tissues during early pregnancy. We investigated the changes of gene expression between three stages (day 12, 18, and 25) by multiple comparisons. There were 1557, 8951, and 2345 differentially expressed genes (DEGs) revealed between the different periods of implantation. We selected several genes for validation by the use of quantitative real-time RT-PCR. Bioinformatic analysis of differentially expressed genes in the endometrium revealed a number of biological processes and pathways potentially involved in embryo implantation in the pig, most noticeably cell proliferation, regulation of immune response, interaction of cytokine-cytokine receptors, and cell adhesion. These results showed that specific gene expression patterns reflect the different functions of the endometrium in three stages (maternal recognition, conceptus attachment, and embryo implantation). This study identified comprehensive transcriptomic profile in the porcine endometrium and thus could be a foundation for targeted studies of genes and pathways potentially involved in abnormal endometrial receptivity and embryo loss in early pregnancy. PMID:26703736

  11. PFOA INDUCES DYSMORPHOGENESIS IN MOUSE WHOLE EMBRYO CULTURE

    EPA Science Inventory

    PFOA Induces Dysmorphogenesis In Mouse Whole Embryo Culture.

    MR Blanton1, JM Padowski2, ES Hunter1, JM Rogers1, and C Lau1. 1Reproductive Toxicology Division, NHEERL, ORD, US EPA, RTP, NC, USA. 2Curriculum in Toxicology, UNC, Chapel Hill, NC, USA

    Perfluorooctanoa...

  12. Use of infrared imaging for investigation of chicken embryo development

    NASA Astrophysics Data System (ADS)

    Frye, Ryan A.; Hsieh, Sheng-Jen; Girón Palomares, José Benjamín D.

    2011-05-01

    The focus of this study is two-fold: first, to investigate the feasibility of thermal imaging for characterizing the development of chicken embryos; and second, to compare the effects of photo periods of 11 hours of light followed by 11 hours of darkness (11-11) versus 24 hours of darkness (24 dark) during the incubation cycle on embryo development. Previous reported work has used invasive methods, such as ultrasound, tomography, and MRI to study chicken embryos with some success. However, very little work has been reported on use of thermography, which is a non-invasive method. Results suggest that use of a cooling-heating-cooling cycle can reveal the anatomy of chicken embryos. A statistical comparison of image data from the two photo periods found no difference in the average cooling rates. However, the 11-11 group of eggs did hatch earlier overall than 24-dark group. Of the hatched eggs, all the chickens from the 24-dark group appeared to be in normal physical condition. However, two of the chickens from the 11-11 group appeared to have leg weakness shortly after hatching. Of these, one fully recovered the next day and the second remains the same after two days of observation. In addition, the second chicken took about 48 hours to fully emerge from its shell.

  13. Development of bovine embryos derived from reproductive techniques.

    PubMed

    Alberto, Míryan L V; Meirelles, Flavio V; Perecin, Felipe; Ambrósio, Carlos E; Favaron, Phelipe O; Franciolli, André L R; Mess, Andrea M; Dos Santos, José M; Rici, Rose E G; Bertolini, Marcelo; Miglino, Maria A

    2013-01-01

    Assisted reproduction techniques have improved agricultural breeding in the bovine. However, important development steps may differ from the situation in vivo and there is a high mortality rate during the first trimester of gestation. To better understand these events, we investigated the development of embryos and fetal membranes following fixed-time AI (FTAI), IVF and nuclear transfer (NT). The onset of yolk-sac development was not normal in cloned embryos. Later steps differed from conditions in vivo in all three groups; the yolk-sac was yellowish and juxtaposed with the amniotic membrane. Vascularisation of the chorioallantoic membrane was relatively late and low in NT gestations, but normal in the others. The overall development of the embryos was normal, as indicated by morphology and regression analysis of growth rate. However, NT conceptuses were significantly smaller, with the livers in some embryos occupying the abdominal cavity and others exhibiting heart abnormalities. In conclusion, the yolk-sac and the cardiovascular system seem to be vulnerable to morphogenetic alterations. Future studies will focus on gene expression and early vascularisation processes to investigate whether these changes may be responsible for the high incidence of intrauterine mortality, especially in clones.

  14. Comparative phosphoproteomics of zebrafish Fyn/Yes morpholino knockdown embryos.

    PubMed

    Lemeer, Simone; Jopling, Chris; Gouw, Joost; Mohammed, Shabaz; Heck, Albert J R; Slijper, Monique; den Hertog, Jeroen

    2008-11-01

    The coordinated movement of cells is indispensable for normal vertebrate gastrulation. Several important players and signaling pathways have been identified in convergence and extension (CE) cell movements during gastrulation, including non-canonical Wnt signaling. Fyn and Yes, members of the Src family of kinases, are key regulators of CE movements as well. Here we investigated signaling pathways in early development by comparison of the phosphoproteome of wild type zebrafish embryos with Fyn/Yes knockdown embryos that display specific CE cell movement defects. For quantitation we used differential stable isotope labeling by reductive amination of peptides. Equal amounts of labeled peptides from wild type and Fyn/Yes knockdown embryos were mixed and analyzed by on-line reversed phase TiO(2)-reversed phase LC-MS/MS. Phosphorylated and non-phosphorylated peptides were quantified, and significant changes in protein expression and/or phosphorylation were detected. We identified 348 phosphoproteins of which 69 showed a decrease in phosphorylation in Fyn/Yes knockdown embryos and 72 showed an increase in phosphorylation. Among these phosphoproteins were known regulators of cell movements, including Adducin and PDLIM5. Our results indicate that quantitative phosphoproteomics combined with morpholino-mediated knockdowns can be used to identify novel signaling pathways that act in zebrafish development in vivo.

  15. Delayed insemination results in embryo mortality in a brooding ascidian.

    PubMed

    Stewart-Savage, J; Phillippi, A; Yund, P O

    2001-08-01

    We explored the effects of temporal variation in sperm availability on fertilization and subsequent larval development in the colonial ascidian Botryllus schlosseri, a brooding hermaphrodite that has a sexual cycle linked to an asexual zooid replacement cycle. We developed a method to quantify the timing of events early in this cycle, and then isolated colonies before the start of the cycle and inseminated them at various times. Colony-wide fertilization levels (assayed by early cleavage) increased from zero to 100% during the period when the siphons of a new generation of zooids were first opening, and remained high for 24 h before slowly declining over the next 48 h. Because embryos are brooded until just before the zooids degenerate at the end of a cycle, delayed fertilization might also affect whether embryos can complete development within the cycle. Consequently, we also determined the effect of delayed insemination on successful embryo development through larval release and metamorphosis. When fertilization was delayed beyond the completion of siphon opening, there was an exponential decline in the percentage of eggs that ultimately produced a metamorphosed larva at the end of the cycle. Thus, even though the majority of oocytes can be fertilized when insemination is delayed for up to 48 h, the resulting embryos cannot complete development before the brooding zooids degenerate.

  16. [Isolation and cultivation of goat embryo stem cells].

    PubMed

    Yan, Long; Lei, Lei; Yang, Chunrong; Gao, Zhimin; Lei, Anmin; Ma, Xiaoling; Dou, Zhongying

    2008-09-01

    Morulaes and blastocysts obtained from Guanzhong dairy goats 6-7 days after mating were treated with whole embryo cultivaton, enzymatic digestion and immunosurgery separately. The goat embryonic stem cells (ESC) were isolated and cultured on a feeder layer of mitomycin-inactivated mouse embryo fibroblasts (MEF). The characteristics of goat ESCs were analyzed by immunohistochemisty, RT-PCR and inducing differentiation in vitro. The results indicated that the embryos were easier to attach the culture dish and form primary colonies with whole embryo method. There were colonies that maintained undifferentiated for 18 passages. The ESCs expressed the protein of Nanog, Oct4 and SSEA-3, whereas the protein of SSEA-4 was absent and the protein of SSEA-1 was weakly expressed. In addition, the genes of Nanog, Oct4, TERT and CD117 were expressed in goat ESCs. The cells also could differentiate to myocardial cells when induced in vitro by DMSO. These results suggest that the goat ESCs have characteristics of ESCs.

  17. Deleterious actions of gossypol on bovine spermatozoa, oocytes, and embryos.

    PubMed

    Brocas, C; Rivera, R M; Paula-Lopes, F F; McDowell, L R; Calhoun, M C; Staples, C R; Wilkinson, N S; Boning, A J; Chenoweth, P J; Hansen, P J

    1997-10-01

    Gossypol (50 and 100 micrograms/ml) decreased the percentage of sperm that completed the swim-up procedure. This effect was not blocked by glutathione monoethyl ester. Cleavage rates were not different between oocytes inseminated with gossypol-treated spermatozoa (10 or 50 micrograms/ml) and oocytes inseminated with control spermatozoa. Development to the blastocyst stage at Day 7 after insemination was reduced when spermatozoa treated with 50 micrograms/ml gossypol were used for fertilization. Gossypol toxicity was evident in cows fed cottonseed meal because erythrocyte fragility was greater than for control cows. However, there were no differences between cottonseed meal and control groups in number of oocytes collected per cow, cleavage rate after in vitro maturation and fertilization, or the proportion of oocytes or embryos that developed to blastocysts. Similarly, exposure of oocytes to 2.5-10 micrograms/ml gossypol during in vitro maturation did not affect cleavage rates or subsequent development. In contrast, addition of 10 micrograms/ml gossypol to embryos reduced cleavage rate. Moreover, development of cleaved embryos was reduced by culture with 5 or 10 micrograms/ml gossypol and tended to be reduced by 2.5 micrograms/ml gossypol. In conclusion, bovine gametes are resistant to gossypol at concentrations similar to those in blood of cows fed cottonseed meal. In contrast, the developing embryo is sensitive to gossypol.

  18. Lipid transport to avian oocytes and to the developing embryo

    PubMed Central

    Schneider, Wolfgang J.

    2016-01-01

    Abstract Studies of receptor-mediated lipoprotein metabolic pathways in avian species have revealed that physiological intricacies of specific cell types are highly analogous to those in mammals. A prime example for the power of comparative studies across different animal kingdoms, elucidated in the chicken, is that the expression of different lipoprotein receptors in somatic cells and oocytes are the key to oocyte growth. In avian species, yolk precursor transport from the hen's liver to rapidly growing oocytes and the subsequent transfer of yolk nutrients via the yolk sac to the developing embryo are highly efficient processes. Oocytes grow from a diameter of 5 mm to 2.5-3 cm in only 7 days, and the yolk sac transfers nutrients from the yolk stored in the mature oocyte to the embryo within just 2 weeks. The underlying key transport mechanism is receptor-mediated endocytosis of macromolecules, i.e., of hepatically synthesized yolk precursors for oocyte growth, and of mature yolk components for embryo nutrition, respectively. Recently, the receptors involved, as well as the role of lipoprotein synthesis in the yolk sac have been identified. As outlined here, lipoprotein degradation/resynthesis cycles and the expression of lipoprotein receptors are not only coordinated with the establishment of the follicular architecture embedding the oocyte, but also with the generation of the yolk sac vasculature essential for nutrient transfer to the embryo. PMID:26585559

  19. The Syncytial Drosophila Embryo as a Mechanically Excitable Medium

    PubMed Central

    Idema, Timon; Dubuis, Julien O.; Kang, Louis; Manning, M. Lisa; Nelson, Philip C.; Lubensky, Tom C.; Liu, Andrea J.

    2013-01-01

    Mitosis in the early syncytial Drosophila embryo is highly correlated in space and time, as manifested in mitotic wavefronts that propagate across the embryo. In this paper we investigate the idea that the embryo can be considered a mechanically-excitable medium, and that mitotic wavefronts can be understood as nonlinear wavefronts that propagate through this medium. We study the wavefronts via both image analysis of confocal microscopy videos and theoretical models. We find that the mitotic waves travel across the embryo at a well-defined speed that decreases with replication cycle. We find two markers of the wavefront in each cycle, corresponding to the onsets of metaphase and anaphase. Each of these onsets is followed by displacements of the nuclei that obey the same wavefront pattern. To understand the mitotic wavefronts theoretically we analyze wavefront propagation in excitable media. We study two classes of models, one with biochemical signaling and one with mechanical signaling. We find that the dependence of wavefront speed on cycle number is most naturally explained by mechanical signaling, and that the entire process suggests a scenario in which biochemical and mechanical signaling are coupled. PMID:24204774

  20. Poisonous plants: Effects on embryo and fetal development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The impact of natural toxins from poisonous plants on the embryo, fetus, and neonate are dramatic and economically significant to livestock producers worldwide. In livestock, reproductive success is the single most important economic multiplier for livestock producers in the U.S. followed by carcas...

  1. Short latency vestibular evoked potentials in the chicken embryo

    NASA Technical Reports Server (NTRS)

    Jones, S. M.; Jones, T. A.

    1996-01-01

    Electrophysiological responses to pulsed linear acceleration stimuli were recorded in chicken embryos incubated for 19 or 20 days (E19/E20). Responses occurred within the first 16 ms following the stimulus onset. The evoked potentials disappeared following bilateral labyrinthectomy, but persisted following cochlear destruction alone, thus demonstrating that the responses were vestibular. Approximately 8 to 10 response peaks could be identified. The first 4 positive and corresponding negative components (early peaks with latencies < 6.0 ms) were scored and latencies and amplitudes quantified. Vestibular response latencies were significantly longer (P < 0.01) and amplitudes significantly smaller (P < 0.001) than those observed in 2-week-old birds. Mean response threshold for anesthetized embryos was -15.9dBre 1.0 g/ms, which was significantly higher (P < 0.03) than those observed in 2-week-old birds (-23.0dBre 1.0 g/ms). Latency/intensity functions (that is, slopes) were not significantly different between embryos and 2-week-old animals, but amplitude/intensity functions for embryos were significantly shallower than those for 2-week-old birds (P < 0.001). We presume that these differences reflect the refinement of sensory function that occurs following 19 to 20 days of incubation. The recording of vestibular evoked potentials provides an objective, direct and noninvasive measure of peripheral vestibular function in the embryo and, as such, the method shows promise as an investigative tool. The results of the present study form the definitive basis for using vestibular evoked potentials in the detailed study of avian vestibular ontogeny and factors that may influence it.

  2. Pollination and embryo development in Brassica rapa L. in microgravity

    NASA Technical Reports Server (NTRS)

    Kuang, A.; Popova, A.; Xiao, Y.; Musgrave, M. E.

    2000-01-01

    Plant reproduction under spaceflight conditions has been problematic in the past. In order to determine what aspect of reproductive development is affected by microgravity, we studied pollination and embryo development in Brassica rapa L. during 16 d in microgravity on the space shuttle (STS-87). Brassica is self-incompatible and requires mechanical transfer of pollen. Short-duration access to microgravity during parabolic flights on the KC-135A aircraft was used initially to confirm that equal numbers of pollen grains could be collected and transferred in the absence of gravity. Brassica was grown in the Plant Growth Facility flight hardware as follows. Three chambers each contained six plants that were 13 d old at launch. As these plants flowered, thin colored tape was used to indicate the date of hand pollination, resulting in silique populations aged 8-15 d postpollination at the end of the 16-d mission. The remaining three chambers contained dry seeds that germinated on orbit to produce 14-d-old plants just beginning to flower at the time of landing. Pollen produced by these plants had comparable viability (93%) with that produced in the 2-d-delayed ground control. Matched-age siliques yielded embryos of equivalent developmental stage in the spaceflight and ground control treatments. Carbohydrate and protein storage reserves in the embryos, assessed by cytochemical localization, were also comparable. In the spaceflight material, growth and development by embryos rescued from siliques 15 d after pollination lagged behind the ground controls by 12 d; however, in the subsequent generation, no differences between the two treatments were found. The results demonstrate that while no stage of reproductive development in Brassica is absolutely dependent upon gravity, lower embryo quality may result following development in microgravity.

  3. Factors affecting the cryosurvival of mouse two-cell embryos.

    PubMed

    Critser, J K; Arneson, B W; Aaker, D V; Huse-Benda, A R; Ball, G D

    1988-01-01

    A series of 4 experiments was conducted to examine factors affecting the survival of frozen-thawed 2-cell mouse embryos. Rapid addition of 1.5 M-DMSO (20 min equilibration at 25 degrees C) and immediate, rapid removal using 0.5 M-sucrose did not alter the frequency (mean +/- s.e.m.) of blastocyst development in vitro when compared to untreated controls (90.5 +/- 2.7% vs 95.3 +/- 2.8%). There was an interaction between the temperature at which slow cooling was terminated and thawing rate. Termination of slow cooling (-0.3 degrees C/min) at -40 degrees C with subsequent rapid thawing (approximately 1500 degrees C/min) resulted in a lower frequency of blastocyst development than did termination of slow cooling at -80 degrees C with subsequent slow thawing (+8 degrees C/min) (36.8 +/- 5.6% vs 63.9 +/- 5.7%). When slow cooling was terminated between -40 and -60 degrees C, higher survival rates were achieved with rapid thawing. When slow cooling was terminated below -60 degrees C, higher survival rates were obtained with slow thawing rates. In these comparisons absolute survival rates were highest among embryos cooled below -60 degrees C and thawed slowly. However, when slow cooling was terminated at -32 degrees C, with subsequent rapid warming, survival rates were not different from those obtained when embryos were cooled to -80 degrees C and thawed slowly (52.4 +/- 9.5%, 59.5 +/- 8.6%). These results suggest that optimal cryosurvival rates may be obtained from 2-cell mouse embryos by a rapid or slow thawing procedure, as has been found for mouse preimplantation embryos at later stages.(ABSTRACT TRUNCATED AT 250 WORDS)

  4. [Cryopreservation study on seeds and embryos in Dalbergia odorifera].

    PubMed

    Zeng, Lin; He, Ming-Jun; Chen, Kui; Wei, Jian-He

    2014-06-01

    The mature seeds and excised embryos of Dalbergia odorifera were used as materials to study the effect of moisture content on their survival, as well as the effect of rapid freezing and vitrification freezing method on seeds and in vitro embryos cryopreservation. The results showed that the germination rate and vigor decreased from 82.67%, 85% to 18.35%, 25% respectively, when the seed moisture content decreased from 15.04% to 8.14%; and the germination rate decreased from 82.67% to 37.50%, 25.37% respectively by vitrification freezing method and rapid freezing method, when the seed moisture content decreased from 15.04% to 9.37%. Among all the moisture content gradient, 12.35% moisture reached the maximal germination rate, which were 63.58% and 50.45% respectively by vitrification freezing and rapid freezing; and when the embryo moisture content was 26.32%, the germination rate decreased from 95.67% to 58.31% and 33.82% respectively by vitrification freezing and rapid freezing. And when the moisture content was in the range of 14.17% -21.34%, the germination rate was a bit of decrease. The experiment results showed that the optimum conditions of seed cryopreservation were: moisture content 12.35%, vitrification freezing; and the optimum conditions of in vitro embryo cryopreservation were: moisture 15.04%, vitrification freezing. In conclusion, the effects of moisture content on germination rate after cryopreservation in D. odorifera seeds and embryo were significant, and vitrification freezing method is much better than rapid freezing method.

  5. Transcriptome analysis of zebrafish embryos exposed to deltamethrin.

    PubMed

    Chueh, Tsung-Cheng; Hsu, Li-Sung; Kao, Chin-Ming; Hsu, Tung-Wei; Liao, Hung-Yu; Wang, Kuan-Yi; Chen, Ssu Ching

    2016-10-27

    Deltamethrin (DTM), a type II pyrethroid, is one of the most commonly used insecticides. The increased use of pyrethroid leads to potential adverse effects, particularly in sensitive populations such as children and pregnant women. None of the related studies was focused on the transcriptome responses in zebrafish embryos after treatment with DTM; therefore, RNA-seq, a high-throughput method, was performed to analyze the global expression of differential expressed genes (DEGs) in zebrafish embryos treated with DTM (40 and 80 μg/L) from fertilization to 48 h postfertilization (hpf) as compared with that in the control group (without DTM treatment). Two cDNA libraries were generated from treated embryos and one cDNA library from nontreated embryos, respectively. Over 92% of reads mapped to the reference in these three libraries. It was observed that many differential genes were expressed in comparison with embryos before and after DTM. The 20 most differentially expressed upregulated or downregulated genes were majorly involved in the signaling transduction. Validation of selected nine genes expression using qRT-PCR confirmed RNA-seq results. The transcriptome sequences were further subjected to gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, showing G-protein-coupled receptor signaling pathway and neuroactive ligand-receptor interaction, respectively, were most enriched. The data from this study contributed to a better understanding of the potential consequences of fish exposed to DTM, to an evaluation of the potential threat of DTM to fish populations in aquatic environments. © 2016 Wiley Periodicals, Inc. Environ Toxicol, 2016.

  6. Identification of emergent motion compartments in the amniote embryo

    PubMed Central

    Loganathan, Rajprasad; Little, Charles D; Joshi, Pranav; Filla, Michael B; Cheuvront, Tracey J; Lansford, Rusty; Rongish, Brenda J

    2014-01-01

    Abstract The tissue scale deformations (≥1mm) required to form an amniote embryo are poorly understood. Here, we studied ∼400 μm-sized explant units from gastrulating quail embryos. The explants deformed in a reproducible manner when grown using a novel vitelline membrane-based culture method. Time-lapse recordings of latent embryonic motion patterns were analyzed after disk-shaped tissue explants were excised from three specific regions near the primitive streak: 1) anterolateral epiblast, 2) posterolateral epiblast, and 3) the avian organizer (Hensen's node). The explants were cultured for 8 hours—an interval equivalent to gastrulation. Both the anterolateral and the posterolateral epiblastic explants engaged in concentric radial/centrifugal tissue expansion. In sharp contrast, Hensen's node explants displayed Cartesian-like, elongated, bipolar deformations—a pattern reminiscent of axis elongation. Time-lapse analysis of explant tissue motion patterns indicated that both cellular motility and extracellular matrix fiber (tissue) remodeling take place during the observed morphogenetic deformations. As expected, treatment of tissue explants with a selective Rho-Kinase (p160ROCK) signaling inhibitor, Y27632, completely arrested all morphogenetic movements. Microsurgical experiments revealed that lateral epiblastic tissue was dispensable for the generation of an elongated midline axis— provided that an intact organizer (node) is present. Our computational analyses suggest the possibility of delineating tissue-scale morphogenetic movements at anatomically discrete locations in the embryo. Further, tissue deformation patterns, as well as the mechanical state of the tissue, require normal actomyosin function. We conclude that amniote embryos contain tissue-scale, regionalized morphogenetic motion generators, which can be assessed using our novel computational time-lapse imaging approach. These data and future studies—using explants excised from overlapping

  7. Pollination and embryo development in Brassica rapa L. in microgravity.

    PubMed

    Kuang, A; Popova, A; Xiao, Y; Musgrave, M E

    2000-03-01

    Plant reproduction under spaceflight conditions has been problematic in the past. In order to determine what aspect of reproductive development is affected by microgravity, we studied pollination and embryo development in Brassica rapa L. during 16 d in microgravity on the space shuttle (STS-87). Brassica is self-incompatible and requires mechanical transfer of pollen. Short-duration access to microgravity during parabolic flights on the KC-135A aircraft was used initially to confirm that equal numbers of pollen grains could be collected and transferred in the absence of gravity. Brassica was grown in the Plant Growth Facility flight hardware as follows. Three chambers each contained six plants that were 13 d old at launch. As these plants flowered, thin colored tape was used to indicate the date of hand pollination, resulting in silique populations aged 8-15 d postpollination at the end of the 16-d mission. The remaining three chambers contained dry seeds that germinated on orbit to produce 14-d-old plants just beginning to flower at the time of landing. Pollen produced by these plants had comparable viability (93%) with that produced in the 2-d-delayed ground control. Matched-age siliques yielded embryos of equivalent developmental stage in the spaceflight and ground control treatments. Carbohydrate and protein storage reserves in the embryos, assessed by cytochemical localization, were also comparable. In the spaceflight material, growth and development by embryos rescued from siliques 15 d after pollination lagged behind the ground controls by 12 d; however, in the subsequent generation, no differences between the two treatments were found. The results demonstrate that while no stage of reproductive development in Brassica is absolutely dependent upon gravity, lower embryo quality may result following development in microgravity.

  8. A Review of The Society for Assisted Reproductive Technology Embryo Grading System and Proposed Modification

    PubMed Central

    Hossain, Amjad; Phelps, John; Agarwal, Ashok; Sanz, Eduardo; Mahadevan, Maha

    2016-01-01

    The Society for Assisted Reproductive Technology (SART) method of embryo grad- ing is unique, simple, and widely practiced, and its use has been mandatory for SART membership programs since 2010. Developed by SART in 2006, the current embryo grading system categories, “good, fair, and poor,” are limited because they do not describe the best 1-2 embryos in the interest of keeping pace with the shift in clinical practice to be more selective and to transfer fewer embryos. This inspired us to conduct a review on the SART embryo grading system. In this retrospective study, the literature on evaluation of human embryo quality in gen- eral, and the SART method of evaluation in particular, were reviewed for the period of 2000 to 2014. A multifaceted search pertaining to methods of embryo grading and trans- fer using a combination of relevant terms [embryo, mammalian, embryo transfer, grade, grading, morphology, biomarkers, SART, and in vitro fertilization (IVF)] was performed. The inclusion and exclusion in this review were dictated by the aim and scope of the study. Two investigators independently assessed the studies and extracted information. A total of 61 articles were reviewed. Very few studies have evaluated the efficacy of the SART embryo grading method. The present study suggests the necessity for revision of the current SART grading system. The system, as it is now, lacks criteria for describing the cohort specific best embryo and thus is of limited use in single embryo transfer. The study foresees heightened descriptive efficiency of the SART system by implementing the proposed changes. Strengths and weaknesses of the SART embryo grading were identified. Ideas for selecting the best cohort-specific embryo have been discussed, which may trigger methodological improvement in SART and other embryo grading systems. PMID:27441045

  9. Zygotic and somatic embryo morphogenesis in Pinus pinaster: comparative histological and histochemical study.

    PubMed

    Tereso, Susana; Zoglauer, Kurt; Milhinhos, Ana; Miguel, Célia; Oliveira, M Margarida

    2007-05-01

    We compared morphogenesis and accumulation of storage proteins and starch in Pinus pinaster Ait. zygotic embryos with those in somatic embryos grown with different carbohydrate sources. The maturation medium for somatic embryos included 80 microM abscisic acid (ABA), 9 g l(-1) gellam gum and either glucose, sucrose or maltose at 44, 88, 175 or 263 mM in the presence or absence of 6% (w/v) polyethylene glycol (PEG) 4000 MW. Maturation medium containing 44 or 88 mM of a carbohydrate source produced only one or no cotyledonary somatic embryos per 0.6 g fresh mass of culture. The addition of PEG to the basal maturation medium resulted in a low yield of cotyledonary somatic embryos that generally showed incomplete development and anatomical abnormalities such as large intercellular spaces and large vacuoles. High concentrations of maltose also induced large intercellular spaces in the somatic embryonic cells, and 263 mM sucrose produced fewer and less developed cotyledonary somatic embryos compared with 175 mM sucrose, indicating that the effect of carbohydrate source is partially osmotic. Zygotic embryos had a lower dry mass than somatic embryos at the same stage of development. Starch granules followed a similar accumulation pattern in zygotic and somatic embryos. A low starch content was found in cotyledonary zygotic embryos and in somatic embryos developed in the presence of 175 mM maltose or 263 mM glucose. In zygotic embryos and in PEG-treated somatic embryos, protein bodies appeared later and were smaller and fewer than in well-developed somatic embryos grown without PEG. We propose that storage protein concentration might be a marker of embryo quality.

  10. A Review of The Society for Assisted Reproductive Technology Embryo Grading System and Proposed Modification.

    PubMed

    Hossain, Amjad; Phelps, John; Agarwal, Ashok; Sanz, Eduardo; Mahadevan, Maha

    2016-01-01

    The Society for Assisted Reproductive Technology (SART) method of embryo grad- ing is unique, simple, and widely practiced, and its use has been mandatory for SART membership programs since 2010. Developed by SART in 2006, the current embryo grading system categories, "good, fair, and poor," are limited because they do not describe the best 1-2 embryos in the interest of keeping pace with the shift in clinical practice to be more selective and to transfer fewer embryos. This inspired us to conduct a review on the SART embryo grading system. In this retrospective study, the literature on evaluation of human embryo quality in gen- eral, and the SART method of evaluation in particular, were reviewed for the period of 2000 to 2014. A multifaceted search pertaining to methods of embryo grading and trans- fer using a combination of relevant terms [embryo, mammalian, embryo transfer, grade, grading, morphology, biomarkers, SART, and in vitro fertilization (IVF)] was performed. The inclusion and exclusion in this review were dictated by the aim and scope of the study. Two investigators independently assessed the studies and extracted information. A total of 61 articles were reviewed. Very few studies have evaluated the efficacy of the SART embryo grading method. The present study suggests the necessity for revision of the current SART grading system. The system, as it is now, lacks criteria for describing the cohort specific best embryo and thus is of limited use in single embryo transfer. The study foresees heightened descriptive efficiency of the SART system by implementing the proposed changes. Strengths and weaknesses of the SART embryo grading were identified. Ideas for selecting the best cohort-specific embryo have been discussed, which may trigger methodological improvement in SART and other embryo grading systems.

  11. Effect of the microenvironment and embryo density on developmental characteristics and gene expression profile of bovine preimplantative embryos cultured in vitro.

    PubMed

    Hoelker, Michael; Rings, Franka; Lund, Qamaruddin; Ghanem, Nasser; Phatsara, Chirawath; Griese, Josef; Schellander, Karl; Tesfaye, Dawit

    2009-03-01

    The Well of the Well (WOW) system has been developed to culture embryos in small groups or to track the development of single embryos. In the present study, we aimed to examine the effects of the microenvironment provided by the WOW system and embryo density on developmental rates, embryo quality and preimplantative gene expression profile of the resulting embryos. Embryos cultured in a group of 16 reached the blastocyst stage at a significantly lower level than zygotes cultured in a group of 50 (22.2 vs 30.3%), whereas zygotes cultured in WOW were able to compensate against low embryo densities, reaching a blastocyst rate as high as embryos cultured in a group of 50 (31.3 vs 30.3%). Moreover, embryos derived from WOW culture did not differ in terms of differential cell counts and apoptotic cell index compared with controls. The gene expression analysis revealed 62 transcripts to be upregulated and 33 transcripts to be downregulated by WOW culture. Comparing the in vivo derived blastocysts with the blastocysts derived from WOW culture, and group culture, expression of ATP5A1, PLAC8 and KRT8 was more similar to the embryos derived from WOW culture, whereas expression of S100A10 and ZP3 genes was more similar to blastocysts cultured in a group. In conclusion, microenvironment as well as embryo density significantly affected developmental rates. While subsequent blastocysts did not differ in terms of differential cell counts and apoptotic cell index, significant differences were observed in terms of the relative abundance of transcripts in the resulting embryos.

  12. Full term delivery following cryopreservation of human embryos for 7. 5 years.

    PubMed

    Ben-Ozer, S; Vermesh, M

    1999-06-01

    Successful pregnancy in a 44 year old woman is described following the transfer of embryos which were cryopreserved for 7.5 years. The embryos were obtained during a gamete intra-Fallopian transfer (GIFT) procedure in 1989. To our knowledge this is one of the longest published periods of cryopreservation of embryos which has resulted in a healthy baby. This report illustrates the previously presumed viability and normality of human embryos undergoing long-term cryopreservation. Additionally, it emphasizes the importance for advanced reproductive technique programmes and patients to review and update their embryo status.

  13. Supravital fluorometric apoptosis detection in a single mouse embryo using lab-on-a-chip.

    PubMed

    Walczak, Rafał; Śniadek, Patrycja; Dziuban, Jan A; Kluger, Joanna; Soyta, Anna Chełmońska

    2011-10-07

    Detection of apoptosis is one of the main criteria of preimplantation embryo growth potential assessment. Recent developments in lab-on-a-chip techniques has led to apoptosis detection and monitoring on a single cell or embryo level. However, single embryo apoptosis detection without a change in embryo developmental competence and post-examination "recovery" still remains a challenge. In this paper we present a lab-on-a-chip, co-working with miniaturized optical instrumentation, which allows supravital examination of single embryos for the presence of apoptotic blastomers with full after lab-on-a-chip study "recovery" and maintenance of their further developmental capacity.

  14. Analysis of factors influencing morphokinetic characteristics of embryos in ART cycles.

    PubMed

    Gryshchenko, Mykola Grygorievich; Pravdyuk, Alexey Igorovich; Parashchyuk, Valentin Yurievich

    2014-10-01

    In this article, some factors were evaluated for their impact on embryo morphokinetics during assisted reproductive technology (ART) cycles. We detected significant differences in the fourth cell division time (t5) of embryos obtained after controlled ovarian stimulation in long GnRH agonists and GnRH antagonist protocols. We also found that higher gonadotropin dose may slow down the development of embryos. However, both male and female age, the number of oocytes and number of normal forms of sperm in the ejaculate did not affect the kinetic parameters of embryo development. Further research is needed to identify all the spectrum of factors, which can affect the rate of embryo development.

  15. Phosphatized polar lobe-forming embryos from the Precambrian of southwest China.

    PubMed

    Chen, Jun-Yuan; Bottjer, David J; Davidson, Eric H; Dornbos, Stephen Q; Gao, Xiang; Yang, Yong-Hua; Li, Chia-Wei; Li, Gang; Wang, Xiu-Qiang; Xian, Ding-Chang; Wu, Hung-Jen; Hwu, Yeu-Kuang; Tafforeau, Paul

    2006-06-16

    In developing embryos of some extant spiralian animals, polar lobe formation is one of the symmetry-breaking mechanisms for segregation of maternal cytoplasmic substances to certain blastomeres and not others. Polar lobe formation leads to unique early cleavage morphologies that include trilobed, J-shaped, and five-lobed structures. Fossil embryos similar to modern lobeforming embryos are recognized from the Precambrian Doushantuo Formation phosphates, Weng'an, Guizhou Province, China. These embryos are abundant and form a developmental sequence comparable to different developing stages observed in lobe-forming embryos of extant spiralians. These data imply that lobe formation is an evolutionarily ancient process of embryonic specification.

  16. Embryo quality before and after slow freezing: Viability, implantation and pregnancy rates in 627 single frozen-thawed embryo replacement cycles following failure of fresh transfer.

    PubMed

    Capodanno, Francesco; De Feo, Gaetano; Gizzo, Salvatore; Nicoli, Alessia; Palomba, Stefano; La Sala, Giovanni Battista

    2016-06-01

    Frozen embryo transfer cycles are now common practice, however, various aspects regarding the potential of frozen embryos remain unclear. The main goal of the present study was to assess embryo quality before and after slow freezing procedure, and more specifically blastomere loss and embryo quality as indicator of viability. A single center retrospective analysis of single frozen-thawed embryo replacements (s-FER) was performed. The embryo quality before and after slow freezing and thawing, implantation, and pregnancy rates were recorded. One hundred and twenty seven s-FER were included in the final analysis. The probability of achieving an ongoing pregnancy was significantly associated with embryo quality and the percentage of blastomere loss after thawing. Considering thawed embryos, a non-significant difference in term of implantation rate was observed, regardless to their post-thawing quality and the percentage of blastomeres loss. In conclusion, current data suggest that thawed embryos are capable of implantation regardless of their morphological quality and the degree of cryoinjury sustained.

  17. Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing

    PubMed Central

    Liu, Wenqiang; Liu, Xiaoyu; Wang, Chenfei; Gao, Yawei; Gao, Rui; Kou, Xiaochen; Zhao, Yanhong; Li, Jingyi; Wu, You; Xiu, Wenchao; Wang, Su; Yin, Jiqing; Liu, Wei; Cai, Tao; Wang, Hong; Zhang, Yong; Gao, Shaorong

    2016-01-01

    Differentiated somatic cells can be reprogrammed into totipotent embryos through somatic cell nuclear transfer. However, most cloned embryos arrest at early stages and the underlying molecular mechanism remains largely unexplored. Here, we first developed a somatic cell nuclear transfer embryo biopsy system at two- or four-cell stage, which allows us to trace the developmental fate of the biopsied embryos precisely. Then, through single-cell transcriptome sequencing of somatic cell nuclear transfer embryos with different developmental fates, we identified that inactivation of Kdm4b, a histone H3 lysine 9 trimethylation demethylase, functions as a barrier for two-cell arrest of cloned embryos. Moreover, we discovered that inactivation of another histone demethylase Kdm5b accounts for the arrest of cloned embryos at the four-cell stage through single-cell analysis. Co-injection of Kdm4b and Kdm5b can restore transcriptional profiles of somatic cell nuclear transfer embryos and greatly improve the blastocyst development (over 95%) as well as the production of cloned mice. Our study therefore provides an effective approach to identify key factors responsible for the developmental arrest of somatic cell cloned embryos. PMID:27462457

  18. Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing.

    PubMed

    Liu, Wenqiang; Liu, Xiaoyu; Wang, Chenfei; Gao, Yawei; Gao, Rui; Kou, Xiaochen; Zhao, Yanhong; Li, Jingyi; Wu, You; Xiu, Wenchao; Wang, Su; Yin, Jiqing; Liu, Wei; Cai, Tao; Wang, Hong; Zhang, Yong; Gao, Shaorong

    2016-01-01

    Differentiated somatic cells can be reprogrammed into totipotent embryos through somatic cell nuclear transfer. However, most cloned embryos arrest at early stages and the underlying molecular mechanism remains largely unexplored. Here, we first developed a somatic cell nuclear transfer embryo biopsy system at two- or four-cell stage, which allows us to trace the developmental fate of the biopsied embryos precisely. Then, through single-cell transcriptome sequencing of somatic cell nuclear transfer embryos with different developmental fates, we identified that inactivation of Kdm4b, a histone H3 lysine 9 trimethylation demethylase, functions as a barrier for two-cell arrest of cloned embryos. Moreover, we discovered that inactivation of another histone demethylase Kdm5b accounts for the arrest of cloned embryos at the four-cell stage through single-cell analysis. Co-injection of Kdm4b and Kdm5b can restore transcriptional profiles of somatic cell nuclear transfer embryos and greatly improve the blastocyst development (over 95%) as well as the production of cloned mice. Our study therefore provides an effective approach to identify key factors responsible for the developmental arrest of somatic cell cloned embryos.

  19. Migratory ability of gonadal germ cells (GGCs) isolated from Ciconia boyciana and Geronticus eremita embryos into the gonad of developing chicken embryos

    PubMed Central

    NAKAJIMA, Yuki; FUKUDA, Haruka; ONUMA, Manabu; MURATA, Koichi; UEDA, Miya; SUNAGA, Emi; SHIRAISHI, Toshirou; TAJIMA, Atsushi

    2016-01-01

    We conducted experiments to evaluate the ability of gonadal germ cells (GGCs), isolated from the embryonic gonads of Ciconia boyciana or Geronticus eremita, to migrate into the gonads of developing chicken embryos. Fluorescently labeled GGCs, isolated by the PBS (−) method, were transferred into the dorsal aorta of 2-day-old chicken embryos. Five days after transfer, fluorescent GGCs were detected in the gonads of recipient embryos. Our results indicate that GGCs from Ciconia boyciana and Geronticus eremita are capable of migrating into the gonads of developing chicken embryos. PMID:26922915

  20. Digital Microfluidic Dynamic Culture of Mammalian Embryos on an Electrowetting on Dielectric (EWOD) Chip

    PubMed Central

    Huang, Hong-Yuan; Shen, Hsien-Hua; Tien, Chang-Hung; Li, Chin-Jung; Fan, Shih-Kang; Liu, Cheng-Hsien; Hsu, Wen-Syang; Yao, Da-Jeng

    2015-01-01

    Current human fertilization in vitro (IVF) bypasses the female oviduct and manually inseminates, fertilizes and cultivates embryos in a static microdrop containing appropriate chemical compounds. A microfluidic microchannel system for IVF is considered to provide an improved in-vivo-mimicking environment to enhance the development in a culture system for an embryo before implantation. We demonstrate a novel digitalized microfluidic device powered with electrowetting on a dielectric (EWOD) to culture an embryo in vitro in a single droplet in a microfluidic environment to mimic the environment in vivo for development of the embryo and to culture the embryos with good development and live births. Our results show that the dynamic culture powered with EWOD can manipulate a single droplet containing one mouse embryo and culture to the blastocyst stage. The rate of embryo cleavage to a hatching blastocyst with a dynamic culture is significantly greater than that with a traditional static culture (p<0.05). The EWOD chip enhances the culture of mouse embryos in a dynamic environment. To test the reproductive outcome of the embryos collected from an EWOD chip as a culture system, we transferred embryos to pseudo-pregnant female mice and produced live births. These results demonstrate that an EWOD-based microfluidic device is capable of culturing mammalian embryos in a microfluidic biological manner, presaging future clinical application. PMID:25933003

  1. Maternal restraint stress negatively influences growth capacity of preimplantation mouse embryos.

    PubMed

    Burkuš, Ján; Cikoš, Stefan; Fabian, Dušan; Kubandová, Janka; Czikková, Soňa; Koppel, Juraj

    2013-03-01

    In our study we investigated the effect of maternal restraint stress on preimplantation embryo development using a mouse model. We exposed hormonally stimulated (superovulated) and unstimulated (i.e. spontaneously ovulating) mouse females to restraint stress for 30 min three times a day during the preimplantation period. The stress exposure caused significant increase in blood plasma corticosterone concentration. Microscopical evaluation of embryos isolated from spontaneously ovulating females showed that maternal stress significantly increased the proportion of embryos with lower cell numbers (≤32 cells) and decreased the proportion of embryos with higher cell numbers (65-96 cells and 97-128 cells). Moreover maternal restraint stress decreased the cell counts per embryo and per blastocyst. After an additional 24 h in vitro culture we did not find any difference in the embryo distribution or in the cell counts per embryo/blastocyst between embryos isolated from stressed and control mothers. The exposure to restraint stress did not affect the incidence of apoptosis in blastocysts isolated from spontaneously ovulated dams. In gonadotropin stimulated dams, the hormonal treatment itself notably changed embryo distribution (increasing the proportion of degenerated embryos) and increased the occurrence of apoptotic cells. Our results indicate that psychical stress exposure in very early pregnancy can significantly influence the developmental capacity of preimplantation embryos.

  2. Utero-tubal embryo transfer and vasectomy in the mouse model.

    PubMed

    Bermejo-Alvarez, Pablo; Park, Ki-Eun; Telugu, Bhanu P

    2014-02-28

    The transfer of preimplantation embryos to a surrogate female is a required step for the production of genetically modified mice or to study the effects of epigenetic alterations originated during preimplantation development on subsequent fetal development and adult health. The use of an effective and consistent embryo transfer technique is crucial to enhance the generation of genetically modified animals and to determine the effect of different treatments on implantation rates and survival to term. Embryos at the blastocyst stage are usually transferred by uterine transfer, performing a puncture in the uterine wall to introduce the embryo manipulation pipette. The orifice performed in the uterus does not close after the pipette has been withdrawn, and the embryos can outflow to the abdominal cavity due to the positive pressure of the uterus. The puncture can also produce a hemorrhage that impairs implantation, blocks the transfer pipette and may affect embryo development, especially when embryos without zona are transferred. Consequently, this technique often results in very variable and overall low embryo survival rates. Avoiding these negative effects, utero-tubal embryo transfer take advantage of the utero-tubal junction as a natural barrier that impedes embryo outflow and avoid the puncture of the uterine wall. Vasectomized males are required for obtaining pseudopregnant recipients. A technique to perform vasectomy is described as a complement to the utero-tubal embryo transfer.

  3. Effective vitrification and warming of porcine embryos using a pH-stable, chemically defined medium

    PubMed Central

    Cuello, Cristina; Martinez, Cristina A.; Nohalez, Alicia; Parrilla, Inmaculada; Roca, Jordi; Gil, Maria A.; Martinez, Emilio A.

    2016-01-01

    The use of pH-stable media would simplify embryo vitrification and the warming of porcine embryos and might facilitate the application of embryo transfer in practice. In this work, we investigated whether a pH-stable basal medium constituted of Tyrode’s lactate medium, polyvinyl alcohol, and HEPES for buffering was suitable for porcine embryo vitrification warming in place of the conventional gas-equilibrated media. A high percentage (>90%) of embryos survived vitrification and warming in this medium, achieving in vitro survival rates similar to embryos vitrified-warmed using the conventional protocol and their fresh counterparts. The pH-stable medium did not affect the in vivo developmental competence of the vitrified-warmed embryos. A farrowing rate of 71.4% (5/7) with 10.4 ± 3.1 piglets born was obtained for the embryos vitrified and warmed in this medium and transferred to selected recipients. This medium will enable the use of simple, safe and standardized protocols for the vitrification and warming of porcine embryos for optimal embryo survival and quality when applied under field conditions. This study opens new possibilities for the widespread use of embryo transfer in pigs. PMID:27666294

  4. Cryopreservation of human embryos and its contribution to in vitro fertilization success rates.

    PubMed

    Wong, Kai Mee; Mastenbroek, Sebastiaan; Repping, Sjoerd

    2014-07-01

    Cryopreservation of human embryos is now a routine procedure in assisted reproductive technologies laboratories. There is no consensus on the superiority of any protocol, and substantial differences exist among centers in day of embryo cryopreservation, freezing method, selection criteria for which embryos to freeze, method of embryo thawing, and endometrial preparation for transfer of frozen-thawed embryos. In the past decade, the number of frozen-thawed embryo transfer cycles per started in vitro fertilization (IVF) cycle increased steadily, and at the same time the percentage of frozen-thawed embryo transfers that resulted in live births increased. Currently, cryopreservation of human embryos is more important than ever for the cumulative pregnancy rate after IVF. Interestingly, success rates after frozen-thawed embryo transfer are now nearing the success rates of fresh embryo transfer. This supports the hypothesis of so called freeze-all strategies in IVF, in which all embryos are frozen and no fresh transfer is conducted, to optimize success rates. High-quality randomized controlled trials should be pursued to find out which cryopreservation protocol is best and whether the time has come to completely abandon fresh transfers.

  5. A system to evaluate the quality of frozen embryos through short-term culture.

    PubMed

    Contreras, D A; Galina, C S; Avila, J G; Aspron, M P; Moreno-Mendoza, N

    2008-07-01

    The aim of the present study was to evaluate a culture system as a non-invasive approach intended for assessing the viability of recently thawed embryos prior to transfer. Embryos (n=51) were collected seven days after insemination out of 20 cows that had been treated to synchronize estrus and induce superovulation. Embryos were classified as good, fair, and poor and frozen. All embryos were cultured in McCoy medium. Morphology was monitored for a period of 24h to register the development stage every 30 min for the first 2h, and every hour thereafter. A sample of four embryos of each classification was separated at 4h, another four at 12h, and the remaining seven at 24h and the degree of apoptosis was determined for all the embryos using the TUNEL technique. Embryos of good and fair quality did not undergo major detrimental changes in development even after 7h of incubation, whereas poor quality embryos experienced changes as early as 2h after incubation. Good quality embryos invariably had fewer numbers of apoptotic cells than those of fair and poor quality suggesting that embryo culture can be a useful method to assess viability and to confirm the quality of thawed embryos previously stored in liquid nitrogen prior to transfer.

  6. Influence of embryo handling and transfer method on pig cloning efficiency.

    PubMed

    Shi, Junsong; Zhou, Rong; Luo, Lvhua; Mai, Ranbiao; Zeng, Haiyu; He, Xiaoyan; Liu, Dewu; Zeng, Fang; Cai, Gengyuan; Ji, Hongmei; Tang, Fei; Wang, Qinglai; Wu, Zhenfang; Li, Zicong

    2015-03-01

    The somatic cell nuclear transfer (SCNT) technique could be used to produce genetically superior or genetically engineered cloned pigs that have wide application in agriculture and bioscience research. However, the efficiency of porcine SCNT currently is very low. Embryo transfer (ET) is a key step for the success of SCNT. In this study, the effects of several ET-related factors, including cloned embryo culture time, recipient's ovulation status, co-transferred helper embryos and ET position, on the success rate of pig cloning were investigated. The results indicated that transfer of cloned embryos cultured for a longer time (22-24h vs. 4-6h) into pre-ovulatory sows decreased recipient's pregnancy rate and farrowing rate, and use of pre-ovulatory and post-ovulatory sows as recipients for SCNT embryos cultured for 22-24h resulted in a similar porcine SCNT efficiency. Use of insemination-produced in vivo fertilized, parthenogenetically activated and in vitro fertilized embryos as helper embryos to establish and/or maintain pregnancy of SCNT embryos recipients could not improve the success rate of porcine SCNT. Transfer of cloned embryos into double oviducts of surrogates significantly increased pregnancy rate as well as farrowing rate of recipients, and the developmental rate of transferred cloned embryos, as compared to unilateral oviduct transfer. This study provided useful information for optimization of the embryo handling and transfer protocol, which will help to improve the ability to generate cloned pigs.

  7. Studies on weak electromagnetic fields effects in chick embryos. Annual report, June 1985-June 1986

    SciTech Connect

    Not Available

    1986-05-31

    This research was directed to test some experimental conditions of the Henhouse project and to enforce a previous study on VLF electromagnetic fields effects on chick embryos. Henhouse Project: the response of White Leghorn Hisex embryos to field exposures effective on the Shaver breed, was studied. 1) A 48-hour exposure, in vivo, to a pulsed horizontal field of 100-Hz frequency, 1.0 micro T intensity, 500-microsecond pulse duration and 2-microsecond rise time induced a significant increase of developmental abnormalities in Hisex embryos. 2) A five-hour exposure of stage 7 Hisex embryos changed the Mitotic Index of their neural tissue. So, the early development of Hisex embryos, like Shaver embryos, can be modified by VLF pulsed electromagnetic fields. In the protocol of the Henhouse project, it was suggested a temperature of 38 C for eggs incubation. Studying the development of chick embryos in relation to the temperature, in the range of 37.4-40 C, it was confirmed that a 48-hour incubation at 38 C (with 55% humidity) does not induce abnormalities and allows a convenient developmental growth rate of the chick embryos. Electromagnetic fields effects in relation to the embryos orientation: preliminary results on the induction of abnormalities in field exposed embryos in relation to their orientation were confirmed. In a East-West oriented horizontal pulsed field, the organisms oriented to Southwest and Southeast showed a significant increase of developmental abnormalities. No effect was appreciable among the embryos Southward oriented.

  8. Growth and metabolism of murine and bovine embryos in bovine uterine flushing-supplemented culture media.

    PubMed Central

    Rondeau, M; Guay, P; Goff, A K; Cooke, G M

    1996-01-01

    The aim of this study was to compare the development and metabolic activity of cultured murine and bovine embryos in 2 standard media (HAM F-10 and RPMI) in the presence or absence of bovine uterine flushings. Murine morulae (n = 653) and day 7 bovine embryos (n = 273) were cultured for 18 h or 36 h in either HAM F-10 or RPMI in the presence or absence of bovine uterine flushings. After culture, the development, quality, and metabolic activity (glucose utilization or methionine uptake and incorporation) of embryos was assessed. It was found that HAM F-10 (without uterine flushings) was a more suitable medium than RPMI for optimal development and metabolism of murine and bovine embryos. Poor quality and development, as well as decreased metabolism, were evident after culture of murine embryos in RPMI; in contrast, this medium had no adverse effects on bovine embryos in culture. Supplementation of HAM F-10 with bovine uterine flushings improved the growth of murine embryos and the protein synthesis (as measured by an increased methionine incorporation) for both murine and bovine embryos. However, supplementation with bovine uterine flushings could not overcome deficiencies of an inappropriate medium (RPMI) for murine embryos. Supplementation of a well-defined culture medium with uterine flushings increased metabolism of embryos in culture, and thus might help to increase pregnancy rates after transfer of such embryos to recipient cows. PMID:8825988

  9. 104 FACTORS AFFECTING PREGNANCY RATES AND EMBRYO/FETAL LOSSES IN RECIPIENTS RECEIVING IN VITRO-PRODUCED EMBRYOS BY FIXED-TIME EMBRYO TRANSFER.

    PubMed

    Tribulo, A; Cedeño, A; Bernal, B; Andrada, S; Barajas, J L; Ortega, J; Oviedo, J M; Tribulo, H; Tribulo, R; Mapletoft, R J; Bó, G A

    2016-01-01

    A retrospective analysis evaluated pregnancy rates and embryo losses with in vitro-produced embryos in a commercial embryo transfer program on 15 different beef farms. Recipients were beef cows and heifers (n=1841) that were synchronized with 5 different protocols and transferred at a fixed-time (FTET). Recipients were examined by ultrasonography on Day 0, and those with a corpus luteum (CL) or a follicle ≥8mm in diameter and with body condition score 2 to 4 (1 to 5 scale) were synchronized. The synchronization treatments were as follows. (T1) Recipients received an intravaginal device with 0.5g of progesterone plus 2mg of oestradiol benzoate on Day 0; device removal, plus 500μg of cloprostenol (prostaglandin F2α), 400IU of eCG, and 0.5mg of oestradiol cypionate on Day 8; and FTET on Day 17. (T2) This treatment was similar to T1 but 1mg of oestradiol cypionate was injected at device removal instead of 0.5mg of oestradiol cypionate. (T3) This treatment was similar to T1 except that animals were tail-painted on Day 8 and observed on Day 10. Those with the tail-paint intact on Day 10 received 100μg of gonadorelin (gonadotropin-releasing hormone) and all recipients were FTET on Day 17. (T4) Recipients received a progesterone device on Day 0; device removal, prostaglandin F2α, and eCG on Day 5; gonadotropin-releasing hormone on Day 8; and FTET on Day 15. (T5) Recipients received a progesterone device and 2mg of oestradiol benzoate on Day 0; device removal, prostaglandin F2α, and eCG on Day 6; gonadotropin-releasing hormone on Day 9; and FTET on Day 16. On the day of FTET all recipients with CL ≥18mm in diameter (G1), ≥16 and <18mm in diameter (G2), and ≥14mm and <16mm in diameter (G3) received in vitro-produced fresh embryos. Pregnancy was diagnosed by ultrasonography at 30 and 60 days of gestation, and data were analysed by logistic regression. The overall proportion of recipients synchronized that were FTET was 80.8% (1487/1841), with a 30-day pregnancy

  10. In vitro culture of mouse embryos reduces differential gene expression between inner cell mass and trophectoderm.

    PubMed

    Giritharan, G; Delle Piane, L; Donjacour, A; Esteban, F J; Horcajadas, J A; Maltepe, E; Rinaudo, P

    2012-03-01

    Differences in gene expression and imprinting have been reported, comparing in vivo versus in vitro generated preimplantation embryos. Furthermore, mouse studies have shown that placenta development is altered following in vitro culture. However, the molecular mechanisms underlying these findings are unknown. We therefore isolated trophectoderm (TE) and inner cell mass (ICM) cells from in vivo and in vitro fertilization (IVF) embryos and evaluated their transcriptome using microarrays. We found that the transcriptomes of in vitro produced ICM and TE cells showed remarkably few differences compared to ICM and TE cells of in vivo generated embryos. In vitro fertilization embryos showed a reduced number of TE cells compared to in vivo embryos. In addition, TE of IVF embryos showed significant downregulation of solute transporter genes and of genes involved in placenta formation (Eomesodermin, Socs3) or implantation (Hbegf). In summary, IVF and embryo culture significantly affects the transcriptome of ICM and TE cells.

  11. In Vitro Culture of Mouse Embryos Reduces Differential Gene Expression Between Inner Cell Mass and Trophectoderm

    PubMed Central

    Giritharan, G.; Piane, L. Delle; Donjacour, A.; Esteban, F. J.; Horcajadas, J. A.; Maltepe, E.; Rinaudo, P.

    2012-01-01

    Differences in gene expression and imprinting have been reported, comparing in vivo versus in vitro generated preimplantation embryos. Furthermore, mouse studies have shown that placenta development is altered following in vitro culture. However, the molecular mechanisms underlying these findings are unknown. We therefore isolated trophectoderm (TE) and inner cell mass (ICM) cells from in vivo and in vitro fertilization (IVF) embryos and evaluated their transcriptome using microarrays. We found that the transcriptomes of in vitro produced ICM and TE cells showed remarkably few differences compared to ICM and TE cells of in vivo generated embryos. In vitro fertilization embryos showed a reduced number of TE cells compared to in vivo embryos. In addition, TE of IVF embryos showed significant downregulation of solute transporter genes and of genes involved in placenta formation (Eomesodermin, Socs3) or implantation (Hbegf). In summary, IVF and embryo culture significantly affects the transcriptome of ICM and TE cells. PMID:22383776

  12. Rescue of keratin 18/19 doubly deficient mice using aggregation with tetraploid embryos.

    PubMed

    Hesse, Michael; Watson, Erica D; Schwaluk, Tanja; Magin, Thomas M

    2005-03-01

    We have previously shown that the targeted deletions of both type I keratins (K) 18 and 19 cause lethality by embryonic day (e) 9.5 due to fragility and cytolysis of trophoblast giant cells. The development of the embryo proper appeared to be unaffected and its death was caused by nutrient deficiency. In order to address the function of keratins within the embryo proper, lethality due to extraembryonic tissue failure must be overcome. One approach to rescue doubly deficient embryos is by aggregating knockout embryos with tetraploid wild-type embryos. As a general tool, tetraploid aggregation can be used to rescue embryonic lethality caused by defects in extraembryonic tissues like the placenta, trophoblast or yolk sac. We rescued K18-/- K19-/- embryos until e11.5, using this approach, proving that the loss of the keratin cytoskeleton causes defects in the trophoblast giant cell layer, but has no effect on early development of the embryo proper.

  13. BioMEMS for high-throughput handling and microinjection of embryos

    NASA Astrophysics Data System (ADS)

    Bernstein, Ralph W.; Scott, Matthew; Solgaard, Olav

    2004-12-01

    Technologies for handling, sorting, and positioning of embryos are increasingly important in biomedicine. In this paper the status for ongoing projects aimed at developing instrumentation for high-throughput treatment of embryos is reviewed. Techniques for positioning of Drosophila (fruit-fly) embryos in 2-D arrays for use in microinjection experiments are especially focused. A method based on fluidic micro assembly is discussed, and important parameters such as immobilization yield, the number of misplaced embryos, and adhesion force of the embryos are reported. A model for the assembly process is described, and simulation results are in good agreement with adhesion force measurements. A fully automated MEMS based system for fruit-fly embryo injection has recently been demonstrated at Stanford University. The first experiments with double-stranded RNA injection proved successful, and the expected genetic modification of the embryos was observed.

  14. 9 CFR 85.10 - Interstate movement of swine semen and swine embryos for insemination of or implantation into swine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... swine embryos for insemination of or implantation into swine. 85.10 Section 85.10 Animals and Animal... and swine embryos for insemination of or implantation into swine. Swine semen and swine embryos moved... collection of the semen or embryos or were members of a qualified pseudorabies negative herd, and had...

  15. 9 CFR 85.10 - Interstate movement of swine semen and swine embryos for insemination of or implantation into swine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... swine embryos for insemination of or implantation into swine. 85.10 Section 85.10 Animals and Animal... and swine embryos for insemination of or implantation into swine. Swine semen and swine embryos moved... collection of the semen or embryos or were members of a qualified pseudorabies negative herd, and had...

  16. 9 CFR 85.10 - Interstate movement of swine semen and swine embryos for insemination of or implantation into swine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... swine embryos for insemination of or implantation into swine. 85.10 Section 85.10 Animals and Animal... and swine embryos for insemination of or implantation into swine. Swine semen and swine embryos moved... collection of the semen or embryos or were members of a qualified pseudorabies negative herd, and had...

  17. 9 CFR 98.21 - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.21 Embryos from...

  18. 9 CFR 98.10a - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses...

  19. 9 CFR 98.21 - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.21 Embryos from...

  20. 9 CFR 98.10a - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses...

  1. 9 CFR 98.10a - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses...

  2. 9 CFR 98.21 - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.21 Embryos from...

  3. 9 CFR 98.10a - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses...

  4. 9 CFR 85.10 - Interstate movement of swine semen and swine embryos for insemination of or implantation into swine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... swine embryos for insemination of or implantation into swine. 85.10 Section 85.10 Animals and Animal... and swine embryos for insemination of or implantation into swine. Swine semen and swine embryos moved... collection of the semen or embryos or were members of a qualified pseudorabies negative herd, and had...

  5. 9 CFR 98.21 - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.21 Embryos from...

  6. 9 CFR 85.10 - Interstate movement of swine semen and swine embryos for insemination of or implantation into swine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... swine embryos for insemination of or implantation into swine. 85.10 Section 85.10 Animals and Animal... and swine embryos for insemination of or implantation into swine. Swine semen and swine embryos moved... collection of the semen or embryos or were members of a qualified pseudorabies negative herd, and had...

  7. 9 CFR 98.10a - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos from Regions Free of Rinderpest and Foot-and-Mouth Disease; and Embryos of Horses...

  8. 9 CFR 98.21 - Embryos from sheep in regions other than Australia, Canada, and New Zealand.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Embryos from sheep in regions other... (INCLUDING POULTRY) AND ANIMAL PRODUCTS IMPORTATION OF CERTAIN ANIMAL EMBRYOS AND ANIMAL SEMEN Ruminant and Swine Embryos From Regions Where Rinderpest or Foot-and-Mouth Disease Exists § 98.21 Embryos from...

  9. Evaluation of quail and chicken embryos for the detection of botulism toxin serotypes A, B E and F activity.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Comparison of quail (Coturnix japonica) and chicken (Gallus domesticus) embryos for the detection of BoNT/A activity was conducted using equal dosages of toxin/g of embryo (quail at 7 g and chickens at 48 g). Quail embryos were injected at 0, 0.5 to 50 ng and chicken embryos at 0, 3.4 to 342 ng and...

  10. Evaluation of quail and chicken embryos for the detection of botulinum toxin serotypes A, B, E and F activity.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Comparison of quail (Coturnix japonica) and chicken (Gallus domesticus) embryos for the detection of BoNT/A activity was conducted using equal dosages of toxin/g of embryo (quail at 7 g and chickens at 48 g). Quail embryos were injected at 0, 0.5 to 50 ng adn chicken embryos at 0, 3.4 to 342 ng and...

  11. HSPC117 deficiency in cloned embryos causes placental abnormality and fetal death

    SciTech Connect

    Wang, Yingying; Hai, Tang; Liu, Zichuan; Zhou, Shuya; Lv, Zhuo; Ding, Chenhui; Liu, Lei; Niu, Yuyu; Zhao, Xiaoyang; Tong, Man; Wang, Liu; Jouneau, Alice; Zhang, Xun; Ji, Weizhi; Zhou, Qi

    2010-07-02

    Somatic cell nuclear transfer (SCNT) has been successfully used in many species to produce live cloned offspring, albeit with low efficiency. The low frequency of successful development has usually been ascribed to incomplete or inappropriate reprogramming of the transferred nuclear genome. Elucidating the genetic differences between normal fertilized and cloned embryos is key to understand the low efficiency of SCNT. Here, we show that expression of HSPC117, which encodes a hypothetical protein of unknown function, was absent or very low in cloned mouse blastocysts. To investigate the role of HSPC117 in embryo development, we knocked-down this gene in normal fertilized embryos using RNA interference. We assessed the post-implantation survival of HSPC117 knock-down embryos at 3 stages: E9 (prior to placenta formation); E12 (after the placenta was fully functional) and E19 (post-natal). Our results show that, although siRNA-treated in vivo fertilized/produced (IVP) embryos could develop to the blastocyst stage and implanted without any difference from control embryos, the knock-down embryos showed substantial fetal death, accompanied by placental blood clotting, at E12. Furthermore, comparison of HSPC117 expression in placentas of nuclear transfer (NT), intracytoplasmic sperm injection (ICSI) and IVP embryos confirmed that HSPC117 deficiency correlates well with failures in embryo development: all NT embryos with a fetus, as well as IVP and ICSI embryos, had normal placental HSPC117 expression while those NT embryos showing reduced or no expression of HSPC117 failed to form a fetus. In conclusion, we show that HSPC117 is an important gene for post-implantation development of embryos, and that HSPC117 deficiency leads to fetal abnormalities after implantation, especially following placental formation. We suggest that defects in HSPC117 expression may be an important contributing factor to loss of cloned NT embryos in vivo.

  12. Embryos produced from fertilization with bovine viral diarrhea virus (BVDV)-infected semen and the risk of disease transmission to embryo transfer (ET) recipients and offspring.

    PubMed

    Bielanski, A; Algire, J; Lalonde, A; Garceac, A

    2013-09-15

    Bovine diarrhea virus (BVDV) causes a variety of economically important enteric and infertility problems in cattle. For that reason, several countries have eradicated the disease, and some others have schemes in progress to achieve freedom. Although there is a considerable amount of information about the risk of BVDV transmission through contaminated semen used for artificial insemination (AI), there is no evidence to indicate whether the resulting embryos, when used for embryo transfer, can lead to the transmission of BVDV to recipients or offspring. For this experiment, semen from a bull persistently infected with BVDV (10(5) 50% tissue culture infective doses/mL NY strain) was used for insemination (two times at estrus) of BVDV-seronegative, superovulated cows (N = 35). Embryos were collected 7 days after insemination and subsequently were washed according to the International Embryo Transfer Society recommendations or left unwashed. Out of 302 collected oocytes and embryos, 173 (57%) were fertilized and the remaining 129 (43%) had degenerated. Infectious BVDV was detected in 24% (17/71) of unwashed and 10% (8/77) of washed embryos, and in all (N = 11) follicular fluid samples, oviductal epithelial cells, endometrium, and corpora lutea tissues as determined by the virus isolation test. After transfer of 39 washed embryos to 27 BVDV-seronegative recipients, 12 (44%) cows became pregnant and 17 calves free of BVDV and BVDV antibodies, including five sets of twins, were born. After embryo transfer, all pregnant and nonpregnant recipients remained free of BVDV and antibodies. In conclusion, results herein suggest that BVDV can be transmitted by AI resulting in the production of some proportion of contaminated embryos. However, it appears that such embryos, when washed according to International Embryo Transfer Society and the World Organization for Animal Health guidelines do not cause BVDV transmission to recipients or their offspring.

  13. ETHICS, EMBRYOS, AND EVIDENCE: A LOOK BACK AT WARNOCK.

    PubMed

    Hammond-Browning, Natasha

    2015-01-01

    The Report of the Committee of Inquiry into Human Fertilisation and Embryology, the Warnock Report, forms the basis of the UK legislation on embryo research, and its influence continues to be felt, even though over 30 years have passed since its publication. The Warnock Committee was the first of its kind to consider how advancements in human fertilisation and embryology should be regulated. This article examines the evidence submitted to the Warnock Committee, upon which its members ultimately reached their conclusions. With ongoing debate as to the status of the human embryo, it is important to recognise that the legislative position is one that was reached after extensive consultation and consideration of submitted evidence by the Warnock Committee. This article considers the differing ethical viewpoints that were expressed by organisations both prior and post-publication of the Warnock Report, and how the Committee used that evidence to reach their conclusions, and ultimately calls for a new Warnock-style committee.

  14. Proteomic analysis of indium embryotoxicity in cultured postimplantation rat embryos.

    PubMed

    Usami, Makoto; Nakajima, Mikio; Mitsunaga, Katsuyoshi; Miyajima, Atsuko; Sunouchi, Momoko; Doi, Osamu

    2009-12-01

    Indium embryotoxicity was investigated by proteomic analysis with two-dimensional electrophoresis of rat embryos cultured from day 10.5 of gestation for 24h in the presence of 50 microM indium trichloride. In the embryo proper, indium increased quantity of several protein spots including those identified as serum albumin, phosphorylated cofilin 1, phosphorylated destrin and tyrosyl-tRNA synthetase. The increased serum albumin, derived from the culture medium composed of rat serum, may decrease the toxicity of indium. The increase of phosphorylated cofilin 1 might be involved in dysmorphogenicity of indium through perturbation of actin functions. In the yolk sac membrane, indium induced quantitative and qualitative changes in the protein spots. Proteins from appeared spots included stress proteins, and those from decreased or disappeared spots included serum proteins, glycolytic pathway enzymes and cytoskeletal proteins, indicating yolk sac dysfunction. Thus, several candidate proteins that might be involved in indium embryotoxicity were identified.

  15. Heterologous embryo transfer: Magisterial answers and metaphysical questions.

    PubMed

    Accad, Michel

    2014-02-01

    The debate regarding the morality of heterologous embryo transfer (HET) as a solution for the fate of cryopreserved embryos remains active. This paper endeavors to show that the magisterial instructions on bioethical issues can only lead to the conclusion that HET is always morally illicit. I begin by showing that the text of Dignitas personae recognizes HET as a procedure accomplishing a procreative function, and I indicate that it is through gestation that this procreative function occurs. I further show that the previous Instruction, Donum vitae, implicitly points to an ontological or spiritual consideration at play during gestation. This consideration is likely related to the procreative function identified in Dignitas personae. Finally, I place these two textual arguments in the context of the debate concerning HET and conclude that metaphysical questions must be clarified in order for the immorality of HET to be understood from a suitable anthropological perspective and gain more widespread acceptance.

  16. Microfluidics for gametes, embryos, and embryonic stem cells.

    PubMed

    Smith, G D; Swain, J E; Bormann, C L

    2011-01-01

    Microfluidics is a young but established field that holds significant potential for scientific discovery. The utility of microfluidics can improve our knowledge of basic biology as well as expand our understanding in specialized areas such as assisted reproduction and stem cell developmental biology. This review describes the technology of microfluidics and discusses applications within assisted reproduction technology and embryonic stem cell growth and directed differentiation. Development of an integrated microfluidic platform for assisted reproduction, which can manipulate gametes, embryos, embryonic stem cells, their culture environment, and incorporate biomarker analysis, could have a dramatic impact on the basic understanding of embryo/embryonic stem cell development, as well as provide significant improvements in current technologies used to treat infertility, preserve fertility, and derive therapeutic cells from stem cells.

  17. Stage dependent susceptibility to lead in Bufo arenarum embryos.

    PubMed

    Pérez-Coll, C S; Herkovits, J

    1990-01-01

    The stage dependent susceptibility to lead in amphibian development was studied by exposing Bufo arenarum embryos during neurulae, neuromuscular activity and gill circulation stages for twenty hours to 1 ppm Pb(2+). Survival, malformations and behavioral disorders were evaluated. The embryonic susceptibility to lead was markedly stage dependent. The survival at the neuromuscular activity stage was approximately half that of the other two periods; concerning malformations, the gill circulation stage was the least sensitive. The observed malformations consisted of failed closure of neural tube, hydropsy, small and cylindrical tails, reduced body size and incurvations in the body axis. Some alterations occurred in all experimental groups and therefore were considered non-dependent on the period of treatment. In all experimental embryos, neurological disorders such as trembles and loss of equilibrium were observed.

  18. Myomaker mediates fusion of fast myocytes in zebrafish embryos.

    PubMed

    Landemaine, Aurélie; Rescan, Pierre-Yves; Gabillard, Jean-Charles

    2014-09-05

    Myomaker (also called Tmem8c), a new membrane activator of myocyte fusion was recently discovered in mice. Using whole mount in situ hybridization on zebrafish embryos at different stages of embryonic development, we show that myomaker is transiently expressed in fast myocytes forming the bulk of zebrafish myotome. Zebrafish embryos injected with morpholino targeted against myomaker were alive after yolk resorption and appeared morphologically normal, but they were unable to swim, even under effect of a tactile stimulation. Confocal observations showed a marked phenotype characterized by the persistence of mononucleated muscle cells in the fast myotome at developmental stages where these cells normally fuse to form multinucleated myotubes. This indicates that myomaker is essential for myocyte fusion in zebrafish. Thus, there is an evolutionary conservation of myomaker expression and function among Teleostomi.

  19. Heterologous embryo transfer: Magisterial answers and metaphysical questions

    PubMed Central

    Accad, Michel

    2014-01-01

    The debate regarding the morality of heterologous embryo transfer (HET) as a solution for the fate of cryopreserved embryos remains active. This paper endeavors to show that the magisterial instructions on bioethical issues can only lead to the conclusion that HET is always morally illicit. I begin by showing that the text of Dignitas personae recognizes HET as a procedure accomplishing a procreative function, and I indicate that it is through gestation that this procreative function occurs. I further show that the previous Instruction, Donum vitae, implicitly points to an ontological or spiritual consideration at play during gestation. This consideration is likely related to the procreative function identified in Dignitas personae. Finally, I place these two textual arguments in the context of the debate concerning HET and conclude that metaphysical questions must be clarified in order for the immorality of HET to be understood from a suitable anthropological perspective and gain more widespread acceptance. PMID:24899737

  20. Ambient UV-B radiation causes deformities in amphibian embryos

    USGS Publications Warehouse

    Blaustein, A.R.; Kiesecker, J.M.; Chivers, D.P.; Anthony, R.G.

    1997-01-01

    There has been a great deal of recent attention on the suspected increase in amphibian deformities. However, most reports of amphibian deformities have been anecdotal, and no experiments in the field under natural conditions have been performed to investigate this phenomenon. Under laboratory conditions, a variety of agents can induce deformities in amphibians. We investigated one of these agents, UV-B radiation, in field experiments, as a cause for amphibian deformities. We monitored hatching success and development in long-toed salamanders under UV-B shields and in regimes that allowed UV-B radiation. Embryos under UV-B shields had a significantly higher hatching rate and fewer deformities, and developed more quickly than those exposed to UV-B. Deformities may contribute directly to embryo mortality, and they may affect an individual's subsequent survival after hatching.