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Sample records for human embryo development

  1. [Association of human chorionic gonadotropin level in embryo culture media with early embryo development].

    Science.gov (United States)

    Wang, Haiying; Zhang, Renli; Han, Dong; Liu, Caixia; Cai, Jiajie; Bi, Yanling; Wen, Anmin; Quan, Song

    2014-06-01

    To investigate the association of human chorionic gonadotropin (HCG) level on day 3 of embryo culture with embryo development. Spent culture media were collected from individually cultured embryos on day 3 of in vitro fertilization and embryo transfer (IVF-ET) cycles. HCG concentration in the culture media was measured using an ELISA kit and its association with embryo development was assessed. In the 163 samples of embryo culture media from 60 patients, HCG was positive in 153 sample (93.8%) with a mean level of 0.85 ± 0.43 mIU/ml. The concentration of hCG in the culture media increased gradually as the number of blastomeres increased (F=2.273, P=0.03), and decreased as the morphological grade of the embryo was lowered (F=3.900, P=0.02). ELISA is capable of detecting HCG levels in spent culture media of embryos on day 3 of in vitro culture. The concentration of HCG in spent culture media is positively correlated with the status of early embryo development and implantation rate and thus serves as a useful marker for embryo selection in IVF-ET procedure.

  2. Effects of fluoxetine on human embryo development

    NARCIS (Netherlands)

    Kaihola, Helena; Yaldir, Fatma G.; Hreinsson, Julius; Hornaeus, Katarina; Bergquist, Jonas; Olivier, Jocelien D. A.; Akerud, Helena; Sundstrom-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,

  3. Development of the ventral body wall in the human embryo

    NARCIS (Netherlands)

    Mekonen, Hayelom K.; Hikspoors, Jill P. J. M.; Mommen, Greet; Köhler, S. Eleonore; Lamers, Wouter H.

    2015-01-01

    Migratory failure of somitic cells is the commonest explanation for ventral body wall defects. However, the embryo increases ~ 25-fold in volume in the period that the ventral body wall forms, so that differential growth may, instead, account for the observed changes in topography. Human embryos

  4. Expression of Aquaporins in Human Embryos and Potential Role of AQP3 and AQP7 in Preimplantation Mouse Embryo Development

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    Yun Xiong

    2013-05-01

    Full Text Available Background/Aims: Water channels, also named aquaporins (AQPs, play crucial roles in cellular water homeostasis. Methods: RT-PCR indicated the mRNA expression of AQPs 1-5, 7, 9, and 11-12, but not AQPs 0, 6, 8, and 10 in the 2∼8-cell stage human embryos. AQP3 and AQP7 were further analyzed for their mRNA expression and protein expression in the oocyte, zygote, 2-cell embryo, 4-cell embryo, 8-cell embryo, morula, and blastocyst from both human and mouse using RT-PCR and immunofluorescence, respectively. Results: AQP3 and AQP7 were detected in all these stages. Knockdown of either AQP3 or AQP7 by targeted siRNA injection into 2-cell mouse embryos significantly inhibited preimplantation embryo development. However, knockdown of AQP3 in JAr spheroid did not affect its attachment to Ishikawa cells. Conclusion: These data demonstrate that multiple aquaporins are expressed in the early stage human embryos and that AQP3 and AQP7 may play a role in preimplantation mouse embryo development.

  5. Development of the ventral body wall in the human embryo.

    Science.gov (United States)

    Mekonen, Hayelom K; Hikspoors, Jill P J M; Mommen, Greet; Köhler, S Eleonore; Lamers, Wouter H

    2015-11-01

    Migratory failure of somitic cells is the commonest explanation for ventral body wall defects. However, the embryo increases ~ 25-fold in volume in the period that the ventral body wall forms, so that differential growth may, instead, account for the observed changes in topography. Human embryos between 4 and 10 weeks of development were studied, using amira reconstruction and cinema 4D remodeling software for visualization. Initially, vertebrae and ribs had formed medially, and primordia of sternum and hypaxial flank muscle primordium laterally in the body wall at Carnegie Stage (CS)15 (5.5 weeks). The next week, ribs and muscle primordium expanded in ventrolateral direction only. At CS18 (6.5 weeks), separate intercostal and abdominal wall muscles differentiated, and ribs, sterna, and muscles began to expand ventromedially and caudally, with the bilateral sternal bars fusing in the midline after CS20 (7 weeks) and the rectus muscles reaching the umbilicus at CS23 (8 weeks). The near-constant absolute distance between both rectus muscles and approximately fivefold decline of this distance relative to body circumference between 6 and 10 weeks identified dorsoventral growth in the dorsal body wall as determinant of the 'closure' of the ventral body wall. Concomitant with the straightening of the embryonic body axis after the 6th week, the abdominal muscles expanded ventrally and caudally to form the infraumbilical body wall. Our data, therefore, show that the ventral body wall is formed by differential dorsoventral growth in the dorsal part of the body. © 2015 Anatomical Society.

  6. Development of the ventral body wall in the human embryo

    Science.gov (United States)

    Mekonen, Hayelom K; Hikspoors, Jill P J M; Mommen, Greet; Köhler, S Eleonore; Lamers, Wouter H

    2015-01-01

    Migratory failure of somitic cells is the commonest explanation for ventral body wall defects. However, the embryo increases ∼ 25-fold in volume in the period that the ventral body wall forms, so that differential growth may, instead, account for the observed changes in topography. Human embryos between 4 and 10 weeks of development were studied, using amira® reconstruction and cinema 4D® remodeling software for visualization. Initially, vertebrae and ribs had formed medially, and primordia of sternum and hypaxial flank muscle primordium laterally in the body wall at Carnegie Stage (CS)15 (5.5 weeks). The next week, ribs and muscle primordium expanded in ventrolateral direction only. At CS18 (6.5 weeks), separate intercostal and abdominal wall muscles differentiated, and ribs, sterna, and muscles began to expand ventromedially and caudally, with the bilateral sternal bars fusing in the midline after CS20 (7 weeks) and the rectus muscles reaching the umbilicus at CS23 (8 weeks). The near-constant absolute distance between both rectus muscles and approximately fivefold decline of this distance relative to body circumference between 6 and 10 weeks identified dorsoventral growth in the dorsal body wall as determinant of the ‘closure’ of the ventral body wall. Concomitant with the straightening of the embryonic body axis after the 6th week, the abdominal muscles expanded ventrally and caudally to form the infraumbilical body wall. Our data, therefore, show that the ventral body wall is formed by differential dorsoventral growth in the dorsal part of the body. PMID:26467243

  7. The fate of the mosaic embryo: Chromosomal constitution and development of Day 4, 5 and 8 human embryos

    NARCIS (Netherlands)

    M.A. Santos; G. Teklenburg (Gijs); N.S. Macklon (Nick); D. van Opstal (Diane); G.H. Schuring-Blom (Heleen); P-J. Krijtenburg (Pieter-Jaap); J. de Vreeden-Elbertse (Johanna); B.C.J.M. Fauser (Bart); E.B. Baart (Esther)

    2010-01-01

    textabstractBackground: Post-zygotic chromosome segregation errors are very common in human embryos after in vitro fertilization, resulting in mosaic embryos. However, the significance of mosaicism for the developmental potential of early embryos is unknown. We assessed chromosomal constitution and

  8. Human endometrial cell coculture reduces the endocrine disruptor toxicity on mouse embryo development

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    Lee Myeong-Seop

    2012-04-01

    Full Text Available Abstract Backgrounds Previous studies suggested that endocrine disruptors (ED are toxic on preimplantation embryos and inhibit development of embryos in vitro culture. However, information about the toxicity of endocrine disruptors on preimplantation development of embryo in human reproductive environment is lacking. Methods Bisphenol A (BPA and Aroclor 1254 (polychlorinated biphenyls were used as endocrine disruptors in this study. Mouse 2-cell embryos were cultured in medium alone or vehicle or co-cultured with human endometrial epithelial layers in increasing ED concentrations. Results At 72 hours the percentage of normal blastocyst were decreased by ED in a dose-dependent manner while the co-culture system significantly enhanced the rate and reduced the toxicity of endocrine disruptors on the embryonic development in vitro. Conclusions In conclusion, although EDs have the toxic effect on embryo development, the co-culture with human endometrial cell reduced the preimplantation embryo from it thereby making human reproductive environment protective to preimplantation embryo from the toxicity of endocrine disruptors.

  9. Global gene expression profiling of individual human oocytes and embryos demonstrates heterogeneity in early development.

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    Lisa Shaw

    Full Text Available Early development in humans is characterised by low and variable embryonic viability, reflected in low fecundity and high rates of miscarriage, relative to other mammals. Data from assisted reproduction programmes provides additional evidence that this is largely mediated at the level of embryonic competence and is highly heterogeneous among embryos. Understanding the basis of this heterogeneity has important implications in a number of areas including: the regulation of early human development, disorders of pregnancy, assisted reproduction programmes, the long term health of children which may be programmed in early development, and the molecular basis of pluripotency in human stem cell populations. We have therefore investigated global gene expression profiles using polyAPCR amplification and microarray technology applied to individual human oocytes and 4-cell and blastocyst stage embryos. In order to explore the basis of any variability in detail, each developmental stage is replicated in triplicate. Our data show that although transcript profiles are highly stage-specific, within each stage they are relatively variable. We describe expression of a number of gene families and pathways including apoptosis, cell cycle and amino acid metabolism, which are variably expressed and may be reflective of embryonic developmental competence. Overall, our data suggest that heterogeneity in human embryo developmental competence is reflected in global transcript profiles, and that the vast majority of existing human embryo gene expression data based on pooled oocytes and embryos need to be reinterpreted.

  10. Trichostatin A (TSA) improves the development of rabbit-rabbit intraspecies cloned embryos, but not rabbit-human interspecies cloned embryos.

    Science.gov (United States)

    Shi, Li-Hong; Miao, Yi-Liang; Ouyang, Ying-Chun; Huang, Jun-Cheng; Lei, Zi-Li; Yang, Ji-Wen; Han, Zhi-Ming; Song, Xiang-Fen; Sun, Qing-Yuan; Chen, Da-Yuan

    2008-03-01

    The interspecies somatic cell nuclear transfer (iSCNT) technique for therapeutic cloning gives great promise for treatment of many human diseases. However, the incomplete nuclear reprogramming and the low blastocyst rate of iSCNT are still big problems. Herein, we observed the effect of TSA on the development of rabbit-rabbit intraspecies and rabbit-human interspecies cloned embryos. After treatment with TSA for 6 hr during activation, we found that the blastocyst rate of rabbit-rabbit cloned embryos was more than two times higher than that of untreated embryos; however, the blastocyst rate of TSA-treated rabbit-human interspecies cloned embryos decreased. We also found evident time-dependent histone deacetylation-reacetylation changes in rabbit-rabbit cloned embryos, but not in rabbit-human cloned embryos from fusion to 6 hr after activation. Our results suggest that TSA-treatment does not improve blastocyst development of rabbit-human iSCNT embryos and that abnormal histone deacetylation-reacetylation changes in iSCNT embryos may account for their poor blastocyst development. (c) 2008 Wiley-Liss, Inc.

  11. Effect of oxygen concentration on human embryo development evaluated by time-lapse monitoring

    DEFF Research Database (Denmark)

    Ingerslev, Hans Jakob; Hindkjær, Johnny Juhl; Kirkegaard, Kirstine

    2012-01-01

    was to evaluate the influence of oxygen tension on human pre-implantation development using time-lapse monitoring. Materials and methods: Human embryos were cultured to the blastocyst stage in a time-lapse incubator (EmbryoScope™) in 20% O2 (group 1), 20% O2 for 24 hours followed by culture in 5% O2 (group 2......) or in 5% O2 (group 3). Eligible were patients with age 8 oocytes retrieved. Group 1 consisted of 120 IVF/ICSI embryos from 26 patients recruited to a study conducted to evaluate the safety of the time-lapse incubator by randomising 1:1 embryos from a patient to culture......-points for each cell division and blastocyst stages were registered until 120 hours after oocyte retrieval. Only 2PN embryos completing the first cleavage were evaluated. The groups were compared using one-way ANOVA or Kruskall-Wallis test. Estimates are reported as medians with 95% confidence intervals. Time...

  12. The First Human Cloned Embryo.

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    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)

  13. Timing of human preimplantation embryonic development is confounded by embryo origin

    DEFF Research Database (Denmark)

    Kirkegaard, Kirstine Kjær; Sundvall Germeys, Linda Karin M; Erlandsen, M.

    2016-01-01

    STUDY QUESTION To what extent do patient- and treatment-related factors explain the variation in morphokinetic parameters proposed as embryo viability markers? SUMMARY ANSWER Up to 31% of the observed variation in timing of embryo development can be explained by embryo origin, but no single facto...... by a grant from the Danish Council for Independent Research Medical Sciences. The authors declare no competing interest....... embryos from one patient as independent observations, and only very few studies that evaluate the influence from patient- and treatment-related factors on timing of development or time-lapse parameters as predictors of viability have controlled for confounding, which implies a high risk of overestimating...

  14. Correlation analysis of human embryo LeY glycan antigen expression and embryo quality.

    Science.gov (United States)

    Gu, Juan; Sui, Linlin; Ma, Yanni; Guo, Zhenzhen; Zhang, Man; Zhu, Chenyang; Cai, Zhu; Kong, Ying

    2017-07-01

    This study assessed the feasibility of using LeY glycan secretion level in human embryos as a method of judging embryo quality. Embryo culture media from patients receiving in vitro fertilization-embryo transfer was collected, and quality scores of embryos were recorded. Secretions of LeY in the culture media in different development stages (from 4-cell to 10-cell), embryos in the same development stage of the same patients (8-cell/I) and embryos in the same development stage of different patients (8-cell/I) were examined by dot-blot. Embryos were divided into a hypersecretion group and hyposecretion group, based on their LeY secretion level. The embryo quality was evaluated by clinical observations, the number which developed to D3 cell stage and the number of successful embryo transplantations. LeY secretion increased as embryos developed from 4-cell to 10-cell (PLeY of 8/I is not identical; development speed of embryos with different secretion level of LeY was also different. The number of embryos which developed to 6-cell or higher was 82.2% in the LeY hypersecretion group but only 60% in the hyposecretion group. The rate of successful transplantation was significantly higher in the hypersecretion group (71.1 vs. 40%). In conclusion, LeY glycan secretion level in human embryos is closely related to embryo quality. LeY may become a useful measure to evaluate embryo quality in the future.

  15. Human embryos commonly form abnormal nuclei during development: a mechanism of DNA damage, embryonic aneuploidy, and developmental arrest.

    Science.gov (United States)

    Kort, Daniel H; Chia, Gloryn; Treff, Nathan R; Tanaka, Akemi J; Xing, Tongji; Vensand, Lauren Bauer; Micucci, Stephanie; Prosser, Robert; Lobo, Roger A; Sauer, Mark V; Egli, Dieter

    2016-02-01

    What is the prevalence and developmental significance of morphologic nuclear abnormalities in human preimplantation embryos? Nuclear abnormalities are commonly found in human IVF embryos and are associated with DNA damage, aneuploidy, and decreased developmental potential. Early human embryonic development is complicated by genomic errors that occur after fertilization. The appearance of extra-nuclear DNA, which has been observed in IVF, may be a result of such errors. However, the mechanism by which abnormal nuclei form and the impact on DNA integrity and embryonic development is not understood. Cryopreserved human cleavage-stage embryos (n = 150) and cryopreserved blastocysts (n = 105) from clinical IVF cycles performed between 1997 and 2008 were donated for research. Fresh embryos (n = 60) of poor quality that were slated for discard were also used. Immunohistochemical, microscopic and cytogenetic analyses at different developmental stages and morphologic grades were performed. Embryos were fixed and stained for DNA, centromeres, mitotic activity and DNA damage and imaged using confocal microscopy. Rates of abnormal nuclear formation were compared between morphologically normal cleavage-stage embryos, morphologically normal blastocysts, and poor quality embryos. To control for clinical and IVF history of oocytes donors, and quality of frozen embryos within our sample, cleavage-stage embryos (n = 52) were thawed and fixed at different stages of development and then analyzed microscopically. Cleavage-stage embryos (n = 9) were thawed and all blastomeres (n = 62) were disaggregated, imaged and analyzed for karyotype. Correlations were made between microscopic and cytogenetic findings of individual blastomeres and whole embryos. The frequency of microscopic nuclear abnormalities was lower in blastocysts (5%; 177/3737 cells) than in cleavage-stage embryos (16%, 103/640 blastomeres, P < 0.05) and highest in arrested embryos (65%; 44/68 blastomeres, P < 0.05). DNA

  16. In vitro development of donated frozen-thawed human embryos in a prototype static microfluidic device: a randomized controlled trial

    NARCIS (Netherlands)

    Kieslinger, Dorit C.; Hao, Zhenxia; Vergouw, Carlijn G.; Kostelijk, Elisabeth H.; Lambalk, Cornelis B.; le Gac, Severine

    Objective: To compare the development of human embryos in microfluidic devices with culture in standard microdrop dishes, both under static conditions. Design: Prospective randomized controlled trial. Setting: In vitro fertilization laboratory. Patient(s): One hundred eighteen donated frozen-thawed

  17. [Cryopreservation of early human embryo stages].

    Science.gov (United States)

    Vökler, T; Fliess, F R

    1988-01-01

    A short review of freezing procedures applied to early human embryos is given. It is noted that human embryos survived freezing and thawing at a developmental stage of 1. cell to blastocyst. But it seems to be necessary to use for any developmental stage of early embryo a special freezing and thawing method. Embryo survival is correlated with their morphologic features where as neither age of embryos nor developmental stage were involved in freezing and thawing ability.

  18. Sourcing human embryos for embryonic stem cell lines: Problems & perspectives

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    Rajvi H Mehta

    2014-01-01

    Full Text Available 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.

  19. Distruption of retinoid and CYP systems and embryo development in marine organisms: a potential model for humans

    DEFF Research Database (Denmark)

    Tairova, Zhanna; Strand, Jakob; Jørgensen, Eva Cecilie Bonefeld

    Some environmental persistent organic pollutants (POPs) can be highly toxic and pose risk for both natural fauna populations and humans. POPs can disrupt an array of molecular and cellular mechanisms causing endocrine disruptions, cancer and teratogenic effects. Potentially, POPs can interfere...... with embryo development and reproduction. At present, there is only limited knowledge of the potential effects of dioxin-like compounds and polycyclic aromatic hydrocarbons in the Danish environment. The Ph.D. project is expected to link exposure to POPs such as dioxin-like compounds and PAHs to effects...

  20. The human embryo: ethical and legal aspects.

    Science.gov (United States)

    Knoppers, Bartha Maria; Bordet, Sylvie; Isasi, Rosario

    2009-01-01

    This paper analyses the status of the embryo in Canadian law. First, a brief overview of some ethical issues raised by research with embryos, focusing on the moral status of the embryo, is presented. A survey of the regulatory framework applicable to embryo research in Canada follows, so as to delineate the legal status of the embryo in Canada and its ethical underpinnings. A summary of applicable regulation in Germany, the United Kingdom, and the United States is also undertaken, illustrating the lack of consensus on this issue in Western countries. Finally, recent developments in stem cell research are considered, focusing on current alternatives to embryo destruction.

  1. Arabidopsis mitochondrial protein slow embryo development1 is essential for embryo development

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    Ju, Yan; Liu, Chunying; Lu, Wenwen; Zhang, Quan; Sodmergen, E-mail: sodmergn@pku.edu.cn

    2016-05-27

    The plant seeds formation are crucial parts in reproductive process in seed plants as well as food source for humans. Proper embryo development ensure viable seed formation. Here, we showed an Arabidopsis T-DNA insertion mutant slow embryo development1 (sed1) which exhibited retarded embryogenesis, led to aborted seeds. Embryo without SED1 developed slower compared to normal one and could be recognized at early globular stage by its white appearance. In later development stage, storage accumulated poorly with less protein and lipid body production. In vitro culture did not rescue albino embryo. SED1 encoded a protein targeted to mitochondria. Transmission electron microscopic analysis revealed that mitochondria developed abnormally, and more strikingly plastid failed to construct grana in time in sed1/sed1 embryo. These data indicated that SED1 is indispensable for embryogenesis in Arabidopsis, and the mitochondria may be involved in the regulation of many aspects of seed development. -- Highlights: •Arabidopsis SED1 is essential for embryo development. •The sed1 embryo accumulates less storage and has abnormal ultrastructure. •SED1 localizes to the mitochondrion.

  2. Human stem cell ethics: beyond the embryo.

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    Sugarman, Jeremy

    2008-06-05

    Human embryonic stem cell research has elicited powerful debates about the morality of destroying human embryos. However, there are important ethical issues related to stem cell research that are unrelated to embryo destruction. These include particular issues involving different types of cells used, the procurement of such cells, in vivo use of stem cells, intellectual property, and conflicts of interest.

  3. Human amniotic fluid contaminants alter thyroid hormone signalling and early brain development in Xenopus embryos

    Science.gov (United States)

    Fini, Jean-Baptiste; Mughal, Bilal B.; Le Mével, Sébastien; Leemans, Michelle; Lettmann, Mélodie; Spirhanzlova, Petra; Affaticati, Pierre; Jenett, Arnim; Demeneix, Barbara A.

    2017-03-01

    Thyroid hormones are essential for normal brain development in vertebrates. In humans, abnormal maternal thyroid hormone levels during early pregnancy are associated with decreased offspring IQ and modified brain structure. As numerous environmental chemicals disrupt thyroid hormone signalling, we questioned whether exposure to ubiquitous chemicals affects thyroid hormone responses during early neurogenesis. We established a mixture of 15 common chemicals at concentrations reported in human amniotic fluid. An in vivo larval reporter (GFP) assay served to determine integrated thyroid hormone transcriptional responses. Dose-dependent effects of short-term (72 h) exposure to single chemicals and the mixture were found. qPCR on dissected brains showed significant changes in thyroid hormone-related genes including receptors, deiodinases and neural differentiation markers. Further, exposure to mixture also modified neural proliferation as well as neuron and oligodendrocyte size. Finally, exposed tadpoles showed behavioural responses with dose-dependent reductions in mobility. In conclusion, exposure to a mixture of ubiquitous chemicals at concentrations found in human amniotic fluid affect thyroid hormone-dependent transcription, gene expression, brain development and behaviour in early embryogenesis. As thyroid hormone signalling is strongly conserved across vertebrates the results suggest that ubiquitous chemical mixtures could be exerting adverse effects on foetal human brain development.

  4. Human embryo cloning prohibited in Hong Kong.

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    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.

  5. Advances in embryo culture platforms: novel approaches to improve preimplantation embryo development through modifications of the microenvironment.

    Science.gov (United States)

    Swain, J E; Smith, G D

    2011-01-01

    The majority of research aimed at improving embryo development in vitro has focused on manipulation of the chemical environment, examining details such as energy substrate composition and impact of various growth factors or other supplements. In comparison, relatively little work has been done examining the physical requirements of preimplantation embryos and the role culture platforms or devices can play in influencing embryo development. Electronic searches were performed using keywords centered on embryo culture techniques using PUBMED through June 2010 and references were searched for additional research articles. Various approaches to in vitro embryo culture that involve manipulations of the physical culture environment are emerging. Novel culture platforms being developed examine issues such as media volume and embryo spacing. Furthermore, methods to permit dynamic embryo culture with fluid flow and embryo movement are now available, and novel culture surfaces are being tested. Although several factors remain to be studied to optimize efficiency, manipulations of the embryo culture microenvironment through novel culture devices may offer a means to improve embryo development in vitro. Reduced volume systems that reduce embryo spacing, such as the well-of-the-well approach, appear beneficial, although more work is needed to verify the source of their true benefit in human embryos. Emerging microfluidic technology appears to be a promising approach. However, along with the work on specialized culture surfaces, more information is required to determine the impact on human embryo development.

  6. Effect of co-culture human endothelial progenitor cells with porcine oocytes during maturation subsequent embryo development of parthenotes in vitro.

    Science.gov (United States)

    Lee, Seok Hee; Oh, Hyun Ju; Kim, Min Jung; Setyawan, Erif Maha Nugraha; Choi, Yoo Bin; Lee, Byeong Chun

    2018-02-14

    Human endothelial progenitor cells (EPCs) have been applied to regenerative medicine for their roles in angiogenesis as well as neovascularization, and these angiogenetic functions have beneficial effects on maturation of ovarian follicles. However, little information is available on whether EPCs on culture systems affect oocyte maturation and subsequent embryo development. Therefore, the objective of this study was to investigate the effect of EPC co-culture on porcine oocytes during in vitro maturation (IVM) and subsequent embryo development, and to examine gene expression in cumulus cells, oocytes and blastocysts. The effect of co-culture using EPC on porcine oocyte IVM was investigated. Oocytes were activated using electrical stimulation and embryo developmental competence was estimated. The expression of the genes related to cumulus expansion, oocyte maturation, embryo development and apoptosis were analyzed. In result, there was a significantly increased maturation rate in EPC group compared with control (p cultured with EPCs exhibited significantly improved blastocyst formation rates (p culture group showed significantly higher SOX2, OCT4 and NANOG levels. In conclusion, co-culturing porcine oocytes with EPCs improves their maturation by regulating genes involved in cumulus cell expansion, oocyte maturation and apoptosis. Moreover, EPC co-culture during IVM enhanced embryo development as shown by increased blastocyst formation rate and pluripotency-related gene expression. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  7. Adoption first? The disposition of human embryos.

    Science.gov (United States)

    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. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

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

    Science.gov (United States)

    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.

  9. THE EFFECT OF RECOMBINANT HUMAN LEUKEMIA INHIBITORY FACTOR (rhLIF ON IN VITRO DEVELOPMENT OF MOUSE 2-CELL EMBRYOS AND THEIR ISOLATED BLASTOMERES

    Directory of Open Access Journals (Sweden)

    MOHAMMAD AKBARI

    2004-09-01

    Full Text Available In this study effect of recombinant human leukemia inhibitory factor on invitro development of 2 cells embryos and isolated blastomeres derived from mouse 2 cell embryos were investigated. Female ICR mice that were between 8 to 10 weeks old received intraperitoneal injection of 7.5 IU of PMSG for super ovulation followed by intraperitoneal administration of 7.5 IU of HCG 48 hours later. The mice were then mated to mature ICR male mice and were checked for vaginal plugs after 13-14 hours. Mice were killed 46-48 hours after HCG injection by cervical dislocation, their oviducts were removed and flushing 2 cell embryos were collected. The zona pellucida of 2 cell embryos were removed by Acid Tyrod solution and blastomeres separated with oocyte preparation pipette and then all embryos and blastomeres were cultured in Potassium Simplex Optimized Medium (KSOM +Aminoacid (AA different amounts of rhLIF (500IU/ml, 1000IU/ml and 1500IU/ml. Some embryos and individual blastomere also were cultured without rhLIF as control group. All samples were cultured in an incubator at 370C with 0.05 CO2 for 120 hours. The rate of embryo and individual blastomeres which reached to 2 cell, 4 cell, 8 cell and 9-16 cell were the same in all groups. However in further developmental stages, morula and blastocyst between experimental and control groups were significantly different. Therefore it may be concluded that: cultivation of isolated blastomers up to the blastocyst stage with rhLIF has stimulatory effect on the preimplantation stage (morula and blastocyst but it has no stimulatory and inhibitory effects when was added to culture media at the early cleavage stage.

  10. Development of a new clinically applicable device for embryo evaluation which measures embryo oxygen consumption.

    Science.gov (United States)

    Kurosawa, Hiroki; Utsunomiya, Hiroki; Shiga, Naomi; Takahashi, Aiko; Ihara, Motomasa; Ishibashi, Masumi; Nishimoto, Mitsuo; Watanabe, Zen; Abe, Hiroyuki; Kumagai, Jin; Terada, Yukihiro; Igarashi, Hideki; Takahashi, Toshifumi; Fukui, Atsushi; Suganuma, Ryota; Tachibana, Masahito; Yaegashi, Nobuo

    2016-10-01

    Does a new system-the chip-sensing embryo respiration monitoring system (CERMs)-enable evaluation of embryo viability for potential application in a clinical IVF setting? The system enabled the oxygen consumption rate of spheroids, bovine embryos and frozen-thawed human embryos to be measured, and this rate corresponded to the developmental potential of embryos. To date, no reliable and clinically suitable objective evaluation methods for embryos are available, which circumvent the differences in inter-observer subjective view. Existing systems such as the scanning electrochemical microscopy (SECM) technique, which enables the measurement of oxygen consumption rate in embryos, need improvement in usability before they can be applied to a clinical setting. This is a prospective original research study. The feasibility of measuring the oxygen consumption rate was assessed using CERMs for 9 spheroids, 9 bovine embryos and 30 redundant frozen-thawed human embryos. The endpoints for the study were whether CERMs could detect a dissolved oxygen gradient with high sensitivity, had comparable accuracy to the SECM measuring system with improved usability, and could predict the development of an embryo to a blastocyst by measuring the oxygen consumption rate. The relationship between the oxygen consumption rate and standard morphological evaluation was also examined. We developed a new CERMs, which enables the oxygen consumption rate to be measured automatically using an electrochemical method. The device was initially used for measuring a dissolved oxygen concentration gradient in order to calculate oxygen consumption rate using nine spheroids. Next, we evaluated data correlation between the CERMs and the SECM measuring systems using nine bovine embryos. Finally, the oxygen consumption rates of 30 human embryos, which were frozen-thawed on 2nd day after fertilization, were measured by CERMs at 6, 24, 48, 72 and 96 h after thawing with standard morphological evaluation

  11. Lack of carbon air filtration impacts early embryo development.

    Science.gov (United States)

    Munch, Erika M; Sparks, Amy E; Duran, Hakan E; Van Voorhis, Bradley J

    2015-07-01

    To assess human fertilization and preimplantation embryo development in the presence and in the absence of carbon filtration This is a retrospective cohort analysis of fresh, controlled ovarian hyperstimulation cycles as well as previously cryopreserved pronuclear stage embryo transfer cycles in a single IVF center. Embryo development and cycle-based outcomes were compared among three groups: 1) when carbon filtration was present, 2) when carbon filtration was absent, and 3) when carbon filtration had been restored. A total of 524 fresh cycles and 156 cryopreserved embryo cycles were analyzed. Fertilization, cleavage, and blastocyst conversion rates for fresh cycles all declined during the period of absent carbon filtration and recovered after the restoration of carbon filtration. Cryopreserved embryos that were thawed and cultured during the period of absent filtration did not have changes in cleavage or blastocyst conversion rates compared to periods where carbon filtration was present. Clinical pregnancy and live birth rates were unchanged among the three time periods. The absence of carbon filtration in an IVF laboratory air handler is associated with poor fertilization and early embryo development for fresh cycles. Because development of previously frozen pronuclear stage embryos was unaffected, the lack of carbon filtration may preferentially affect embryos in the peri-fertilization period. Carbon filtration is an integral part to a successful human in-vitro fertilization laboratory.

  12. Embryo splitting

    Directory of Open Access Journals (Sweden)

    Karl Illmensee

    2010-04-01

    Full Text Available Mammalian embryo splitting has successfully been established in farm animals. Embryo splitting is safely and efficiently used for assisted reproduction in several livestock species. In the mouse, efficient embryo splitting as well as single blastomere cloning have been developed in this animal system. In nonhuman primates embryo splitting has resulted in several pregnancies. Human embryo splitting has been reported recently. Microsurgical embryo splitting under Institutional Review Board approval has been carried out to determine its efficiency for blastocyst development. Embryo splitting at the 6–8 cell stage provided a much higher developmental efficiency compared to splitting at the 2–5 cell stage. Embryo splitting may be advantageous for providing additional embryos to be cryopreserved and for patients with low response to hormonal stimulation in assisted reproduction programs. Social and ethical issues concerning embryo splitting are included regarding ethics committee guidelines. Prognostic perspectives are presented for human embryo splitting in reproductive medicine.

  13. Rape embryogenesis. III. Embryo development in time

    Directory of Open Access Journals (Sweden)

    Teresa Tykarska

    2014-01-01

    Full Text Available It was found that the growth curve of the rape embryo axis is of triple sigmoid type. Embryo growth occurs in 3 phases corresponding to 3 different periods of development. Phase I includes growth of the apical cell up to it's division into two layers of octants. Phase II comprises the increase of the spherical proembryo to the change of its symmetry from radial to bilateral. Phase III includes, growth of the embryo from the heart stage up to the end of embryogenesis. In each phase the relative growth rate increases drastically and then diminishes. The differences in growth intensity during the same phase are several-fold. The growth intensity maximum of the embryo axis occurs in phase II. The phasic growth intensity maxima occur: in phase I during apical cell elongation, :before its division, and in phases II and III in the periods of cell division ;growth in globular and torpedo-shaped -shaped embryos.

  14. Potential of human twin embryos generated by embryo splitting in assisted reproduction and research.

    Science.gov (United States)

    Noli, Laila; Ogilvie, Caroline; Khalaf, Yacoub; Ilic, Dusko

    2017-03-01

    Embryo splitting or twinning has been widely used in veterinary medicine over 20 years to generate monozygotic twins with desirable genetic characteristics. The first human embryo splitting, reported in 1993, triggered fierce ethical debate on human embryo cloning. Since Dolly the sheep was born in 1997, the international community has acknowledged the complexity of the moral arguments related to this research and has expressed concerns about the potential for reproductive cloning in humans. A number of countries have formulated bans either through laws, decrees or official statements. However, in general, these laws specifically define cloning as an embryo that is generated via nuclear transfer (NT) and do not mention embryo splitting. Only the UK includes under cloning both embryo splitting and NT in the same legislation. On the contrary, the Ethics Committee of the American Society for Reproductive Medicine does not have a major ethical objection to transferring two or more artificially created embryos with the same genome with the aim of producing a single pregnancy, stating that 'since embryo splitting has the potential to improve the efficacy of IVF treatments for infertility, research to investigate the technique is ethically acceptable'. Embryo splitting has been introduced successfully to the veterinary medicine several decades ago and today is a part of standard practice. We present here an overview of embryo splitting experiments in humans and non-human primates and discuss the potential of this technology in assisted reproduction and research. A comprehensive literature search was carried out using PUBMED and Google Scholar databases to identify studies on embryo splitting in humans and non-human primates. 'Embryo splitting' and 'embryo twinning' were used as the keywords, alone or in combination with other search phrases relevant to the topics of biology of preimplantation embryos. A very limited number of studies have been conducted in humans and non-human

  15. Oviduct: roles in fertilization and early embryo development.

    Science.gov (United States)

    Li, Shuai; Winuthayanon, Wipawee

    2017-01-01

    Animal oviducts and human Fallopian tubes are a part of the female reproductive tract that hosts fertilization and pre-implantation development of the embryo. With an increasing understanding of roles of the oviduct at the cellular and molecular levels, current research signifies the importance of the oviduct on naturally conceived fertilization and pre-implantation embryo development. This review highlights the physiological conditions within the oviduct during fertilization, environmental regulation, oviductal fluid composition and its role in protecting embryos and supplying nutrients. Finally, the review compares different aspects of naturally occurring fertilization and assisted reproductive technology (ART)-achieved fertilization and embryo development, giving insight into potential areas for improvement in this technology. © 2017 Society for Endocrinology.

  16. Radial extracorporeal shock wave treatment harms developing chicken embryos

    Science.gov (United States)

    Kiessling, Maren C.; Milz, Stefan; Frank, Hans-Georg; Korbel, Rüdiger; Schmitz, Christoph

    2015-01-01

    Radial extracorporeal shock wave treatment (rESWT) has became one of the best investigated treatment modalities for cellulite, including the abdomen as a treatment site. Notably, pregnancy is considered a contraindication for rESWT, and concerns have been raised about possible harm to the embryo when a woman treated with rESWT for cellulite is not aware of her pregnancy. Here we tested the hypothesis that rESWT may cause serious physical harm to embryos. To this end, chicken embryos were exposed in ovo to various doses of radial shock waves on either day 3 or day 4 of development, resembling the developmental stage of four- to six-week-old human embryos. We found a dose-dependent increase in the number of embryos that died after radial shock wave exposure on either day 3 or day 4 of development. Among the embryos that survived the shock wave exposure a few showed severe congenital defects such as missing eyes. Evidently, our data cannot directly be used to draw conclusions about potential harm to the embryo of a pregnant woman treated for cellulite with rESWT. However, to avoid any risks we strongly recommend applying radial shock waves in the treatment of cellulite only if a pregnancy is ruled out. PMID:25655309

  17. The endometrial factor in human embryo implantation

    NARCIS (Netherlands)

    Boomsma, C.M.

    2009-01-01

    The studies presented in this thesis aimed to explore the role of the endometrium in the implantation process. At present, embryo implantation is the major rate-limiting step for success in fertility treatment. Clinicians have sought to develop clinical interventions aimed at enhancing implantation

  18. Biosensors for detecting stress in developing embryos

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  20. Morphometric analysis of human embryos to predict developmental competence

    DEFF Research Database (Denmark)

    Ziebe, Søren

    2013-01-01

    Morphometric and morphokinetic approaches toward embryo quality assessment have for many years been difficult due to technical limitations. Today, with improvements in laboratory techniques and subsequent quality, we have a better understanding of the morphometric and kinetics of embryo development...

  1. Addressing the ethical issues raised by synthetic human entities with embryo-like features

    NARCIS (Netherlands)

    Aach, John; Lunshof, Jeantine; Iyer, Eswar; Church, George M.

    2017-01-01

    The "14-day rule" for embryo research stipulates that experiments with intact human embryos must not allow them to develop beyond 14 days or the appearance of the primitive streak. However, recent experiments showing that suitably cultured human pluripotent stem cells can self organize and

  2. The Early Stages of Heart Development: Insights from Chicken Embryos

    Directory of Open Access Journals (Sweden)

    Johannes G. Wittig

    2016-04-01

    Full Text Available The heart is the first functioning organ in the developing embryo and a detailed understanding of the molecular and cellular mechanisms involved in its formation provides insights into congenital malformations affecting its function and therefore the survival of the organism. Because many developmental mechanisms are highly conserved, it is possible to extrapolate from observations made in invertebrate and vertebrate model organisms to humans. This review will highlight the contributions made through studying heart development in avian embryos, particularly the chicken. The major advantage of chick embryos is their accessibility for surgical manipulation and functional interference approaches, both gain- and loss-of-function. In addition to experiments performed in ovo, the dissection of tissues for ex vivo culture, genomic, or biochemical approaches is straightforward. Furthermore, embryos can be cultured for time-lapse imaging, which enables tracking of fluorescently labeled cells and detailed analysis of tissue morphogenesis. Owing to these features, investigations in chick embryos have led to important discoveries, often complementing genetic studies in mice and zebrafish. As well as including some historical aspects, we cover here some of the crucial advances made in understanding early heart development using the chicken model.

  3. Differences in gene expression profiles between human preimplantation embryos cultured in two different IVF culture media.

    Science.gov (United States)

    Kleijkers, Sander H M; Eijssen, Lars M T; Coonen, Edith; Derhaag, Josien G; Mantikou, Eleni; Jonker, Martijs J; Mastenbroek, Sebastiaan; Repping, Sjoerd; Evers, Johannes L H; Dumoulin, John C M; van Montfoort, Aafke P A

    2015-10-01

    Is gene expression in human preimplantation embryos affected by the medium used for embryo culture in vitro during an IVF treatment? Six days of in vitro culture of human preimplantation embryos resulted in medium-dependent differences in expression level of genes involved in apoptosis, protein degradation, metabolism and cell-cycle regulation. Several human studies have shown an effect of culture medium on embryo development, pregnancy outcome and birthweight. However, the underlying mechanisms in human embryos are still unknown. In animal models of human development, it has been demonstrated that culture of preimplantation embryos in vitro affects gene expression. In humans, it has been found that culture medium affects gene expression of cryopreserved embryos that, after thawing, were cultured in two different media for 2 more days. In a multicenter trial, women were randomly assigned to two culture medium groups [G5 and human tubal fluid (HTF)]. Data on embryonic development were collected for all embryos. In one center, embryos originating from two pronuclei (2PN) zygotes that were not selected for transfer or cryopreservation on Day 2 or 3 because of lower morphological quality, were cultured until Day 6 and used in this study, if couples consented. Ten blastocysts each from the G5 and HTF study groups, matched for fertilization method, maternal age and blastocyst quality, were selected and their mRNA was isolated and amplified. Embryos were examined individually for genome-wide gene expression using Agilent microarrays and PathVisio was used to identify the pathways that showed a culture medium-dependent activity. Expression of 951 genes differed significantly (P differences observed between the study groups are caused by factors that we did not investigate. Extrapolation of these results to embryos used for transfer demands caution as in the present study embryos that were not selected for either embryo transfer or cryopreservation have been used for the

  4. Toxicity testing of human assisted reproduction devices using the mouse embryo assay.

    NARCIS (Netherlands)

    Punt-Van der Zalm, J.P.; Hendriks, J.C.M.; Westphal, J.R.; Kremer, J.A.M.; Teerenstra, S.; Wetzels, A.M.M.

    2009-01-01

    Systems to assess the toxicity of materials used in human assisted reproduction currently lack efficiency and/or sufficient discriminatory power. The development of 1-cell CBA/B6 F1 hybrid mouse embryos to blastocysts, expressed as blastocyst rate (BR), is used to measure toxicity. The embryos were

  5. In vitro maturation, fertilization, embryo development & clinical outcome of human metaphase-I oocytes retrieved from stimulated intracytoplasmic sperm injection cycles

    Directory of Open Access Journals (Sweden)

    Cristina Álvarez

    2013-01-01

    Full Text Available Background & objectives: The major cause of fertilisation failure after ICSI is failure of the oocyte to initiate the biochemical processes necessary for activation. This inability could be ascribed to cytoplasmic immaturity of those gametes even if they had reached nuclear maturity. The activation of a mature oocyte is characterised by release from metaphase II (MII arrest and extrusion of the second polar body, followed by pro-nuclear formation. The aim of this study was to evaluate the fate of in vitro matured (IVM metaphase I (MI oocytes subjected to intracytoplasmic sperm injection (ICSI at different time intervals after extrusion of the first polar body (1PB in in vitro fertilization (IVF cycles. Methods: A total of 8030 oocytes were collected from 1400 ICSI cycles, 5504 MII at the time of cumulus retrieval. Four hundred eight metaphase II (MII (27.1% matured to MII after in vitro culture for 2-26 h and 5389 sibling MII in the moment of oocyte denudation were injected. On the other hand, 49 ICSI cycles containing only MI oocytes at retrieval were injected at three different time intervals after reaching the MII. The intervals were as follows: 2-6 h (n=10, 8-11 h (n=4 and 23-26 h (n=10. Fertilization and development potential were evaluated in both studies. Results: Fertilization, embryo cleavage and quality were significantly lower in IVM MI compared to MII at time of denudation. Pregnancy rate was higher in group MII. Pregnancy was achieved in three embryo transfers when ICSI was performed within 2-6 h (group I and 8-11 h (group II after PB extrusion. One pregnancy was obtained in group I and a healthy neonate was born. Interpretation & conclusions: Immature oocytes from women whose ovaries have been stimulated could be matured, fertilized by ICSI, cleaved in vitro and to give rise to a live birth. However, the developmental competence of embryos derived from immature oocytes is reduced, compared with sibling in vivo matured oocytes

  6. The ethics of cloning and human embryo research.

    Science.gov (United States)

    Saran, Madeleine

    2002-01-01

    The successful cloning experiments that led to Dolly in 1997 have raised many ethical and policy questions. This paper will focus on cloning research in human embryonic cells. The possible gains of the research will be judged against the moral issues of doing research on a person. This paper concludes that while the embryo has some moral status, its moral status is outweighed by the multitude of benefits that embryonic stem cell research will bring to humanity. Policy suggestions are given for dealing with this new and developing field of stem cell research.

  7. Human cloning and embryo research: the 2003 John J. Conley Lecture on medical ethics.

    Science.gov (United States)

    George, Robert P

    2004-01-01

    The author, a member of the U.S. President's Council on Bioethics, discusses ethical issues raised by human cloning, whether for purposes of bringing babies to birth or for research purposes. He first argues that every cloned human embryo is a new, distinct, and enduring organism, belonging to the species Homo sapiens, and directing its own development toward maturity. He then distinguishes between two types of capacities belonging to individual organisms belonging to this species, an immediately exerciseable capacity and a basic natural capacity that develops over time. He argues that it is the second type of capacity that is the ground for full moral respect, and that this capacity (and its concomitant degree of respect) belongs to cloned human embryos no less than to adult human beings. He then considers and rejects counter-arguments to his position, including the suggestion that the capacity of embryos is equivalent to the capacity of somatic cells, that full human rights are afforded only to human organisms with functioning brains, that the possibility of twinning diminishes the moral status of embryos, that the fact that people do not typically mourn the loss of early embryos implies that they have a diminished moral status, that the fact that early spontaneous abortions occur frequently diminishes the moral status of embryos, and that his arguments depend upon a concept of ensoulment. He concludes that if the moral status of cloned human embryos is equivalent to that of adults, then public policy should be based upon this assumption.

  8. Closure of the vertebral canal in human embryos and fetuses

    NARCIS (Netherlands)

    Mekonen, Hayelom K.; Hikspoors, Jill P. J. M.; Mommen, Greet; Kruepunga, Nutmethee; Köhler, S. Eleonore; Lamers, Wouter H.

    2017-01-01

    The vertebral column is the paradigm of the metameric architecture of the vertebrate body. Because the number of somites is a convenient parameter to stage early human embryos, we explored whether the closure of the vertebral canal could be used similarly for staging embryos between 7 and 10weeks of

  9. Chromosomal mosaicism in human preimplantation embryos : a systematic review

    NARCIS (Netherlands)

    van Echten-Arends, Jannie; Mastenbroek, Sebastiaan; Sikkema-Raddatz, Birgit; Korevaar, Johanna C.; Heineman, Maas Jan; van der Veen, Fulco; Repping, Sjoerd

    2011-01-01

    BACKGROUND: Although chromosomal mosaicism in human preimplantation embryos has been described for almost two decades, its exact prevalence is still unknown. The prevalence of mosaicism is important in the context of preimplantation genetic screening in which the chromosomal status of an embryo is

  10. Human embryo-conditioned medium stimulates in vitro endometrial angiogenesis

    NARCIS (Netherlands)

    Kapiteijn, K.; Koolwijk, P.; Weiden, R.M.F. van der; Nieuw Amerongen, G. van; Plaisier, M.; Hinsbergh, V.W.M. van; Helmerhorst, F.M.

    2006-01-01

    Objective: Successful implantation and placentation depend on the interaction between the endometrium and the embryo. Angiogenesis is crucial at this time. In this article we investigate the direct influence of the human embryo on in vitro endometrial angiogenesis. Design: In vitro study. Setting:

  11. Expression of microRNAs in bovine and human pre-implantation embryo culture media

    Science.gov (United States)

    Kropp, Jenna; Salih, Sana M.; Khatib, Hasan

    2014-01-01

    MicroRNAs (miRNA) are short non-coding RNAs which act to regulate expression of genes driving numerous cellular processes. These RNAs are secreted within exosomes from cells into the extracellular environment where they may act as signaling molecules. In addition, they are relatively stable and are specifically expressed in association to certain cancers making them strong candidates as biological markers. Moreover, miRNAs have been detected in body fluids including urine, milk, saliva, semen, and blood plasma. However, it is unknown whether they are secreted by embryonic cells into the culture media. Given that miRNAs are expressed throughout embryonic cellular divisions and embryonic genome activation, we hypothesized that they are secreted from the embryo into the extracellular environment and may play a role in the developmental competence of bovine embryos. To test this hypothesis, bovine embryos were cultured individually from day 5 to day 8 of development in an in vitro fertilization system and gene expression of 5 miRNAs was analyzed in both embryos and culture media. Differential miRNA gene expression was observed between embryos that developed to the blastocyst stage and those that failed to develop from the morula to blastocyst stage, deemed degenerate embryos. MiR-25, miR-302c, miR-196a2, and miR-181a expression was found to be higher in degenerate embryos compared to blastocyst embryos. Interestingly, these miRNAs were also found to be expressed in the culture media of both bovine and human pre-implantation embryos. Overall, our results show for the first time that miRNAs are secreted from pre-implantation embryos into culture media and that miRNA expression may correlate with developmental competence of the embryo. Expression of miRNAs in in vitro culture media could allow for the development of biological markers for selection of better quality embryos and for subsequent successful pregnancy. PMID:24795753

  12. Expression of microRNAs in bovine and human pre-implantation embryo culture media

    Directory of Open Access Journals (Sweden)

    Jenna eKropp

    2014-04-01

    Full Text Available MicroRNAs (miRNA are short non-coding RNAs which act to regulate expression of genes driving numerous cellular processes. These RNAs are secreted within exosomes from cells into the extracellular environment where they may act as signaling molecules. In addition, they are relatively stable and are specifically expressed in association to certain cancers making them strong candidates as biological markers. Moreover, miRNAs have been detected in body fluids including urine, milk, saliva, semen, and blood plasma. However, it is unknown whether they are secreted by embryonic cells into the culture media. Given that miRNAs are expressed throughout embryonic cellular divisions and embryonic genome activation, we hypothesized that they are secreted from the embryo into the extracellular environment and may play a role in the developmental competence of bovine embryos. To test this hypothesis, bovine embryos were cultured individually from day 5 to day 8 of development in an in vitro fertilization system and gene expression of 5 miRNAs was analyzed in both embryos and culture media. Differential miRNA gene expression was observed between embryos that developed to the blastocyst stage and those that failed to develop from the morula to blastocyst stage, deemed degenerate embryos. MiR-25, mir-302c, miR-196a2, and miR-181a expression was found to be higher in degenerate embryos compared to blastocyst embryos. Interestingly, these miRNAs were also found to be expressed in the culture media of both bovine and human pre-implantation embryos. Overall, our results show for the first time that miRNAs are secreted from pre-implantation embryos into culture media and that miRNA expression may correlate with developmental competence of the embryo. Expression of miRNAs in in vitro culture media could allow for the development of biological markers for selection of better quality embryos and for subsequent successful pregnancy.

  13. Humanization of Learning: A Mission in Embryo

    Science.gov (United States)

    Fritz, John O.

    1971-01-01

    The Humanization of Learning Mission is directed toward developing program alternatives which would accept as critical priorities for schooling the development of self awareness, empathic understanding, and the enhancement of social responsibility. (Author)

  14. In vitro maturation is associated with increased early embryo arrest without impairing morphokinetic development of useable embryos progressing to blastocysts.

    Science.gov (United States)

    Walls, M L; Ryan, J P; Keelan, J A; Hart, R

    2015-08-01

    Does polycystic ovarian syndrome (PCOS) or in vitro maturation (IVM) treatment affect embryo development events and morphokinetic parameters after time-lapse incubation? There was an increase in some abnormal phenotypic events in PCOS-IVM embryos as well as an increase in early arrest of PCOS-IVM and PCOS-ICSI embryos; however, IVM treatment or PCOS status did not alter morphokinetic development of embryos suitable for transfer of vitrification. IVM has been less successful than standard IVF in terms of clinical pregnancy, implantation and live birth rates. There is currently no information available about the development of IVM embryos according to time-lapse analysis. This article represents a prospective case-control study. The study involved 93 participants who underwent 93 treatment cycles. Cycles were completed between January 2013 and July 2014. Participants were recruited for the study at Fertility Specialists of WA and Fertility Specialists South, Perth, Western Australia. Of the PCOS diagnosed patients, 32 underwent IVM treatment (PCOS-IVM) and 23 had standard ICSI treatment (PCOS-ICSI). There were 38 patients without PCOS who underwent standard ICSI treatment comprising the control group (control-ICSI). The PCOS-IVM group showed significantly more embryos with multinucleated two cells (P = 0.041), multinucleated four cells (P = 0.001) and uneven two cells (P = 0.033) compared with the control-ICSI group, but not the PCOS-ICSI group. There were no significant differences in the rates of any abnormal events between the PCOS-ICSI and control-ICSI groups. Embryo arrest between Days 2 and 3 was higher in the PCOS-IVM and PCOS-ICSI groups compared with the control-ICSI group (P events from embryos generated using this approach for patients diagnosed with PCOS and shows that embryos generated from IVM have an increased rate of early embryo arrest, however; morphokinetic development is not impaired in embryos that progress to the useable blastocyst stage. The

  15. The Impact of Biopsy on Human Embryo Developmental Potential during Preimplantation Genetic Diagnosis

    Directory of Open Access Journals (Sweden)

    Danilo Cimadomo

    2016-01-01

    Full Text Available Preimplantation Genetic Diagnosis and Screening (PGD/PGS for monogenic diseases and/or numerical/structural chromosomal abnormalities is a tool for embryo testing aimed at identifying nonaffected and/or euploid embryos in a cohort produced during an IVF cycle. A critical aspect of this technology is the potential detrimental effect that the biopsy itself can have upon the embryo. Different embryo biopsy strategies have been proposed. Cleavage stage blastomere biopsy still represents the most commonly used method in Europe nowadays, although this approach has been shown to have a negative impact on embryo viability and implantation potential. Polar body biopsy has been proposed as an alternative to embryo biopsy especially for aneuploidy testing. However, to date no sufficiently powered study has clarified the impact of this procedure on embryo reproductive competence. Blastocyst stage biopsy represents nowadays the safest approach not to impact embryo implantation potential. For this reason, as well as for the evidences of a higher consistency of the molecular analysis when performed on trophectoderm cells, blastocyst biopsy implementation is gradually increasing worldwide. The aim of this review is to present the evidences published to date on the impact of the biopsy at different stages of preimplantation development upon human embryos reproductive potential.

  16. The Impact of Biopsy on Human Embryo Developmental Potential during Preimplantation Genetic Diagnosis

    Science.gov (United States)

    Cimadomo, Danilo; Capalbo, Antonio; Ubaldi, Filippo Maria; Scarica, Catello; Palagiano, Antonio; Canipari, Rita; Rienzi, Laura

    2016-01-01

    Preimplantation Genetic Diagnosis and Screening (PGD/PGS) for monogenic diseases and/or numerical/structural chromosomal abnormalities is a tool for embryo testing aimed at identifying nonaffected and/or euploid embryos in a cohort produced during an IVF cycle. A critical aspect of this technology is the potential detrimental effect that the biopsy itself can have upon the embryo. Different embryo biopsy strategies have been proposed. Cleavage stage blastomere biopsy still represents the most commonly used method in Europe nowadays, although this approach has been shown to have a negative impact on embryo viability and implantation potential. Polar body biopsy has been proposed as an alternative to embryo biopsy especially for aneuploidy testing. However, to date no sufficiently powered study has clarified the impact of this procedure on embryo reproductive competence. Blastocyst stage biopsy represents nowadays the safest approach not to impact embryo implantation potential. For this reason, as well as for the evidences of a higher consistency of the molecular analysis when performed on trophectoderm cells, blastocyst biopsy implementation is gradually increasing worldwide. The aim of this review is to present the evidences published to date on the impact of the biopsy at different stages of preimplantation development upon human embryos reproductive potential. PMID:26942198

  17. Where does New Zealand stand on permitting research on human embryos?

    Science.gov (United States)

    Jones, D Gareth

    2014-08-01

    In many respects New Zealand has responded to the assisted reproductive technologies (ARTs) as positively as many comparable societies, such as Australia and the UK. Consequently, in vitro fertilisation (IVF) and pre-implantation genetic diagnosis (PGD) are widely available, as is non-commercial surrogacy utilising IVF. These developments have been made possible by the Human Assisted Reproductive Technology (HART) Act 2004, overseen by its two committees, the Advisory Committee on Assisted Reproductive Technology (ACART) and the Ethics Committee (ECART). However, New Zealand stands apart from many of these other societies by the lack of permission for scientists to conduct research using human embryos. There is no doubt this reflects strongly held viewpoints on the part of some that embryos should be protected and not exploited. Legitimate as this stance is, the resulting situation is problematic when IVF is already designated as an established procedure. This is because the development of IVF involved embryo research, and continuing improvements in procedures depend upon ongoing embryo research. While prohibition of research on human embryos gives the impression of protecting embryos, it fails to do this and also fails to enhance the health and wellbeing of children born using IVF. This situation will not be rectified until research is allowed on human embryos.

  18. The spectrum of DNA damage in human sperm assessed by single cell gel electrophoresis (Comet assay) and its relationship to fertilization and embryo development.

    Science.gov (United States)

    Morris, I D; Ilott, S; Dixon, L; Brison, D R

    2002-04-01

    The integrity of sperm DNA is important for the success of natural or assisted fertilization, as well as normal development of the embryo, fetus and child. ICSI, by bypassing sperm selection mechanisms, increases the risk of transmitting damaged DNA and the significance of this requires investigation. DNA damage in sperm from an unselected group of 60 men undergoing IVF treatment was measured by single cell gel electrophoresis (Comet assay) and correlated with semen and treatment cycle parameters. Wide spectra of sperm DNA damage were found both within and between men but no specific subgroups were identified. Semen and treatment cycle parameters were not different in men grouped according to high or low sperm DNA damage. However, regression analysis showed that DNA damage was positively associated with age (29-44 years), abnormal sperm and motility and negatively associated with sperm concentration. In ICSI cycles DNA damage was positively associated with impairment of post-fertilization embryo cleavage. This study contributes to the evidence of DNA damage within sperm. High loads of DNA damage measured by the Comet assay were predictive of failure of embryo development after ICSI. As it is likely that sperm with DNA damage contributed to successful fertilization and in-vitro development, potential adverse effects remain to be clarified.

  19. Addition of ascorbate during cryopreservation stimulates subsequent embryo development.

    Science.gov (United States)

    Lane, Michelle; Maybach, Jeffery M; Gardner, David K

    2002-10-01

    Embryo development following cryopreservation is reduced compared with fresh embryos. One of the traumas that cryopreservation imparts on embryos is an increase in oxidative stress. Therefore, this study investigated the effects of the addition of the antioxidant ascorbate to the cryopreservation solutions on subsequent embryo development. Mouse embryos at the 2-cell and blastocyst stages were either slow-frozen or vitrified in solutions containing either no ascorbate or 0.1 or 0.5 mmol/l ascorbate. The effects on the levels of hydrogen peroxide and subsequent embryo development and physiology were assessed. Addition of ascorbate to the cryopreservation solutions reduced the levels of hydrogen peroxide in embryos. Furthermore, addition of 0.1 mmol/l ascorbate significantly enhanced inner cell mass development in blastocysts. Embryos cryopreserved with ascorbate had significantly lower levels of lactate dehydrogenase leakage, and increased rates of metabolism compared with those cryopreserved in the absence of ascorbate. The benefits of ascorbate were significantly greater in embryos that were slow-frozen compared with those that were vitrified. These data indicate that the addition of 0.1 mmol/l ascorbate to the cryopreservation solutions for the mammalian embryo would be of significant value.

  20. Ovarian hyperstimulation syndrome and prophylactic human embryo cryopreservation: analysis of reproductive outcome following thawed embryo transfer

    Directory of Open Access Journals (Sweden)

    Sills Eric

    2008-11-01

    Full Text Available Abstract Objective To review utilisation of elective embryo cryopreservation in the expectant management of patients at risk for developing ovarian hyperstimulation syndrome (OHSS, and report on reproductive outcome following transfer of thawed embryos. Materials and methods Medical records were reviewed for patients undergoing IVF from 2000–2008 to identify cases at risk for OHSS where cryopreservation was electively performed on all embryos at the 2 pn stage. Patient age, total number of oocytes retrieved, number of 2 pn embryos cryopreserved, interval between retrieval and thaw/transfer, number (and developmental stage of embryos transferred (ET, and delivery rate after IVF were recorded for all patients. Results From a total of 2892 IVF cycles undertaken during the study period, 51 IVF cases (1.8% were noted where follicle number exceeded 20 and pelvic fluid collection was present. Elective embryo freeze was performed as OHSS prophylaxis in each instance. Mean (± SD age of these patients was 32 ± 3.8 yrs. Average number of oocytes retrieved in this group was 23 ± 8.7, which after fertilisation yielded an average of 14 ± 5.7 embryos cryopreserved per patient. Thaw and ET was performed an average of 115 ± 65 d (range 30–377 d after oocyte retrieval with a mean of 2 ± 0.6 embryos transferred. Grow-out to blastocyst stage was achieved in 88.2% of cases. Delivery/livebirth rate was 33.3% per initiated cycle and 43.6% per transfer. Non-transferred blastocysts remained in cryostorage for 24 of 51 patients (46.1% after ET, with an average of 3 ± 3 blastocysts refrozen per patient. Conclusion OHSS prophylaxis was used in 1.8% of IVF cycles at this institution; no serious OHSS complications were encountered during the study period. Management based on elective 2 pn embryo cryopreservation with subsequent thaw and grow-out to blastocyst stage for transfer did not appear to compromise embryo viability or overall reproductive outcome. For

  1. Embryo development and embryo transfer in the European mink (Mustela lutreola), an endangered mustelid species.

    Science.gov (United States)

    Amstislavsky, S; Kizilova, E; Ternovskaya, Y; Zudova, G; Lindeberg, H; Aalto, J; Valtonen, M

    2006-01-01

    The European mink is an endangered Mustelidae species and thus requires effective conservation measures, although little is known about reproduction in this species. In particular, preimplantation development has not been studied and, therefore, embryonic development and the growth of embryos was documented in the present study for European mink using light and fluorescent microscopy. Embryos develop in the oviducts and then migrate into the uterus on Day 6 post coitum (p.c.) at the morula stage. Embryos expanded as blastocysts from Day 7 until implantation on Day 12 p.c. Based on these findings, the use of embryo transfer for a conservation programme for the European mink was evaluated. Embryos were flushed from European mink resource females and transferred into the uterine horns of recipient hybrid females (honoriks and nohoriks). These hybrids were obtained by mating European polecat males with European mink females and vice versa. A total of 40 embryos was transferred and 20 live kits were born. The rates of pre- and postnatal survival were 50% and 70%, respectively. Both male and female offspring were lighter at birth in the embryo transfer group compared with naturally born controls, but there was no difference at 3 months of age.

  2. Retrograde tubal transfer of human embryos.

    Science.gov (United States)

    Risquez, F; Boyer, P; Rolet, F; Magnani, M; Guichard, A; Cedard, L; Zorn, J R

    1990-02-01

    This preliminary study was designed to evaluate retrograde cannulation of the Fallopian tubes up to the isthmo-interstitial junction using the new technique of tubal embryo stage transfer (TEST). Follicular aspiration was performed under the guidance of a vaginal ultrasound probe in 51 women treated with GnRH + HMG. The oocytes retrieved were inseminated in vitro with 50,000 motile spermatozoa and kept in Menezo B2 medium without serum, at 37 degrees C, in an atmosphere of air + 5% CO2. The eggs were checked 24 and 36 h after insemination. No fertilization occurred in 23 patients. Cleaved embryos were obtained in the 28 other patients. One to seven embryos at the 2-4-cell stage were transferred with the 'Baudelocque Black Catheter' (BBC) into one tube and spare embryos were frozen. Five pregnancies occurred after retrograde TEST, for a pregnancy rate of 9.8% per cycle and 17.9% per transfer. One patient has given birth to a normal full-term baby. One singleton and one twin pregnancy are ongoing (8 months in June 1989). The other two pregnancies were ectopic.

  3. In vitro culture of mouse embryos amniotic fluid ID human

    African Journals Online (AJOL)

    1989-07-15

    Jul 15, 1989 ... Human amniotic fluid was compared with Ham's F-10 culture medium as a possible alternative for use in in vitro fertilisation. The cleavage success of mouse embryos in human amniotic fluid (experimental group) was 92% compared with 86% in. Ham's F-10 medium. It is concluded that human amniotic.

  4. Optimization of a novel nylon mesh container for human embryo ultrarapid vitrification.

    Science.gov (United States)

    Nakashima, Akira; Ino, Nao; Kusumi, Maki; Ohgi, Shirei; Ito, Megumu; Horikawa, Takashi; Nakagawa, Koji; Saito, Takakazu; Kamura, Toshiharu; Saito, Hidekazu

    2010-05-01

    To evaluate the efficacy of a nylon mesh container in vitrification of human embryos and to determine the optimal osmotic pressure of the initial thawing solution. Retrospective analysis. National Center for Child Health and Development, Tokyo, Japan. Infertile patients undergoing either in vitro fertilization or intracytoplasmic sperm injection in our hospital. Embryos, at the cleavage stage, were cryopreserved using the vitrification method in either a plastic straw or a nylon mesh container. The embryos were thawed using an initial osmotic pressure of either 0.5 M or 1.0 M sucrose with subsequent step-wise dilution. After thawing, the embryos were transferred to the uterus. Survival rate of blastomeres, embryo survival rate, implantation, and pregnancy rates, cancellation rate because of embryo damage. Use of nylon mesh and the 1.0 M sucrose thawing solution significantly improved blastomere survival rate (98.0 +/- 1.0%, mean +/- SEM), pregnancy rate (41.0%) and implantation rate (32.3%). Vitrification using a nylon mesh container and subsequent thawing in a 1.0 M sucrose solution is an easy and inexpensive method that improves the reliability of embryo cryopreservation of embryos without adverse effects on clinical outcomes. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  5. Fruit, seed and embryo development of different cassava (Manihot ...

    African Journals Online (AJOL)

    Fruit, seed and embryo developments of different cassava (Manihot esculenta Crantz) genotypes, as well as embryo rescue, were investigated. The fruits of three genotypes after uncontrolled open pollination presented the same progressive development with similar sizes at different stages. There are large differences in ...

  6. Fruit, seed and embryo development of different cassava (Manihot ...

    African Journals Online (AJOL)

    SAM

    2014-03-24

    Mar 24, 2014 ... Fruit, seed and embryo developments of different cassava (Manihot esculenta Crantz) genotypes, as well as embryo rescue, were investigated. The fruits of three genotypes after uncontrolled open pollination presented the same progressive development with similar sizes at different stages. There are large.

  7. Embryo sac development in some South African Lantana species (Verbenaceae

    Directory of Open Access Journals (Sweden)

    J. J. Spies

    1984-12-01

    Full Text Available Evidence that the South African Lantana camara L. complex only produces sexual embryo sacs is provided. It is shown that the archesporium occasionally divides mitotically and that both archesporia form tetrads. The chalazal megaspore of one tetrad and the micropylar megaspore of the second tetrad develop into Polygonum type embryo sacs. L. rugosa Thunb. also forms Polygonum type embryo sacs. The L. rugosa embryo sac has a much more densely packed cytoplasm, smaller vacuole and the position of the polar nuclei differs from that of the L. camara embryo sac. It is possible to distinguish between  L. camara and  L. rugosa on their embryo sac morphology alone.

  8. Developmental stages in human embryos: revised and new measurements.

    Science.gov (United States)

    O'Rahilly, Ronan; Müller, Fabiola

    2010-01-01

    The staging of human embryos, as distinct from seriation, depends on a morphological scheme devised by Streeter and completed by O'Rahilly, who proposed the term Carnegie stages. To avoid misconceptions and errors, and to place new findings in perspective, it is necessary to summarize the essentials of the Carnegie system: (1) Twenty-three stages cover the embryonic period, i. e. the first 8 postfertilizational weeks of development. (2) The system is based on internal as well as external features, and the use of only external criteria is subject to serious limitations. For example, precise delineation of stages 19-23 and of the embryonic-fetal transition depends on histological examination. (3) Prenatal measurements are not an integral component of the staging system, and hence a stage should never be assigned merely on the basis of embryonic length. A 20-mm embryo, for example, could belong to any of three stages. Measurements, however, are important for the assessment of age, and very few measurements are available for staged embryos. Presented here and based on accurate staging are the maximum diameter of the chorionic sac, the crown-heel length, the greatest length exclusive of the lower limbs, the biparietal diameter, the head circumference, the length of the hindbrain, the total length of the brain, and the lengths of the limbs as well as of their segments, including the foot length. (4) Prenatal ages are also not an integral part of the staging system and hence a stage should never be assigned merely on the basis of prenatal age. Ages, however, are of clinical importance and their estimate has been rendered more precise by accurate timing of fertilization followed by ultrasonography. Prenatal age is postfertilizational and hence some 2 weeks less than the postmenstrual interval. The term gestational age is ambiguous and should be discarded. Presented here is a new graph showing proposed estimates of age in relation to stages and based on current information

  9. Early embryo mortality in natural human reproduction: What the data say

    Science.gov (United States)

    Jarvis, Gavin E.

    2017-01-01

    How many human embryos die between fertilisation and birth under natural conditions? It is widely accepted that natural human embryo mortality is high, particularly during the first weeks after fertilisation, with total prenatal losses of 70% and higher frequently claimed. However, the first external sign of pregnancy occurs two weeks after fertilisation with a missed menstrual period, and establishing the fate of embryos before this is challenging. Calculations are additionally hampered by a lack of data on the efficiency of fertilisation under natural conditions. Four distinct sources are used to justify quantitative claims regarding embryo loss: (i) a hypothesis published by Roberts & Lowe in The Lancet  is widely cited but has no practical quantitative value; (ii) life table analyses give consistent assessments of clinical pregnancy loss, but cannot illuminate losses at earlier stages of development; (iii) studies that measure human chorionic gonadotrophin (hCG) reveal losses in the second week of development and beyond, but not before; and (iv) the classic studies of Hertig and Rock offer the only direct insight into the fate of human embryos from fertilisation under natural conditions. Re-examination of Hertig’s data demonstrates that his estimates for fertilisation rate and early embryo loss are highly imprecise and casts doubt on the validity of his numerical analysis. A recent re-analysis of hCG study data concluded that approximately 40-60% of embryos may be lost between fertilisation and birth, although this will vary substantially between individual women. In conclusion, natural human embryo mortality is lower than often claimed and widely accepted. Estimates for total prenatal mortality of 70% or higher are exaggerated and not supported by the available data. PMID:28580126

  10. Heteroparental blastocyst production from microsurgically corrected tripronucleated human embryos.

    Science.gov (United States)

    Escribá, María-José; Martín, Julio; Rubio, Carmen; Valbuena, Diana; Remohí, José; Pellicer, Antonio; Simón, Carlos

    2006-12-01

    To prove the efficiency of identification and removal of one of the surplus paternal pronuclei in dispermic IVF zygotes to obtain heteroparental blastocysts. Experimental. One hundred fourteen tripronucleated (3PN) embryos from conventional IVF. After informed and signed consent, the patients from Instituto Valenciano Infertilidad (IVI), Valencia, donated their abnormally fertilized embryos. Seventy-two embryos were diploidized by microsurgical removal of the pronucleus located at the farthest position to the second polar body. Forty-two 3PN embryos served as controls. Survival and correction rate; in vitro development up to the blastocyst stage; X, Y, and 18 chromosome determination by triple fluorescent in situ hybridization and, inheritance analysis for 10 polymorphic repeat regions using polymerase chain reaction (PCR) amplification and sequencing. Seventy-eight percent of 3PN zygotes (56/72) survived manipulation and eventually 51 zygotes had two pronuclei (71%). Forty-one percent of manipulated embryos progressed in vitro to the blastocyst stage (21/51). Fluorescent in situ hybridization analysis performed on eight manipulated embryos confirmed their diploid state; all four controls were triploid. Heteroparental inheritances were also confirmed in four of six manipulated embryos. Heteroparental blastocysts can be derived from corrected dispermic zygotes.

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

    Science.gov (United States)

    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.

  12. Early embryo development in Fucus distichus is auxin sensitive

    Science.gov (United States)

    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.

  13. [TSA improve transgenic porcine cloned embryo development and transgene expression].

    Science.gov (United States)

    Kong, Qing-Ran; Zhu, Jiang; Huang, Bo; Huan, Yan-Jun; Wang, Feng; Shi, Yong-Qian; Liu, Zhong-Feng; Wu, Mei-Ling; Liu, Zhong-Hua

    2011-07-01

    Uncompleted epigenetic reprogramming is attributed to the low efficiency of producing transgenic cloned animals. Histone modification associated with epigenetics can directly influence the embryo development and transgene expression. Trichostatin A (TSA), as an inhibitor of histone deacetylase, can change the status of histone acetylation, improve somatic cell reprogramming, and enhance cloning efficiency. TSA prevents the chromatin structure from being condensed, so that transcription factor could binds to DNA sequence easily and enhance transgene expression. Our study established the optimal TSA treatment on porcine donor cells and cloned embryos, 250 nmol/L, 24 h and 40 nmol/L, 24 h, respectively. Furthermore, we found that both the cloned embryo and the donor cell treated by TSA resulted in the highest development efficiency. Meanwhile, TSA can improve transgene expression in donor cell and cloned embryo. In summary, TSA can significantly improve porcine reconstructed embryo development and transgene expression.

  14. IVF and embryo transfer: historical origin and development.

    Science.gov (United States)

    Biggers, John D

    2012-08-01

    IVF and embryo transfer for the treatment of human infertility has now resulted in the birth of over 4 million babies. The technique did not arise as a quantum event but was built on the efforts of many earlier workers in the fields of reproductive endocrinology and development. One should remember the famous saying of Isaac Newton: 'If I have seen further than most, it is because I have stood on the shoulder's of giants'. Ethical and moral issues have always arisen when investigators study early mammalian development, particularly human development. This paper documents these earlier studies and also draws attention to the ethical and moral arguments that inevitably arose. Copyright © 2012 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  15. Status of the human embryo: Philosophical Foundations from Phenomenology

    Directory of Open Access Journals (Sweden)

    Maria Emilia de Oliveira Schpallir Silva

    2017-10-01

    Full Text Available Given the difficulty in demonstrating the moment of ontogenesis in which personalization takes place, we sought to define, from a philosophic point of view, the nature of the human embryo regarding its individuality, using Phenomenology, specifically reflections of philosophers Bourghet and Merleau-Ponty on the embryo. Although the statement of their individuality does not entail ethical content in itself, from the point of view of ethical responsibility, it is an extremely important fact to be considered in the bioethical reflection about the moment of ontogeny from which human life must (ethical duty be protected.

  16. Biopsy of human morula-stage embryos: outcome of 215 IVF/ICSI cycles with PGS.

    Directory of Open Access Journals (Sweden)

    Elena E Zakharova

    Full Text Available Preimplantation genetic diagnosis (PGD is commonly performed on biopsies from 6-8-cell-stage embryos or blastocyst trophectoderm obtained on day 3 or 5, respectively. Day 4 human embryos at the morula stage were successfully biopsied. Biopsy was performed on 709 morulae from 215 ICSI cycles with preimplantation genetic screening (PGS, and 3-7 cells were obtained from each embryo. The most common vital aneuploidies (chromosomes X/Y, 21 were screened by fluorescence in situ hybridization (FISH. No aneuploidy was observed in 72.7% of embryos, 91% of those developed to blastocysts. Embryos were transferred on days 5-6. Clinical pregnancy was obtained in 32.8% of cases, and 60 babies were born. Patients who underwent ICSI/PGS treatment were compared with those who underwent standard ICSI treatment by examining the percentage of blastocysts, pregnancy rate, gestational length, birth height and weight. No significant differences in these parameters were observed between the groups. Day 4 biopsy procedure does not adversely affect embryo development in vitro or in vivo. The increased number of cells obtained by biopsy of morulae might facilitate diagnostic screening. There is enough time after biopsy to obtain PGD results for embryo transfer on day 5-6 in the current IVF cycle.

  17. An in vivo culture system for human embryos using an encapsulation technology: a pilot study

    Science.gov (United States)

    Blockeel, C.; Mock, P.; Verheyen, G.; Bouche, N.; Le Goff, Ph.; Heyman, Y.; Wrenzycki, C.; Höffmann, K.; Niemann, H.; Haentjens, P.; de Los Santos, M.J.; Fernandez-Sanchez, M.; Velasco, M.; Aebischer, P.; Devroey, P.; Simón, C.

    2009-01-01

    BACKGROUND Animal studies have demonstrated better embryo development in vivo than in vitro. This pilot study tested the feasibility of using a novel in utero culture system (IUCS) to obtain normal human fertilization and embryo development. METHODS The IUCS device comprised a perforated silicone hollow tube. The study included 13 patients (<36 years) undergoing a first intracytoplasmic sperm injection (ICSI) treatment and 167 metaphase II oocytes in three groups. In Group 1, 1–2 h after ICSI, sibling oocytes were assigned to IUCS or conventional in vitro culture. The device was retrieved on Day 1, and all zygotes were cultured in vitro till Day 5. In Group 2, fertilized oocytes were assigned on Day 1, embryos retrieved on Day 3 and all embryos cultured till Day 5. In Group 3, after Day 0 assignment, embryos were retrieved on Day 3 for blastomere biopsy and fluorescence in situ hybridization (FISH) and cultured until Day 5. The highest quality blastocysts were transferred on Day 5. RESULTS Fertilization and embryo development were comparable in the in vitro and IUCS arms, with a tendency towards better embryo quality in the IUCS. FISH analysis in Group 3 revealed more normal embryos using the IUCS (P = 0.049). Three clinical pregnancies and live births were obtained: two from the IUCS arm and one from the in vitro arm. CONCLUSIONS Our pilot study shows that this new IUCS appears to be feasible and safe, supporting normal fertilization, embryo development and normal chromosomal segregation. Furthermore, live births are possible after the transient presence of a silicone device in the uterus.Clinicaltrials.gov: NCT00480103. PMID:19273881

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

    Science.gov (United States)

    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. Symposium: innovative techniques in human embryo viability assessment. Soluble human leukocyte antigen-G and pregnancy success.

    Science.gov (United States)

    Warner, Carol M; Lampton, Paula W; Newmark, Judith A; Cohen, Jacques

    2008-10-01

    Non-invasive methods of assessing embryo quality are critical for pregnancy success following IVF or intracytoplasmic sperm injection (ICSI). The addition of new non-invasive morphological and biochemical analyses may further improve pregnancy success, allowing the transfer of a single embryo, thereby reducing the risks involved in multiple births following IVF/ICSI. The presence of a protein, soluble human leukocyte antigen-G (sHLA-G), in embryo cultures has been suggested as a way to non-invasively predict embryo quality and pregnancy success, especially when used in conjunction with current embryo quality assessment methods. Detection of sHLA-G in embryo culture medium has been correlated with pregnancy success in 12 studies, but three studies were not able to detect sHLA-G. This is a review of the literature on sHLA-G detection in IVF/ICSI, and reasons are proposed for the reported discrepancies, as well as guidelines for reporting of data in future studies. Furthermore, it is suggested that the use of an HLA-G transgenic mouse model would advance understanding of the mechanism of action of sHLA-G in preimplantation embryos and its correlation to embryo health and viability. Research on a mouse model, combined with clinical studies, should enable the development of a fast and reliable method for utilizing sHLA-G detection to improve pregnancy success after IVF/ICSI.

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

    Science.gov (United States)

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

  1. Human embryo research and the language of moral uncertainty.

    Science.gov (United States)

    Cheshire, William P

    2004-01-01

    In bioethics as in the sciences, enormous discussions often concern the very small. Central to public debate over emerging reproductive and regenerative biotechnologies is the question of the moral status of the human embryo. Because news media have played a prominent role in framing the vocabulary of the debate, this study surveyed the use of language reporting on human embryo research in news articles spanning a two-year period. Terminology that devalued moral status-for example, the descriptors things, property, tissue, or experimental material -was found to outnumber fivefold those that affirmed any degree of moral status above that of inanimate cellular matter; for example, living, human life, or human being. A quarter of the articles failed to note that the embryos under discussion were human. These findings confirm that even among scientific and philosophical experts a diversity of opinion exists on society's moral obligations to nascent human life. The skewed linguistic distribution also indicates a distinct bias. Concerned readers should take notice when any category of humanity becomes subject to prejudicial and disparaging language and the value of vulnerable human life is trivialized alongside sensational assertions of anticipated medical cures. The responsibility for holding the media to a higher standard of truth and fairness falls to us all.

  2. Comparative transcriptomic analysis of developing cotton cotyledons and embryo axis.

    Science.gov (United States)

    Jiao, Xiaoming; Zhao, Xiaochun; Zhou, Xue-Rong; Green, Allan G; Fan, Yunliu; Wang, Lei; Singh, Surinder P; Liu, Qing

    2013-01-01

    As a by product of higher value cotton fibre, cotton seed has been increasingly recognised to have excellent potential as a source of additional food, feed, biofuel stock and even a renewable platform for the production of many diverse biological molecules for agriculture and industrial enterprises. The large size difference between cotyledon and embryo axis that make up a cotton seed results in the under-representation of embryo axis gene transcript levels in whole seed embryo samples. Therefore, the determination of gene transcript levels in the cotyledons and embryo axes separately should lead to a better understanding of metabolism in these two developmentally diverse tissues. A comparative study of transcriptome changes between cotton developing cotyledon and embryo axis has been carried out. 17,384 unigenes (20.74% of all the unigenes) were differentially expressed in the two adjacent embryo tissues, and among them, 7,727 unigenes (44.45%) were down-regulated and 9,657 unigenes (55.55%) were up-regulated in cotyledon. Our study has provided a comprehensive dataset that documents the dynamics of the transcriptome at the mid-maturity of cotton seed development and in discrete seed tissues, including embryo axis and cotyledon tissues. The results showed that cotton seed is subject to many transcriptome variations in these two tissue types and the differential gene expression between cotton embryo axis and cotyledon uncovered in our study should provide an important starting point for understanding how gene activity is coordinated during seed development to make a seed. Further, the identification of genes involved in rapid metabolite accumulation stage of seed development will extend our understanding of the complex molecular and cellular events in these developmental processes and provide a foundation for future studies on the metabolism, embryo differentiation of cotton and other dicot oilseed crops.

  3. Comparative transcriptomic analysis of developing cotton cotyledons and embryo axis.

    Directory of Open Access Journals (Sweden)

    Xiaoming Jiao

    Full Text Available BACKGROUND: As a by product of higher value cotton fibre, cotton seed has been increasingly recognised to have excellent potential as a source of additional food, feed, biofuel stock and even a renewable platform for the production of many diverse biological molecules for agriculture and industrial enterprises. The large size difference between cotyledon and embryo axis that make up a cotton seed results in the under-representation of embryo axis gene transcript levels in whole seed embryo samples. Therefore, the determination of gene transcript levels in the cotyledons and embryo axes separately should lead to a better understanding of metabolism in these two developmentally diverse tissues. RESULTS: A comparative study of transcriptome changes between cotton developing cotyledon and embryo axis has been carried out. 17,384 unigenes (20.74% of all the unigenes were differentially expressed in the two adjacent embryo tissues, and among them, 7,727 unigenes (44.45% were down-regulated and 9,657 unigenes (55.55% were up-regulated in cotyledon. CONCLUSIONS: Our study has provided a comprehensive dataset that documents the dynamics of the transcriptome at the mid-maturity of cotton seed development and in discrete seed tissues, including embryo axis and cotyledon tissues. The results showed that cotton seed is subject to many transcriptome variations in these two tissue types and the differential gene expression between cotton embryo axis and cotyledon uncovered in our study should provide an important starting point for understanding how gene activity is coordinated during seed development to make a seed. Further, the identification of genes involved in rapid metabolite accumulation stage of seed development will extend our understanding of the complex molecular and cellular events in these developmental processes and provide a foundation for future studies on the metabolism, embryo differentiation of cotton and other dicot oilseed crops.

  4. Xenotransplantation of human adipose-derived stem cells in zebrafish embryos.

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    Jin Li

    Full Text Available Zebrafish is a widely used animal model with well-characterized background in developmental biology. The fate of human adipose-derived stem cells (ADSCs after their xenotransplantation into the developing embryos of zebrafish is unknown. Therefore, human ADSCs were firstly isolated, and then transduced with lentiviral vector system carrying a green fluorescent protein (GFP reporter gene, and followed by detection of their cell viability and the expression of cell surface antigens. These GFP-expressing human ADSCs were transplanted into the zebrafish embryos at 3.3-4.3 hour post-fertilization (hpf. Green fluorescent signal, the proliferation and differentiation of human ADSCs in recipient embryos were respectively examined using fluorescent microscopy and immunohistochemical staining. The results indicated that human ADSCs did not change their cell viability and the expression levels of cell surface antigens after GFP transduction. Microscopic examination demonstrated that green fluorescent signals of GFP expressed in the transplanted cells were observed in the embryos and larva fish at post-transplantation. The positive staining of Ki-67 revealed the survival and proliferation of human ADSCs in fish larvae after transplantation. The expression of CD105 was observable in the xenotransplanted ADSCs, but CD31 expression was undetectable. Therefore, our results indicate that human ADSCs xenotransplanted in the zebrafish embryos not only can survive and proliferate at across-species circumstance, but also seem to maintain their undifferentiation status in a short term. This xenograft model of zebrafish embryos may provide a promising and useful technical platform for the investigation of biology and physiology of stem cells in vivo.

  5. On developing a thesis for Reproductive Endocrinology and Infertility fellowship: a case study of ultra-low (2%) oxygen tension for extended culture of human embryos.

    Science.gov (United States)

    Kaser, Daniel J

    2017-03-01

    Fellows in Reproductive Endocrinology and Infertility training are expected to complete 18 months of clinical, basic, or epidemiological research. The goal of this research is not only to provide the basis for the thesis section of the oral board exam but also to spark interest in reproductive medicine research and to provide the next generation of physician-scientists with a foundational experience in research design and implementation. Incoming fellows often have varying degrees of training in research methodology and, likewise, different career goals. Ideally, selection of a thesis topic and mentor should be geared toward defining an "answerable" question and building a practical skill set for future investigation. This contribution to the JARG Young Investigator's Forum revisits the steps of the scientific method through the lens of one recently graduated fellow and his project aimed to test the hypothesis that "sequential oxygen exposure (5% from days 1 to 3, then 2% from days 3 to 5) improves blastocyst yield and quality compared to continuous exposure to 5% oxygen among human preimplantation embryos."

  6. Dose of recombinant FSH and oestradiol concentration on day of HCG affect embryo development kinetics

    DEFF Research Database (Denmark)

    Muñoz, Manuel; Cruz, María; Humaidan, Peter

    2012-01-01

    During follicular growth, the follicle is exposed to an almost ever-changing composition of isoforms of FSH and LH, which causes a number of different and divergent biological effects. Through a time-lapse system, embryo kinetics were examined following the use of FSH only (recombinant FSH, r......FSH) and gonadotrophins containing LH activity (human menopausal gonadotrophin, HMG, and FSH+HMG) in oocyte donors. No significant differences were seen between the three groups (for rFSH, HMG and rFSH+HMG, t2 was 27.8h, 27.9h and 27.5h respectively). Moreover, although embryos obtained with rFSH showed an increase...... in the proportions of optimal timings of development, the differences observed were not significant, as shown by the percentages of embryos inside/outside these kinetic variables. In contrast, for gonadotrophin dosage and oestradiol concentration, this study observed differences in embryo development kinetics...

  7. Blocking connexin channels improves embryo development of vitrified bovine blastocysts

    OpenAIRE

    Ortiz Escribano, Nerea; Szymanska, Katarzyna; BOL, MELISSA; Vandenberghe, Lynn; Decrock, Elke; Van Poucke, Mario; Peelman, Luc; Van den Abbeel, Etienne; Soom, Ann Van; Leybaert, Luc

    2017-01-01

    Connexins (Cxs) are required for normal embryo development and implantation. They form gap junctions (GJs) connecting the cytoplasm of adjacent cells and hemichannels (HCs), which are normally closed but open in response to stress conditions. Excessive HC opening is detrimental for cell function and may lead to cell death. We found that hatching of in vitro-produced bovine embryos, matured in serum-containing conditions, was significantly improved when vitrification/warming was done in the pr...

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

    Science.gov (United States)

    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.

  9. A cutin fluorescence pattern in developing embryos of some angiosperms

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    Ewa Szczuka

    2014-01-01

    Full Text Available A cuticle visualized by auramine O fluorescence appears on the developing embryos of 9 species belonging to Cruciferae, Caryophyllaceae, Plantaginaceae, Linaceae and Papilionaceae. In the investigated species the formation and extent of fluorescing and non-fluorescing embryonic areas follow a similar pattern. At first the cutin fluorescing layer is formed on the apical part of the proembryo without delimited protoderm. This layer extends and at the late globular stage envelops the embryo proper, except for a cell adjoining the suspensor. Fluorescing cutin persists during the heart stage but disappears from the torpedo embryo. During these stages there is no cutine fluorescence on suspensorial cells. Continuous cutin fluorescence appears again on the surface of the whole embryo by the late torpedo stage. Then fluorescence disappears from the radicular part of U-shaped embryos, but persists on the shoot apex, cotyledons and at least on the upper part of hypocotyl. It is assumed that polarization and nutrition of the embryo may be influenced by cuticular changes.

  10. Single-site neural tube closure in human embryos revisited.

    Science.gov (United States)

    de Bakker, Bernadette S; Driessen, Stan; Boukens, Bastiaan J D; van den Hoff, Maurice J B; Oostra, Roelof-Jan

    2017-10-01

    Since the multi-site closure theory was first proposed in 1991 as explanation for the preferential localizations of neural tube defects, the closure of the neural tube has been debated. Although the multi-site closure theory is much cited in clinical literature, single-site closure is most apparent in literature concerning embryology. Inspired by Victor Hamburgers (1900-2001) statement that "our real teacher has been and still is the embryo, who is, incidentally, the only teacher who is always right", we decided to critically review both theories of neural tube closure. To verify the theories of closure, we studied serial histological sections of 10 mouse embryos between 8.5 and 9.5 days of gestation and 18 human embryos of the Carnegie collection between Carnegie stage 9 (19-21 days) and 13 (28-32 days). Neural tube closure was histologically defined by the neuroepithelial remodeling of the two adjoining neural fold tips in the midline. We did not observe multiple fusion sites in neither mouse nor human embryos. A meta-analysis of case reports on neural tube defects showed that defects can occur at any level of the neural axis. Our data indicate that the human neural tube fuses at a single site and, therefore, we propose to reinstate the single-site closure theory for neural tube closure. We showed that neural tube defects are not restricted to a specific location, thereby refuting the reasoning underlying the multi-site closure theory. Clin. Anat. 30:988-999, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  11. Fertilization and Embryo Development of Fresh and Cryopreserved Sibling Oocytes

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    Robert F. Casper

    2010-01-01

    Full Text Available Background: Oocyte cryopreservation is potentially the best way to preserve female fertility forunmarried women or young girls at risk of losing ovarian function. The aim of this study was tocompare fertilization and embryo development in frozen-thawed oocytes to their fresh siblings inwomen undergoing in vitro fertilization (IVF and embryo transfer (ET.Materials and Methods: Eleven infertile women undergoing infertility treatment, between theages of 24 to 37 years (mean ± SD = 31.6 ± 3.5, were included in this study. Mature oocytesfrom each patient were randomized into cryopreserved and fresh groups prior to intracytoplasmicsperm injection (ICSI. One hundred and thirty nine oocytes were retrieved, of which 105 were atmetaphase II (MII. Forty- five fresh MII oocytes were kept in culture whereas their sibling 60 MIIoocytes were cryopreserved using a slow cooling protocol. The frozen oocytes remained in LN2for 2 hours before thawing. ICSI was performed 1-2 hours after thawing for frozen oocytes and 4-5hours after retrieval for fresh oocytes. Fertilization and embryo development were compared.Results: Following thawing, 31 oocytes (51.6 % survived and 22 fertilized (79% while 32 freshoocytes fertilized upon ICSI (71%. The mean ± SE scores for embryos developing from frozenthawedoocytes were significantly lower at 48 and 72 hours post-ICSI than for embryos resultingfrom fresh oocytes (p<0.05.Conclusion: Our data demonstrated that oocyte freezing resulted in acceptable survival ratesfollowing cryopreservation, and similar fertilization rates following ICSI as compared to the freshsibling oocytes. However the number of blastomeres and the embryo quality on day three wassuperior in embryos from fresh oocytes when compared to the frozen oocytes.

  12. The Effects of Progesterone on Oocyte Maturation and Embryo Development

    Directory of Open Access Journals (Sweden)

    Saeed Zavareh

    2013-01-01

    Full Text Available Oocyte maturation and embryo development are controlled by intra-ovarian factors suchas steroid hormones. Progesterone (P4 exists in the follicular fluid that contributes tonormal mammalian ovarian function and has several critical functions during embryodevelopment and implantation, including endometrial receptivity, embryonic survivalduring gestation and transformation of the endometrial stromal cells to decidual cells.It is well known that the physiological effects of P4 during the pre-implantation stages ofsome mammal’s embryos are mediated by P4 receptors and their gene expression is determined.The effects of P4 on oocytes and embryo development have been assessed bysome investigations, with contradictory results. P4, a dominant steroid in follicular fluidat approximately 18 hours after the luteinizing hormone (LH surge may have a criticalrole in maturation of oocytes at the germinal stage. However, it has been shown that differentconcentrations of P4 could not improve in vitro maturation rates of germinal vesicles(GV in cumulus oocyte complexes (COCs and cumulus denuded oocytes (CDOs.Culture media supplemented with P4 significantly improved mouse embryo development.In addition, an in vivo experimental design has shown high blastocyst survival andimplantation rates in P4-treated mice.In this review we explain some of the findings that pertain to the effects of P4 onoocyte maturation and embryo development both in vitro and in vivo.

  13. Expression of cathepsin K in the human embryo and fetus.

    Science.gov (United States)

    Haeckel, C; Krueger, S; Buehling, F; Broemme, D; Franke, K; Schuetze, A; Roese, I; Roessner, A

    1999-10-01

    Cathepsin K is a protease with high collagenolytic and elastinolytic activity. Its cellular expression was previously thought to be restricted to osteoclasts and osteoclast-mediated bone resorption. In this study, the expression of cathepsin K in the human embryo and fetus was demonstrated by immunohistochemistry, in situ hybridization, and by Northern blotting of fetal tissue extracts. Besides osteoclasts and chondroclasts and their precursors, epithelial cells of various organ systems expressed significant amounts of this enzyme. Respiratory and gastrointestinal mucosa, including bile duct epithelia and urothelia, showed high levels of cathepsin K expression. With the exception of the urothelium, showing a more homogenous expression pattern, the protease was usually accentuated in the surface cell layers of pithelia. In summary, these findings in the human embryo and early fetus demonstrated a significant expression of cathepsin K in different epithelial cell types besides osteoclasts. The functional aspects of cathepsin K expression in nonosteoclastic cells and potential conclusions on physiological and pathological conditions in the embryo-fetal or adult organism remain to be investigated. Dev Dyn 1999;216:89-95. Copyright 1999 Wiley-Liss, Inc.

  14. Near-infrared laser irradiation improves the development of mouse pre-implantation embryos.

    Science.gov (United States)

    Yokoo, Masaki; Mori, Miho

    2017-05-27

    The aim of the present study was to assess the effects of near-infrared laser irradiation on the in vitro development of mouse embryos. Female ICR mice were superovulated with pregnant mare serum gonadotropin and human chorionic gonadotropin (hCG), and mated with male mice. Two-cell stage embryos were collected 40 h after administering hCG and cultured in M16 medium. Two-cell embryos (0 h after culture), 8-cell embryos (approx. 30 h after culture), morula (approx. 48 h after culture), and blastocysts (approx. 73 h after culture) were irradiated at 904 nm for 60 s. These embryos were cultured in a time-lapse monitoring system and the timing of blastocyst hatching was evaluated. Some of the irradiated blastocysts were transferred to the uterine horns of pseudopregnant recipients immediately after irradiation. Pregnancy rates, and offspring growth and fertility, were evaluated. Near-infrared laser irradiation increased the speed of in vitro mouse embryo development. In irradiated blastocysts, hatching was faster than in control (non-irradiated) blastocysts (18.4 vs. 28.2 h, P infrared laser irradiation improves the quality of mouse embryo development in vitro, and increases the live birth rate without affecting the normality of the offspring. Thus, the near-infrared laser method may enhance the quality of embryos and contribute to improvements in reproductive technologies in mammals. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Filial cannibalism improves survival and development of beaugregory damselfish embryos.

    Science.gov (United States)

    Payne, Adam G; Smith, Carl; Campbell, Andrew C

    2002-01-01

    Cannibalism of small numbers of offspring by a parent has been proposed as an adaptive parental strategy, by providing energy to support parental care. However, there are few empirical studies to support this hypothesis. We conducted field and laboratory experiments to investigate partial filial cannibalism in Stegastes leucostictus, a coral reef fish with paternal care. Partial cannibalism was shown to be common, and males were found to remove developing embryos from throughout a clutch in a random pattern, rather than in the more aggregated pattern seen during embryo predation. Males that received a diet supplement grew faster than control males, but did not engage in less cannibalism. Also, males did not concentrate cannibalism on early embryonic stages with the highest energetic value. Experimental reduction of embryo densities was found to significantly increase embryo development rate and survival from egg deposition to hatching, and experimental reduction of oxygen levels significantly increased rates of partial filial cannibalism by males. Artificial spawning sites with low oxygen levels were avoided by spawning females, and cannibalism rates by males were higher. We propose that partial filial cannibalism serves as an adaptive parental strategy to low oxygen levels in S. leucostictus by increasing the hatching success of embryos. PMID:12396483

  16. The effects of strontium on skeletal development in zebrafish embryo.

    Science.gov (United States)

    Pasqualetti, Sara; Banfi, Giuseppe; Mariotti, Massimo

    2013-10-01

    The strontium is an alkaline earth metal found in nature as trace element. Chemically similar to calcium, it is known to be involved in the human bone mineral metabolism. The strontium ranelate has been approved in therapy as drug with both anti-resorption and anabolic effects on bone tissues. Since few data in vivo are available, we used Danio rerio as animal model to evaluate the effects of strontium on skeletal development. First, toxicity assay performed on zebrafish embryos estimated the LC50 around 6mM. Since several zebrafish bones are formed from cartilage mineralization, we evaluated whether strontium affects cartilage development during embryogenesis. Strontium does not perturb the development of the cartilage tissues before the endochondral osteogenesis takes place. About the mineralization process, we evidentiated an increase of vertebral mineralization respect to controls at lower strontium concentrations whereas higher concentration inhibited mineral deposition in dose dependent fashion. Our results evidentiated, in addition, that the calcium/strontium rate but not the absolute level of strontium modulates the mineralization process during embryonic osteogenesis. Zebrafish represents an excellent animal model to study the role of micronutrients in the development of the tissues/organs because the ions are not absorbed by intestine but assumed by skin diffusion. Copyright © 2013 Elsevier GmbH. All rights reserved.

  17. Improving metabolic health in obese male mice via diet and exercise restores embryo development and fetal growth.

    Directory of Open Access Journals (Sweden)

    Nicole O McPherson

    Full Text Available Paternal obesity is now clearly associated with or causal of impaired embryo and fetal development and reduced pregnancy rates in humans and rodents. This appears to be a result of reduced blastocyst potential. Whether these adverse embryo and fetal outcomes can be ameliorated by interventions to reduce paternal obesity has not been established. Here, male mice fed a high fat diet (HFD to induce obesity were used, to determine if early embryo and fetal development is improved by interventions of diet (CD and/or exercise to reduce adiposity and improve metabolism. Exercise and to a lesser extent CD in obese males improved embryo development rates, with increased cell to cell contacts in the compacting embryo measured by E-cadherin in exercise interventions and subsequently, increased blastocyst trophectoderm (TE, inner cell mass (ICM and epiblast cell numbers. Implantation rates and fetal development from resulting blastocysts were also improved by exercise in obese males. Additionally, all interventions to obese males increased fetal weight, with CD alone and exercise alone, also increasing fetal crown-rump length. Measures of embryo and fetal development correlated with paternal measures of glycaemia, insulin action and serum lipids regardless of paternal adiposity or intervention, suggesting a link between paternal metabolic health and subsequent embryo and fetal development. This is the first study to show that improvements to metabolic health of obese males through diet and exercise can improve embryo and fetal development, suggesting such interventions are likely to improve offspring health.

  18. Composition of commercial media used for human embryo culture.

    Science.gov (United States)

    Morbeck, Dean E; Krisher, Rebecca L; Herrick, Jason R; Baumann, Nikola A; Matern, Dietrich; Moyer, Thomas

    2014-09-01

    To determine the composition of commercially available culture media and test whether differences in composition are biologically relevant in a murine model. Experimental laboratory study. University-based laboratory. Cryopreserved hybrid mouse one-cell embryos were used in experiments. Amino acid, organic acid, ions, and metal content were determined for two different lots of media from Cook, In Vitro Care, Origio, Sage, Vitrolife, Irvine CSC, and Global. To determine whether differences in the composition of these media are biologically relevant, mouse one-cell embryos were thawed and cultured for 120 hours in each culture media at 5% and 20% oxygen in the presence or absence of protein in an EmbryoScope time-lapse incubator. The compositions of seven culture media were analyzed for concentrations of 39 individual amino acids, organic acids, ions, and elements. Blastocyst rates and cell cycle timings were calculated at 96 hours of culture, and the experiments were repeated in triplicate. Of the 39 analytes, concentrations of glucose, lactate, pyruvate, amino acids, phosphate, calcium, and magnesium were present in variable concentrations, likely reflecting differences in the interpretation of animal studies. Essential trace elements, such as copper and zinc, were not detected. Mouse embryos failed to develop in one culture medium and were differentially affected by oxygen in two other media. Culture media composition varies widely, with differences in pyruvate, lactate, and amino acids especially notable. Blastocyst development was culture media dependent and showed an interaction with oxygen concentration and presence of protein. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  19. Pollination and embryo development in Brassica rapa L. in microgravity

    Science.gov (United States)

    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.

  20. Chromosomal abnormalities and embryo development in recurrent miscarriage couples.

    Science.gov (United States)

    Rubio, C; Simón, C; Vidal, F; Rodrigo, L; Pehlivan, T; Remohí, J; Pellicer, A

    2003-01-01

    Chromosomal abnormalities are an important cause of spontaneous abortion and recurrent miscarriage (RM). Therefore, we have analysed the incidence of chromosomal abnormalities and embryo development in patients with RM. Preimplantation genetic diagnosis (PGD) was performed on 71 couples with RM and 28 couples undergoing PGD for sex-linked diseases (control group). Chromosomes 13, 16, 18, 21, 22, X and Y were analysed by fluorescence in-situ hybridization. The implantation rate in RM patients was 28% and three patients (13%) miscarried. The percentage of abnormal embryos was significantly increased (P abnormal in 19 cycles (22.1%) and repeated PGD cycles yielded similar rates of chromosomal abnormalities in 14 couples. Anomalies for chromosomes 16 and 22 were significantly higher (P abnormal embryos (61.7 versus 24.9%; P chromosomally abnormal embryos, of which some are able to develop to the blastocyst stage. IVF plus PGD is an important step in the management of these couples, but the technique has to move towards a full chromosome analysis.

  1. Early embryo mortality in natural human reproduction: What the data say [version 2; referees: 1 approved, 2 approved with reservations

    Directory of Open Access Journals (Sweden)

    Gavin E. Jarvis

    2017-06-01

    Full Text Available How many human embryos die between fertilisation and birth under natural conditions? It is widely accepted that natural human embryo mortality is high, particularly during the first weeks after fertilisation, with total prenatal losses of 70% and higher frequently claimed. However, the first external sign of pregnancy occurs two weeks after fertilisation with a missed menstrual period, and establishing the fate of embryos before this is challenging. Calculations are additionally hampered by a lack of data on the efficiency of fertilisation under natural conditions. Four distinct sources are used to justify quantitative claims regarding embryo loss: (i a hypothesis published by Roberts & Lowe in The Lancet  is widely cited but has no practical quantitative value; (ii life table analyses give consistent assessments of clinical pregnancy loss, but cannot illuminate losses at earlier stages of development; (iii studies that measure human chorionic gonadotrophin (hCG reveal losses in the second week of development and beyond, but not before; and (iv the classic studies of Hertig and Rock offer the only direct insight into the fate of human embryos from fertilisation under natural conditions. Re-examination of Hertig’s data demonstrates that his estimates for fertilisation rate and early embryo loss are highly imprecise and casts doubt on the validity of his numerical analysis. A recent re-analysis of hCG study data concluded that approximately 40-60% of embryos may be lost between fertilisation and birth, although this will vary substantially between individual women. In conclusion, natural human embryo mortality is lower than often claimed and widely accepted. Estimates for total prenatal mortality of 70% or higher are exaggerated and not supported by the available data.

  2. Dynamics of maternal and paternal effects on embryo and seed development in wild radish (Raphanus sativus)

    Science.gov (United States)

    Diggle, P. K.; Abrahamson, N. J.; Baker, R. L.; Barnes, M. G.; Koontz, T. L.; Lay, C. R.; Medeiros, J. S.; Murgel, J. L.; Shaner, M. G. M.; Simpson, H. L.; Wu, C. C.; Marshall, D. L.

    2010-01-01

    Background and Aims Variability in embryo development can influence the rate of seed maturation and seed size, which may have an impact on offspring fitness. While it is expected that embryo development will be under maternal control, more controversial hypotheses suggest that the pollen donor and the embryo itself may influence development. These latter possibilities are, however, poorly studied. Characteristics of 10-d-old embryos and seeds of wild radish (Raphanus sativus) were examined to address: (a) the effects of maternal plant and pollen donor on development; (b) the effects of earlier reproductive events (pollen tube growth and fertilization) on embryos and seeds, and the influence of embryo size on mature seed mass; (c) the effect of water stress on embryos and seeds; (d) the effect of stress on correlations of embryo and seed characteristics with earlier and later reproductive events and stages; and (e) changes in maternal and paternal effects on embryo and seed characteristics during development. Methods Eight maternal plants (two each from four families) and four pollen donors were crossed and developing gynoecia were collected at 10 d post-pollination. Half of the maternal plants experienced water stress. Characteristics of embryos and seeds were summarized and also compared with earlier and later developmental stages. Key Results In addition to the expected effects of the maternal plants, all embryo characters differed among pollen donors. Paternal effects varied over time, suggesting that there are windows of opportunity for pollen donors to influence embryo development. Water-stress treatment altered embryo characteristics; embryos were smaller and less developed. In addition, correlations of embryo characteristics with earlier and later stages changed dramatically with water stress. Conclusions The expected maternal effects on embryo development were observed, but there was also evidence for an early paternal role. The relative effects of these

  3. Dynamics of maternal and paternal effects on embryo and seed development in wild radish (Raphanus sativus).

    Science.gov (United States)

    Diggle, P K; Abrahamson, N J; Baker, R L; Barnes, M G; Koontz, T L; Lay, C R; Medeiros, J S; Murgel, J L; Shaner, M G M; Simpson, H L; Wu, C C; Marshall, D L

    2010-08-01

    Variability in embryo development can influence the rate of seed maturation and seed size, which may have an impact on offspring fitness. While it is expected that embryo development will be under maternal control, more controversial hypotheses suggest that the pollen donor and the embryo itself may influence development. These latter possibilities are, however, poorly studied. Characteristics of 10-d-old embryos and seeds of wild radish (Raphanus sativus) were examined to address: (a) the effects of maternal plant and pollen donor on development; (b) the effects of earlier reproductive events (pollen tube growth and fertilization) on embryos and seeds, and the influence of embryo size on mature seed mass; (c) the effect of water stress on embryos and seeds; (d) the effect of stress on correlations of embryo and seed characteristics with earlier and later reproductive events and stages; and (e) changes in maternal and paternal effects on embryo and seed characteristics during development. Eight maternal plants (two each from four families) and four pollen donors were crossed and developing gynoecia were collected at 10 d post-pollination. Half of the maternal plants experienced water stress. Characteristics of embryos and seeds were summarized and also compared with earlier and later developmental stages. In addition to the expected effects of the maternal plants, all embryo characters differed among pollen donors. Paternal effects varied over time, suggesting that there are windows of opportunity for pollen donors to influence embryo development. Water-stress treatment altered embryo characteristics; embryos were smaller and less developed. In addition, correlations of embryo characteristics with earlier and later stages changed dramatically with water stress. The expected maternal effects on embryo development were observed, but there was also evidence for an early paternal role. The relative effects of these controls may change over time. Thus, there may be

  4. Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Pang, Daxin, E-mail: pdx@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Ouyang, Hongsheng, E-mail: ouyh@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China)

    2011-07-29

    Highlights: {yields} Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. {yields} The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. {yields} A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 {mu}g/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

  5. Embryonic catalase protects against ethanol embryopathies in acatalasemic mice and transgenic human catalase-expressing mice in embryo culture.

    Science.gov (United States)

    Miller-Pinsler, Lutfiya; Wells, Peter G

    2015-09-15

    Reactive oxygen species (ROS) have been implicated in the mechanism of ethanol (EtOH) teratogenicity, but the protective role of the embryonic antioxidative enzyme catalase is unclear, as embryonic activity is only about 5% of maternal levels. We addressed this question in a whole embryo culture model. C57BL/6 mouse embryos expressing human catalase (hCat) or their wild-type (C57BL/6 WT) controls, and C3Ga.Cg-Cat(b)/J catalase-deficient, acatalasemic (aCat) mouse embryos or their wild-type C3HeB/FeJ (C3H WT) controls, were explanted on gestational day (GD) 9 (plug=GD 1), exposed for 24h to 2 or 4mg/mL EtOH or vehicle, and evaluated for functional and morphological changes. hCat and C57BL/6 WT vehicle-exposed embryos developed normally, while EtOH was embryopathic in C57BL/6 WT embryos, evidenced by decreases in anterior neuropore closure, somites developed, turning and head length, whereas hCat embryos were protected (pcatalase (PEG-cat) 8h prior to embryo culture, which increases embryonic catalase activity, blocked all EtOH embryopathies (pcatalase is a determinant of risk for EtOH embryopathies. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Nanomaterial Toxicity Screening in Developing Zebrafish Embryos

    Science.gov (United States)

    To assess nanomaterial vertebrate toxicity, a high-content screening assay was created using developing zebrafish, Danio rerio. This included a diverse group of nanomaterials (n=42 total) ranging from metallic (Ag, Au) and metal oxide (CeO2, CuO, TiO2, ZnO) nanoparticles, to non...

  7. Changes in RNA Splicing in Developing Soybean (Glycine max Embryos

    Directory of Open Access Journals (Sweden)

    Delasa Aghamirzaie

    2013-11-01

    Full Text Available Developing soybean seeds accumulate oils, proteins, and carbohydrates that are used as oxidizable substrates providing metabolic precursors and energy during seed germination. The accumulation of these storage compounds in developing seeds is highly regulated at multiple levels, including at transcriptional and post-transcriptional regulation. RNA sequencing was used to provide comprehensive information about transcriptional and post-transcriptional events that take place in developing soybean embryos. Bioinformatics analyses lead to the identification of different classes of alternatively spliced isoforms and corresponding changes in their levels on a global scale during soybean embryo development. Alternative splicing was associated with transcripts involved in various metabolic and developmental processes, including central carbon and nitrogen metabolism, induction of maturation and dormancy, and splicing itself. Detailed examination of selected RNA isoforms revealed alterations in individual domains that could result in changes in subcellular localization of the resulting proteins, protein-protein and enzyme-substrate interactions, and regulation of protein activities. Different isoforms may play an important role in regulating developmental and metabolic processes occurring at different stages in developing oilseed embryos.

  8. The politics of human embryo research and the motivation to achieve PGD.

    Science.gov (United States)

    Theodosiou, Anastasia A; Johnson, Martin H

    2011-05-01

    This article reports a historical study of factors influencing the achievement of clinical preimplantation genetic diagnosis (PGD) in 1990, 22 years after its first demonstration in animals. During the 1970s, research on PGD continued in large farm animals, but serious interest in human PGD was not evident until 1986. First, interest in PGD during the 1970s waned with the advent of prenatal testing, which for gynaecologists was clinically more familiar, technically simpler and ethically less challenging than IVF. Indeed, IVF was viewed with widespread suspicion until the first IVF births in 1978. Second, interest in clinical PGD was stimulated by the UK Parliamentary reaction against human embryo research that greeted the Warnock Report in 1984. This hostility led scientists to initiate a pro-research campaign, further galvanized in 1985 by MP Enoch Powell's bid to ban such research. However, while Powell abhorred embryo research, he approved of PGD, a stance that divided the anti-research lobby. Accordingly, the campaigners for research emphasized that it was needed to achieve PGD. Powell demanded evidence of such projects and PGD research increased from 1986. It is concluded that UK political debates on embryo research played a critical role in stimulating the achievement of clinical PGD. Human pregnancies following preimplantation genetic diagnosis (PGD) for embryo sex were announced in 1990, 22 years after the technique was pioneered in animals. PGD in humans required not only technological advances, such as IVF and sensitive diagnostic tests, but also the motivation to develop and apply them. Our historical analysis shows that, although research on PGD continued in large farm animals during the 1970s, and techniques of the required sensitivity were developed on mouse embryo models, interest in clinical PGD was not evident until 1986. Two factors stimulated this sudden change in motivation. First, interest in PGD was depressed during the 1970s by the advent of

  9. Human embryos secrete microRNAs into culture media--a potential biomarker for implantation.

    Science.gov (United States)

    Rosenbluth, Evan M; Shelton, Dawne N; Wells, Lindsay M; Sparks, Amy E T; Van Voorhis, Bradley J

    2014-05-01

    To determine whether human blastocysts secrete microRNA (miRNAs) into culture media and whether these reflect embryonic ploidy status and can predict in vitro fertilization (IVF) outcomes. Experimental study of human embryos and IVF culture media. Academic IVF program. 91 donated, cryopreserved embryos that developed into 28 tested blastocysts, from 13 couples who had previously completed IVF cycles. None. Relative miRNA expression in IVF culture media. Blastocysts were assessed by chromosomal comparative genomic hybridization analysis, and the culture media from 55 single-embryo transfer cycles was tested for miRNA expression using an array-based quantitative real-time polymerase chain reaction analysis. The expression of the identified miRNA was correlated with pregnancy outcomes. Ten miRNA were identified in the culture media; two were specific to spent media (miR-191 and miR-372), and one was only present in media before the embryos had been cultured (miR-645). MicroRNA-191 was more highly concentrated in media from aneuploid embryos, and miR-191, miR-372, and miR-645 were more highly concentrated in media from failed IVF/non-intracytoplasmic sperm injection cycles. Additionally, miRNA were found to be more highly concentrated in ICSI and day-5 media samples when compared with regularly inseminated and day-4 samples, respectively. MicroRNA can be detected in IVF culture media. Some of these miRNA are differentially expressed according to the fertilization method, chromosomal status, and pregnancy outcome, which makes them potential biomarkers for predicting IVF success. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  10. Human Development

    NARCIS (Netherlands)

    D.R. Gasper (Des)

    2009-01-01

    textabstract‘Human development’ language spread gradually in circles of national and international development policy and planning from the 1970s and acquired a definitive form in the 1990s in the United Nations Development Programme’s Human Development Reports (HDRs). Human development was defined

  11. Myoinositol Improves Embryo Development in PCOS Patients Undergoing ICSI

    Directory of Open Access Journals (Sweden)

    Artur Wdowiak

    2016-01-01

    Full Text Available The aim of this study was to investigate the activity of myoinositol, in a court of 217 PCOS women undergoing intracytoplasmic sperm injection (ICSI, on pregnancy rate, embryo development, estradiol, and progesterone concentration in blood serum, superoxide dismutase (SOD, and catalase (CAT in follicular fluid. Concerning the court of patient, 112 (groups I and II out of 217 were PCOS women, whereas group III consisted of healthy subjects (not PCOS. Group I patients were treated with 400 μg of folic acid per day for 3 months before ICSI, whereas group II patients received 4000 mg of myoinositol and 400 μg of folic acid per day for 3 months before ICSI. Group II revealed a shorter embryo/blastocyst development period between microinjection and 5-cell stage compared to group I. The difference in SOD concentration between groups I and II and between groups II and III was statistically significant. In group II, 34.62% of pregnancies were obtained, whereas in group I this number reached 20% (NS. Myoinositol increased embryo development dynamics and accelerated blastocyst stage reaching time; however, no effect was shown on clinical pregnancy. Furthermore, it restored SOD concentration, lowered in PCOS women, but did not exert any effect on CAT concentration.

  12. Human embryos from induced pluripotent stem cell-derived gametes: ethical and quality considerations.

    Science.gov (United States)

    Ilic, Dusko; Ogilvie, Caroline; Noli, Laila; Kolundzic, Nikola; Khalaf, Yacoub

    2017-09-01

    Protocols for successful differentiation of male and female gametes from induced pluripotent stem cells have been published. Although culture of precursor cells in a natural microenvironment remains necessary to achieve terminal differentiation, the creation of human preimplantation embryos from induced pluripotent stem cell-derived gametes is technically feasible. Such embryos could provide a solution to the scarcity of human cleavage-stage embryos donated for research. Here, we discuss current technology, major research-related ethical concerns and propose the norms that would assure the quality and reliability of such embryos.

  13. Polyamine levels during the development of zygotic and somatic embryos of Pinus radiata

    Science.gov (United States)

    Rakesh Minocha; Dale R. Smith; Cathie Reeves; Kevin D. Steele; Subhash C. Minocha

    1999-01-01

    Changes in the cellular content of three polyamines (putrescine, spermidine and spermine) were compared at different stages of development in zygotic and somatic embryos of Pinus radiata D. Don. During embryo development, both the zygotic and the somatic embryos showed a steady increase in spermidine content, with either a small decrease or no...

  14. Cryopreservation of murine embryos, human spermatozoa and embryonic stem cells using a liquid nitrogen-free, controlled rate freezer.

    Science.gov (United States)

    Morris, G J; Acton, E; Faszer, K; Franklin, A; Yin, H; Bodine, R; Pareja, J; Zaninovic, N; Gosden, R

    2006-09-01

    A Stirling Cycle Cryocooler has been developed as an alternative to conventional liquid nitrogen controlled rate freezers. Unlike liquid nitrogen systems, the Stirling Cycle freezer does not pose a contamination risk, can be used in sterile conditions and has no need for a constant supply of cryogen. Three types of samples from two species (murine embryos, human spermatozoa and embryonic stem cells), each requiring different cooling protocols, were cryopreserved in the Stirling Cycle freezer. For comparison, cells were also frozen in a conventional liquid nitrogen controlled rate freezer. Upon thawing, the rates of survival of viable cells were generally greater than 50% for mouse embryos and human embryonic stem cells, based on morphology (mouse embryos) and staining and colony formation (human embryonic stem cells). Survival rates of human spermatozoa frozen in the Stirling Cycle freezer, based on motility and dead cell staining, were similar to those of samples frozen in a conventional controlled rate freezer using liquid nitrogen.

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

    Science.gov (United States)

    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.

  16. Oral Morphine Consumption Reduces Lens Development in Rat Embryos

    Directory of Open Access Journals (Sweden)

    Hossein Bahadoran

    2012-07-01

    Full Text Available Objective: Consumption of morphine, during pregnancy, in addition to inducing defects in the mother’s nervous system function, caused defects or delays in the formation and evolution of embryonic visual system. In the present study, changes in lens development was assessed in embryos exposed in utero to morphine. Material and Methods: Female Wistar rats (250-300 g were mated with male rats and pregnancy was determined by sperm observation in vaginal smear. This day was considered as embryonic day zero (E0. The females were then divided randomly into the experimental and the control groups. The control group received tap water and the experimental group received morphine (0.05 mg/ml in their water. On embryonic day 13 ( E13, blood samples were collected from the retro-orbital sinus of all animals for plasma corticosterone detection. On embryonic day 17(E17, the animals were killed by an overdose of chloroform and the embryos were taken out surgically. The embryos were fixed in 10% formalin for 30 days. At this time, the head of the embryos were removed for tissue processing and Hematoxylin- Eosin (H&E staining. The samples were evaluated using light microscope and MOTIC software. Results: Our data indicated that plasma corticosterone level was dramatically increased and the lens was thinner in the experimental group. (Although the proliferation of lens cells increased in the experiment group but that lens had delay in removing the proliferated and elongation cells with abnormal density in the lateral part of the lens in compare with control group. I have no idea what the authors are stating here. Moreover, the opening of the eyelids was delayed in the off springs of the mothers who received morphine. Conclusions: This study showed that morphine consumption during pregnancy leads to defects in fetal visual system development, particularly in the lens, and eyelids.

  17. Jewish points of views on the animation of the human embryo

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    Ks. Artur Aleksiejuk

    2014-11-01

    Full Text Available According to biblical anthropology, human beings are composed of body and soul. The question arises, however, at what moment does the body of the embryo possess a spiritual element? Can the breath of God visit the created and already developing in its own biological rhythm embryo? The key issue here is the moment of animation – the origin of a living being, which is created in the image and likeness of God. This article presents various Jewish points of views on the animation of the human embryo, all of which attempt to determine the exact moment at which the soul is breathed into the human body. Rabbinical authorities distinguish five different moments in this process: conception, the forty first day after conception, the birth of the child, the moment of circumcision and the moment in which the child is able to say “Amen.” The first three mentioned cases have the most supporters. The first refers to the simultaneous animation, while the other theories argue for successive animation.

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

    Energy Technology Data Exchange (ETDEWEB)

    Osychenko, A A; Zalesskii, A D; Krivokharchenko, A S; Zhakhbazyan, A K; Nadtochenko, V A [N N Semenov Institute of Chemical Physics, Russian Academy of Sciences, Moscow (Russian Federation); Ryabova, A V [A M Prokhorov General Physics Institute, Russian Academy of Sciences, Moscow (Russian Federation)

    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)

  19. Exploring polyamines: Functions in embryo/fetal development

    Directory of Open Access Journals (Sweden)

    Tarique Hussain

    2017-03-01

    Full Text Available Polyamines such as putrescine, spermidine, spermine and agmatine are aliphatic polycationic compounds present in all living cells, and are derived from amino acids, intestinal bacteria, exfoliated enterocytes and supported from diet. Polyamines as the key compounds play essential role in cell proliferation, growth and differentiation. They also exert significant effects on embryonic development, implantation, embryonic diapause, placentation, angiogensis and fetal development. This review paper summarizes the functions of polyamines and embryo/fetus development and its regulatory mechanism which should help to provide some evidences for clinic.

  20. Specific gene-regulation networks during the pre-implantation development of the pig embryo as revealed by deep sequencing.

    Science.gov (United States)

    Cao, Suying; Han, Jianyong; Wu, Jun; Li, Qiuyan; Liu, Shichao; Zhang, Wei; Pei, Yangli; Ruan, Xiaoan; Liu, Zhonghua; Wang, Xumin; Lim, Bing; Li, Ning

    2014-01-03

    Because few studies exist to describe the unique molecular network regulation behind pig pre-implantation embryonic development (PED), genetic engineering in the pig embryo is limited. Also, this lack of research has hindered derivation and application of porcine embryonic stem cells and porcine induced pluripotent stem cells (iPSCs). We identified and analyzed the genome wide transcriptomes of pig in vivo-derived and somatic cell nuclear transferred (SCNT) as well as mouse in vivo-derived pre-implantation embryos at different stages using mRNA deep sequencing. Comparison of the pig embryonic transcriptomes with those of mouse and human pre-implantation embryos revealed unique gene expression patterns during pig PED. Pig zygotic genome activation was confirmed to occur at the 4-cell stage via genome-wide gene expression analysis. This activation was delayed to the 8-cell stage in SCNT embryos. Specific gene expression analysis of the putative inner cell mass (ICM) and the trophectoderm (TE) revealed that pig and mouse pre-implantation embryos share regulatory networks during the first lineage segregation and primitive endoderm differentiation, but not during ectoderm commitment. Also, fatty acid metabolism appears to be a unique characteristic of pig pre-implantation embryonic development. In addition, the global gene expression patterns in the pig SCNT embryos were different from those in in vivo-derived pig embryos. Our results provide a resource for pluripotent stem cell engineering and for understanding pig development.

  1. Dilemmas encountered with preimplantation diagnosis of aneuploidy in human embryos.

    Science.gov (United States)

    Allan, John; Edirisinghe, Rohini; Anderson, Jasen; Jemmott, Rodney; Nandini, A V; Gattas, Michael

    2004-04-01

    An increased embryo aneuploidy rate is associated with advancing maternal age. Preimplantation genetic diagnosis (PGD) using fluorescence in situ hybridisation (FISH) coupled with in vitro fertilisation (IVF)/embryo biopsy provides a powerful tool to improve the take home baby rates for this poor prognostic group. To report the preliminary findings of a PGD study for aneuploidy screening and to discuss the dilemmas encountered. Preimplantation genetic diagnosis was offered in egg pick up-PGD and frozen embryo transfer-PGD cycles. Embryo biopsy was carried out on day 3 and FISH was used to detect chromosomal abnormalities. The outcome of 75 patients, 100 treatment cycles; 62 egg pick up-PGD and 38 frozen embryo transfer-PGD are presented. The embryo biopsy rate, blastomere survival, presence of nuclei and successful FISH rates for egg pick-up and frozen embryo transfer cycles were similar giving a chromosomal abnormality rate of 57.5 and 51.2% for the respective treatment group. The positive pregnancy, clinical pregnancy and implantation rates were, for egg pick up-PGD 22.7, 13.6 and 21.1% and for frozen embryo transfer-PGD 13.8, 10.3 and 10.0%, respectively. Preimplantation genetic diagnosis coupled with IVF treatment seems to give satisfactory pregnancy rates. The high embryo aneuploidy rates, chromosomal mosaicism and other issues have presented significant ethical and management dilemmas for our physicians and patients alike. These issues highlight the importance of skillful pretreatment counselling for patients considering PGD.

  2. Tongue Growth during Prenatal Development in Korean Fetuses and Embryos

    Directory of Open Access Journals (Sweden)

    Soo Jeong Hong

    2015-11-01

    Full Text Available Background: Prenatal tongue development may affect oral-craniofacial structures, but this muscular organ has rarely been investigated. Methods: In order to document the physiology of prenatal tongue growth, we histologically examined the facial and cranial base structures of 56 embryos and 106 fetuses. Results: In Streeter’s stages 13–14 (fertilization age [FA], 28 to 32 days, the tongue protruded into the stomodeal cavity from the retrohyoid space to the cartilaginous mesenchyme of the primitive cranial base, and in Streeter’s stage 15 (FA, 33 to 36 days, the tongue rapidly swelled and compressed the cranial base to initiate spheno-occipital synchondrosis and continued to swell laterally to occupy most of the stomodeal cavity in Streeter’s stage 16–17 (FA, 37 to 43 days. In Streeter’s stage 18–20 (FA, 44 to 51 days, the tongue was vertically positioned and filled the posterior nasopharyngeal space. As the growth of the mandible and maxilla advanced, the tongue was pulled down and protruded anteriorly to form the linguomandibular complex. Angulation between the anterior cranial base (ACB and the posterior cranial base (PCB was formed by the emerging tongue at FA 4 weeks and became constant at approximately 124°–126° from FA 6 weeks until birth, which was consistent with angulations measured on adult cephalograms. Conclusions: The early clockwise growth of the ACB to the maxillary plane became harmonious with the counter-clockwise growth of the PCB to the tongue axis during the early prenatal period. These observations suggest that human embryonic tongue growth affects ACB and PCB angulation, stimulates maxillary growth, and induces mandibular movement to achieve the essential functions of oral and maxillofacial structures.

  3. A microfluidic system supports single mouse embryo culture leading to full-term development

    NARCIS (Netherlands)

    Esteves, Telma Cristina; van Rossem, F.; Nordhoff, Verena; Schlatt, Stefan; Boiani, Michele; le Gac, Severine

    2013-01-01

    The present study demonstrates the feasibility of application of a microfluidic system for in vitro culture of pre-implantation mouse embryos, with subsequent development to full-term upon embryo transfer. Specifically, embryos cultured in groups in nL volume chambers achieve pre-implantation

  4. DOT1L inhibitor improves early development of porcine somatic cell nuclear transfer embryos

    DEFF Research Database (Denmark)

    Tao, Jia; Zhang, Yu; Zuo, Xiaoyuan

    2017-01-01

    for 12 or 24 h significantly enhanced the blastocyst rate of SCNT embryos and dramatically reduced the level of H3K79me2 during SCNT 1-cell embryonic development. Additionally, H3K79me2 level in the EPZ-treated SCNT embryos was similar to that in in vitro fertilized embryos, suggesting that DOT1L...

  5. Epigenetics and chromosome segregation in human pre-implantation embryos

    NARCIS (Netherlands)

    C. van de Werken (Christine)

    2015-01-01

    markdownabstractAbstract Chapter 1 Currently, the average pregnancy rate per embryo transfer after in vitro fertilization (IVF) is around 32%. In order to achieve better results in the future, we need to gain knowledge on all aspects of the treatment, including pre-implantation embryo

  6. Cryopreservation of human embryos and its contribution to in vitro fertilization success rates

    NARCIS (Netherlands)

    Wong, Kai Mee; Mastenbroek, Sebastiaan; Repping, Sjoerd

    2014-01-01

    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

  7. The effect of unilateral ovariectomy on early embryonic survival and embryo development in rabbits

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    R. Peiró

    2014-06-01

    Full Text Available Unilateral ovariectomy can be used to study uterine capacity in rabbits because an overcrowding of the functional uterine horn is produced. Due to the uterus duplex, the rabbit is the ideal model for such studies. However, this technique may affect embryo survival. The aim of this work is to study the effect of unilateral ovariectomy on early embryo survival and development in rabbit. A total of 101 unilateral ovariectomised females and 52 intact females were compared after slaughter at 30 h post-mating. Early embryo survival was estimated as the ratio between number of embryo recovered and ovulation rate. No differences were found between intact and unilaterally ovariectomised females in this trait. Unilateral ovariectomy did not change embryo development, measured as the number of embryo cells. Variability of embryo development was not affected either. At 30 h post-mating, the majority of embryos (86.2% were 4-cell stage. Embryo quality was evaluated according to morphological criteria. No difference in embryo quality between intact and unilaterally ovariectomised females was found. Therefore, unilateral ovariectomy performed before puberty in rabbit does not modify early embryo survival and development.

  8. Culture media for human pre-implantation embryos in assisted reproductive technology cycles.

    Science.gov (United States)

    Youssef, Mohamed M A; Mantikou, Eleni; van Wely, Madelon; Van der Veen, Fulco; Al-Inany, Hesham G; Repping, Sjoerd; Mastenbroek, Sebastiaan

    2015-11-20

    Many media are commercially available for culturing pre-implantation human embryos in assisted reproductive technology (ART) cycles. It is unknown which culture medium leads to the best success rates after ART. To evaluate the safety and effectiveness of different human pre-implantation embryo culture media in used for in vitro fertilisation (IVF) and intracytoplasmic sperm injection (ICSI) cycles. We searched the Cochrane Menstrual Disorders and Subfertility Group's Trials Register, Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE, the National Research Register, the Medical Research Council's Clinical Trials Register and the NHS Center for Reviews and Dissemination databases from January 1985 to March 2015. We also examined the reference lists of all known primary studies, review articles, citation lists of relevant publications and abstracts of major scientific meetings. We included all randomised controlled trials which randomised women, oocytes or embryos and compared any two commercially available culture media for human pre-implantation embryos in an IVF or ICSI programme. Two review authors independently selected the studies, assessed their risk of bias and extracted data. We sought additional information from the authors if necessary. We assessed the quality of the evidence using Grades of Recommendation, Assessment, Development and Evaluation (GRADE) methods. The primary review outcome was live birth or ongoing pregnancy. We included 32 studies in this review. Seventeen studies randomised women (total 3666), three randomised cycles (total 1018) and twelve randomised oocytes (over 15,230). It was not possible to pool any of the data because each study compared different culture media.Only seven studies reported live birth or ongoing pregnancy. Four of these studies found no evidence of a difference between the media compared, for either day three or day five embryo transfer. The data from the fifth study did not appear reliable

  9. Implantation of the human embryo requires Rac1-dependent endometrial stromal cell migration.

    Science.gov (United States)

    Grewal, Seema; Carver, Janet G; Ridley, Anne J; Mardon, Helen J

    2008-10-21

    Failure of the human embryo to implant into the uterine wall during the early stages of pregnancy is a major cause of infertility. Implantation involves embryo apposition and adhesion to the endometrial epithelium followed by penetration through the epithelium and invasion of the embryonic trophoblast through the endometrial stroma. Although gene-knockdown studies have highlighted several molecules that are important for implantation in the mouse, the molecular mechanisms controlling implantation in the human are unknown. Here, we demonstrate in an in vitro model for human implantation that the Rho GTPases Rac1 and RhoA in human endometrial stromal cells modulate invasion of the human embryo through the endometrial stroma. We show that knockdown of Rac1 expression in human endometrial stromal cells inhibits human embryonic trophoblast invasion into stromal cell monolayers, whereas inhibition of RhoA activity promotes embryo invasion. Furthermore, we demonstrate that Rac1 is required for human endometrial stromal cell migration and that the motility of the stromal cells increases at implantation sites. This increased motility correlates with a localized increase in Rac1 activation and a reciprocal decrease in RacGAP1 levels. These results reveal embryo-induced and localized endometrial responses that may govern implantation of the human embryo.

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

    Science.gov (United States)

    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.

  11. Early embryonic development and in vitro culture of in vivo produced embryos in the farmed European polecat (Mustela putorius).

    Science.gov (United States)

    Lindeberg, H; Järvinen, M

    2003-09-15

    Early embryonic development and in vitro culture of in vivo produced embryos in the farmed European polecat (Mustela putorius) was investigated as a part of an ex situ conservation program of the endangered European mink (Mustela lutreola), using the European polecat as a model species. The oestrus cycles of 34 yearling polecat females were monitored by visual examination of the vulval swelling and, to induce ovulation, the females were mated once daily on two consecutive days. Sixteen yearling males were used for mating. The females were humanely killed 3-14 days after the first mating and the uteri and oviducts were collected for embryo recovery. Uterine and oviductal flushings yielded a total number of 295 embryos, representing developmental stages from the 1-cell stage to large expanded and hatched blastocysts. On Day 3 after the first mating, only 1-16-cell stage embryos were recovered. Between Days 4 and 6 after the first mating, 1-16-cell stage embryos and morulae were found. The first blastocysts were recovered on Day 7 after the first mating. The first implanted blastocysts were detected on Day 11 after the first mating. A total number of 85 embryos were in vitro cultured after recovery. Blastocyst production rates for in vitro cultured 1-16-cell stage embryos and for morulae/compact morulae were 68 and 84%, respectively. For all cultured embryos, the hatching rate was 15%. The in vitro culture requirements for the preimplantation embryos of the farmed European polecat remain to be determined before further utilization of the technique.

  12. Gas6 stimulates angiogenesis of human retinal endothelial cells and of zebrafish embryos via ERK1/2 signaling.

    Directory of Open Access Journals (Sweden)

    Young Sook Kim

    Full Text Available AIM: To determine if growth arrest-specific 6 (Gas6 plays an important role in the regulation of angiogenesis in human retinal microvascular endothelial cells (HRMECs and in vessel development of zebrafish. METHODS: Proliferation, wound-healing cell migration, and tube formation were measured in HRMECs treated with recombinant human Gas6 (rhGas6. Sprague-Dawley rat aortas in Matrigels were treated with rhGas6, and microvessel sprouting emanating from arterial rings was analyzed. Transgenic zebrafish embryos (flk:GFP were microinjected with rhGas6 at 50 hours post-fertilization (hpf, and ectopic sprouting of subintestinal vessels (SIVs was observed under a confocal microscope. Morpholino oligonucleotides (MOs were microinjected to knockdown gas6 in zebrafish embryos, and intersegmental vessel impairment was observed. The effect of the extracellular signal-regulated kinase (ERK1/2 inhibitor on the migration of HRMECs and on vessel development in zebrafish embryos was tested. RESULTS: rhGas6 stimulated proliferation, migration, and tube formation in HRMECs in a dose-dependent manner. In rat aortas, rhGas6 induced vessel outgrowth, and the sprouting length was longer than that of controls. The rhGas6-microinjected zebrafish embryos had significantly increased vessel outgrowth in the SIVs. Recombinant human vascular endothelial growth factor (rhVEGF served as a positive control. Knockdown of gas6 inhibited angiogenesis in the developing vessels of zebrafish. The ERK1/2 inhibitor inhibited HRMEC migration and intersegmental vessel formation in zebrafish embryos. CONCLUSIONS/INTERPRETATIONS: These data suggest that Gas6 plays a pivotal role in proliferation, migration, and sprouting of angiogenic endothelial cells in the retina and in zebrafish embryos. Furthermore, Gas6 induced angiogenic processes are induced via phosphorylation of ERK1/2.

  13. Effect of different ovule isolation times on the embryo development of Campanula hybrids

    DEFF Research Database (Denmark)

    Röper, Anna Catharina; Lütken, Henrik Vlk; Hegelund, Josefine Nymark

    2012-01-01

    , hybridization between plant species is associated with many challenges to enable survival of the developing embryo. Here we present an optimised technique for embryo rescue via ovule isolation in selected intra- and interspecific Campanula hybrids. A frequent problem in embryo rescue is the malformation...... of the endosperm. To circumvent this, embryos were isolated and the optimal ovule isolation time and growth conditions were determined to increase embryo survival. Ovules were isolated one to four weeks after pollination and cultivated on a modified MS medium. When ovules were allowed to stay inside the ovary...... for 2-3 weeks the number of germinating embryos increased as compared to ovules isolated one week after pollination. Additionally, ovules isolated 2-3 weeks after pollination showed an increased embryo germination rate. Among the Campanula hybrids, produced here from both the intraspecific crosses...

  14. Metabolic and Transcriptional Reprogramming in Developing Soybean (Glycine max Embryos

    Directory of Open Access Journals (Sweden)

    Ruth Grene

    2013-05-01

    Full Text Available Soybean (Glycine max seeds are an important source of seed storage compounds, including protein, oil, and sugar used for food, feed, chemical, and biofuel production. We assessed detailed temporal transcriptional and metabolic changes in developing soybean embryos to gain a systems biology view of developmental and metabolic changes and to identify potential targets for metabolic engineering. Two major developmental and metabolic transitions were captured enabling identification of potential metabolic engineering targets specific to seed filling and to desiccation. The first transition involved a switch between different types of metabolism in dividing and elongating cells. The second transition involved the onset of maturation and desiccation tolerance during seed filling and a switch from photoheterotrophic to heterotrophic metabolism. Clustering analyses of metabolite and transcript data revealed clusters of functionally related metabolites and transcripts active in these different developmental and metabolic programs. The gene clusters provide a resource to generate predictions about the associations and interactions of unknown regulators with their targets based on “guilt-by-association” relationships. The inferred regulators also represent potential targets for future metabolic engineering of relevant pathways and steps in central carbon and nitrogen metabolism in soybean embryos and drought and desiccation tolerance in plants.

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

    Science.gov (United States)

    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. Copyright © 2013 Elsevier B.V. All rights reserved.

  16. Embryonic catalase protects against ethanol embryopathies in acatalasemic mice and transgenic human catalase-expressing mice in embryo culture

    Energy Technology Data Exchange (ETDEWEB)

    Miller-Pinsler, Lutfiya [Department of Pharmacology and Toxicology, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Wells, Peter G., E-mail: pg.wells@utoronto.ca [Division of Biomolecular Sciences, Faculty of Pharmacy, University of Toronto, Toronto, Ontario (Canada); Department of Pharmacology and Toxicology, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada)

    2015-09-15

    Reactive oxygen species (ROS) have been implicated in the mechanism of ethanol (EtOH) teratogenicity, but the protective role of the embryonic antioxidative enzyme catalase is unclear, as embryonic activity is only about 5% of maternal levels. We addressed this question in a whole embryo culture model. C57BL/6 mouse embryos expressing human catalase (hCat) or their wild-type (C57BL/6 WT) controls, and C3Ga.Cg-Cat{sup b}/J catalase-deficient, acatalasemic (aCat) mouse embryos or their wild-type C3HeB/FeJ (C3H WT) controls, were explanted on gestational day (GD) 9 (plug = GD 1), exposed for 24 h to 2 or 4 mg/mL EtOH or vehicle, and evaluated for functional and morphological changes. hCat and C57BL/6 WT vehicle-exposed embryos developed normally, while EtOH was embryopathic in C57BL/6 WT embryos, evidenced by decreases in anterior neuropore closure, somites developed, turning and head length, whereas hCat embryos were protected (p < 0.001). Maternal pretreatment of C57BL/6 WT dams with 50 kU/kg PEG-catalase (PEG-cat) 8 h prior to embryo culture, which increases embryonic catalase activity, blocked all EtOH embryopathies (p < 0.001). Vehicle-exposed aCat mouse embryos had lower yolk sac diameters compared to WT controls, suggesting that endogenous ROS are embryopathic. EtOH was more embryopathic in aCat embryos than WT controls, evidenced by reduced head length and somite development (p < 0.01), and trends for reduced anterior neuropore closure, turning and crown–rump length. Maternal pretreatment of aCat dams with PEG-Cat blocked all EtOH embryopathies (p < 0.05). These data suggest that embryonic catalase is a determinant of risk for EtOH embryopathies. - Highlights: • Ethanol (EtOH) exposure causes structural embryopathies in embryo culture. • Genetically enhanced catalase (hCat) protects against EtOH embryopathies. • Genetically deficient catalase (aCat) exacerbates EtOH embryopathies. • Embryonic catalase is developmentally important. • Et

  17. Neurotransmitter signaling pathways required for normal development in Xenopus laevis embryos: a pharmacological survey screen.

    Science.gov (United States)

    Sullivan, Kelly G; Levin, Michael

    2016-10-01

    Neurotransmitters are not only involved in brain function but are also important signaling molecules for many diverse cell types. Neurotransmitters are widely conserved, from evolutionarily ancient organisms lacking nervous systems through man. Here, results are reported from a loss- and gain-of-function survey, using pharmacological modulators of several neurotransmitter pathways to examine possible roles for these pathways in normal embryogenesis. Applying reagents targeting the glutamatergic, adrenergic and dopaminergic pathways to embryos of Xenopus laevis from gastrulation to organogenesis stages, we observed and quantified numerous malformations, including craniofacial defects, hyperpigmentation, muscle mispatterning and miscoiling of the gut. These data implicate several key neurotransmitters in new embryonic patterning roles, reveal novel earlier stages for processes involved in eye development, suggest new targets for subsequent molecular-genetic investigation, and highlight the necessity for in-depth toxicology studies of psychoactive compounds to which human embryos might be exposed during pregnancy. © 2016 Anatomical Society.

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

    Science.gov (United States)

    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 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. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. In vivo and in vitro development of Tibetan antelope (Pantholops hodgsonii interspecific cloned embryos

    Directory of Open Access Journals (Sweden)

    Guanghua SU,Lei CHENG,Yu GAO,Kun LIU,Zhuying WEI,Chunling BAI,Fengxia YIN,Li GAO,Guangpeng LI,Shorgan BOU

    2014-02-01

    Full Text Available The Tibetan antelope is endemic to the Tibetan Plateau, China, and is now considered an endangered species. As a possible rescue strategy, the development of embryos constructed by interspecies somatic cell nuclear transfer (iSCNT was examined. Tibetan antelope fibroblast cells were transferred into enucleated bovine, ovine and caprine oocytes. These cloned embryos were then cultured in vitro or in the oviducts of intermediate animals. Less than 0.5% of the reconstructed antelope-bovine embryos cultured in vitro developed to the blastocyst stage. However, when the cloned antelope-bovine embryos were transferred to caprine oviducts, about 1.6% of the embryos developed to the blastocyst stage. In contrast, only 0.7% of the antelope-ovine embryos developed to the morula stage and none developed to blastocysts in ovine oviducts. The treatment of donor cells and bovine oocytes with trichostatin A did not improve the embryo development even when cultured in the oviducts of ovine and caprine. When the antelope-bovine embryos, constructed from oocytes treated with roscovitine or trichostatin A, were cultured in rabbit oviducts 2.3% and 14.3% developed to blastocysts, respectively. It is concluded that although some success was achieved with the protocols used, interspecies cloning of Tibetan antelope presents difficulties still to be overcome. The mechanisms resulting in the low embryo development need investigation and progress might require a deeper understanding of cellular reprogramming.

  20. Early development of the human pelvic diaphragm

    NARCIS (Netherlands)

    Koch, Wijnandus Franciscus Robertus Maria

    2006-01-01

    The last decade an increasing interest in the pelvic floor can be observed in medical sciences. The lack of data on the development of the human pelvic floor is striking. The early development of the human pelvic diaphragm was studied. Materials and methodsUse was made of 38 human embryos and

  1. Sequential gene activation and gene imprinting during early embryo development in maize.

    Science.gov (United States)

    Meng, Dexuan; Zhao, Jianyu; Zhao, Cheng; Luo, Haishan; Xie, Mujiao; Liu, Renyi; Lai, Jinsheng; Zhang, Xiaolan; Jin, Weiwei

    2017-11-24

    Gene imprinting is a widely observed epigenetic phenomenon in maize endosperm; however, whether it also occurs in maize embryo remains controversial. Here, we used high throughput RNA sequencing on laser capture microdissection (LCM) and manually dissected maize embryos from reciprocal crosses between inbred lines B73 and Mo17 at six time points (3 to 13 days after pollination) to analyze allelic gene expression patterns. Co-expression analysis revealed sequential gene activation during maize embryo development. Gene imprinting was observed in maize embryos and a greater number of imprinted genes were identified at early embryo stages. Sixty-four strongly imprinted genes were identified (at the threshold of 9:1) on 5 DAP to 13 DAP manually dissected embryos (more imprinted genes on 5 DAP). Forty-one strongly imprinted genes were identified from 3 DAP and 5 DAP embryos obtained by LCM (more imprinted genes on 3DAP). Furthermore, of the 56 genes that were completely imprinted (at the threshold of 99:1), 36 were not previously identified as imprinted genes in endosperms and nor in embryos. In situ hybridization demonstrated that most of the imprinted genes were expressed abundantly in maize embryotic tissue. Our results shed lights on early maize embryo development and provide evidence supporting that gene imprinting occurs in maize embryos. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  2. Impact of the Z potential technique on reducing the sperm DNA fragmentation index, fertilization rate and embryo development.

    Science.gov (United States)

    Duarte, Carlos; Núñez, Víctor; Wong, Yat; Vivar, Carlos; Benites, Elder; Rodriguez, Urso; Vergara, Carlos; Ponce, Jorge

    2017-12-01

    In assisted reproduction procedures, we need to develop and enhance new protocols to optimize sperm selection. The aim of this study is to evaluate the ability of the Z potential technique to select sperm with intact DNA in non-normospermic patients and evaluate the impact of this selection on embryonic development. We analyzed a total of 174 human seminal samples with at least one altered parameter. We measured basal, post density gradients, and post density gradients + Z potential DNA fragmentation index. To evaluate the impact of this technique on embryo development, 54 cases were selected. The embryo development parameters evaluated were fertilization rate, cleavage rate, top quality embryos at the third day and blastocysts rate. We found significant differences in the study groups when we compared the sperm fragmentation index by adding the Z potential technique to density gradient selection vs. density gradients alone. Furthermore, there was no significant difference in the embryo development parameters between the low sperm fragmentation index group vs. the moderate and high sperm fragmentation index groups, when selecting sperms with this new technique. The Z potential technique is a very useful tool for sperm selection; it significantly reduces the DNA fragmentation index and improves the parameters of embryo development. This technique could be considered routine for its simplicity and low cost.

  3. Injection of ligand-free gold and silver nanoparticles into murine embryos does not impact pre-implantation development

    Directory of Open Access Journals (Sweden)

    Ulrike Taylor

    2014-05-01

    Full Text Available Intended exposure to gold and silver nanoparticles has increased exponentially over the last decade and will continue to rise due to their use in biomedical applications. In particular, reprotoxicological aspects of these particles still need to be addressed so that the potential impacts of this development on human health can be reliably estimated. Therefore, in this study the toxicity of gold and silver nanoparticles on mammalian preimplantation development was assessed by injecting nanoparticles into one blastomere of murine 2 cell-embryos, while the sister blastomere served as an internal control. After treatment, embryos were cultured and embryo development up to the blastocyst stage was assessed. Development rates did not differ between microinjected and control groups (gold nanoparticles: 67.3%, silver nanoparticles: 61.5%, sham: 66.2%, handling control: 79.4%. Real-time PCR analysis of six developmentally important genes (BAX, BCL2L2, TP53, OCT4, NANOG, DNMT3A did not reveal an influence on gene expression in blastocysts. Contrary to silver nanoparticles, exposure to comparable Ag+-ion concentrations resulted in an immediate arrest of embryo development. In conclusion, the results do not indicate any detrimental effect of colloidal gold or silver nanoparticles on the development of murine embryos.

  4. Antioxidants improve IVF outcome and subsequent embryo development in the mouse.

    Science.gov (United States)

    Truong, T; Gardner, D K

    2017-12-01

    What is the effect of a combination of three antioxidants (Acetyl-L-Carnitine, N-Acetyl-L-Cysteine and α-Lipoic Acid), present in IVF medium during mouse oocyte and sperm collection, on fertilization and subsequent IVF embryo development? A combination of antioxidants resulted in faster developmental times from the 2-cell stage through to expanded blastocyst stage, accompanied by a significant increase in blastocyst cell number and a reduction of intracellular hydrogen peroxide (H2O2) levels. The antioxidant combination Acetyl-L-Carnitine, N-Acetyl-L-Cysteine and α-Lipoic Acid, when present in embryo culture media, has a significant beneficial effect on in vitro fertilized mouse pronucleate oocyte development, especially under oxidative stress. IVF was conducted with combined antioxidants supplemented in IVF medium that was used for mouse oocyte collection and fertilization (oocyte IVF medium, 4 h exposure) and sperm collection and preparation (sperm IVF medium, 1 h exposure). IVF was conducted under 20% oxygen, in the presence or absence of a combination of antioxidants (10 μM Acetyl-L-Carnitine, 10 μM N-Acetyl-L-Cysteine, 5 μM α-Lipoic Acid) and resultant embryos cultured with and without antioxidants under 20% oxygen. Subsequently, the effects of antioxidants on either oocytes or sperm was evaluated. Embryo development was analysed through time-lapse microscopy followed by differential nuclear staining to determine cell allocation in the blastocyst. Intracellular levels of H2O2 were assessed using an aryl boronate probe after 4 h of incubation with antioxidants. Controls were gametes and embryos that had no antioxidants in the medium. In a separate series of experiments, pronucleate oocytes were collected in handling medium with and without antioxidants for 20 min and subsequent cell numbers analysed. Antioxidant treatment during both IVF and culture resulted in significantly faster development times to two cell cleavage (P IVF medium or embryo culture

  5. Long bone development in the Japanese quail (Coturnix coturnixjaponica) embryos.

    Science.gov (United States)

    Ahmed, Yasser A; Soliman, Soha A

    2013-09-15

    The current study was undertaken to describe the main histological development stages of long bones (tibia and femur) from Japanese quail (Coturnix coturnix japonica) embryos. Whole Limbs or just tibia and femur of fifty Japanese quail embryos of different ages were fixed and embedded in paraffin or Spurr's resin. Paraffin and semi-thin, respectively, were undertaken and examined with light microscopy. Limb bud was established at day 5 of incubation. Mesenchymal cells differentiated into chondrocytes forming a cartilage template in the position of the future tibia and femur at day 6 of incubation. At day 7 of incubation, the cartilage template enlarged and had the shape and position of the future tibia and femur. At day 8, central chondrocytes underwent hypertrophy and were surrounded by a periosteal bone collar. Cellular and vascular invasion from the bone collar into the central zone of the cartilage template, cartilage resorption and formation of marrow tunnel and finally peripheral calcification was seen. Vascular cartilage canals penetrating the epiphysis were observed at day 9 and the canals gradually increased in thickness and number toward the hatching day. Articular epiphyseal growth cartilage with resting, proliferative and hypertrophic zones was clearly established by day 10 of incubation. After 17 days of incubation, the zonation of the articular epiphyseal cartilage were much clear, many cartilage canals were present within the epiphyses. In epiphyses of tibia but not femur, foci of chondrocytes hypertrophy were noticed close to the cartilage canals. The current study timed the main histological sequences of development of tibia and femur of embryonic quail.

  6. The development of ovary in quail's embryo | Rong | African Journal ...

    African Journals Online (AJOL)

    The results showed that when embryo was hatched for 4 days, lots of primordial germ cells (PGCs) clustered in the region where gonad would be formed. On the 5th day of hatching, the gonad of the embryo began to be formed and exhibited the feature of ovary or testis. On the 7th hatching day, the right ovary began to ...

  7. Fresh or frozen? Classifying 'spare' embryos for donation to human embryonic stem cell research.

    Science.gov (United States)

    Ehrich, Kathryn; Williams, Clare; Farsides, Bobbie

    2010-12-01

    United Kingdom (UK) funding to build human embryonic stem cell (hESC) derivation labs within assisted conception units (ACU) was intended to facilitate the 'In-vitro fertilisation (IVF)-stem cell interface', including the flow of fresh 'spare' embryos to stem cell labs. However, in the three sites reported on here, which received this funding, most of the embryos used for hESC research came from long term cryopreservation storage and/or outside clinics. In this paper we explore some of the clinical, technical, social and ethical factors that might help to explain this situation. We report from our qualitative study of the ethical frameworks for approaching women/couples for donation of embryos to stem cell research. Members of staff took part in 44 interviews and six ethics discussion groups held at our study sites between February 2008 and October 2009. We focus here on their articulations of social and ethical, as well as scientific, dimensions in the contingent classification of 'spare' embryos, entailing uncertainty, fluidity and naturalisation in classifying work. Social and ethical factors include acknowledging and responding to uncertainty in classifying embryos; retaining 'fluidity' in the grading system to give embryos 'every chance'; tensions between standardisation and variation in enacting a 'fair' grading system; enhancement of patient choice and control, and prevention of regret; and incorporation of patients' values in construction of ethically acceptable embryo 'spareness' ('frozen' embryos, and embryos determined through preimplantation genetic diagnosis (PGD) to be genetically 'affected'). We argue that the success of the 'built moral environment' of ACU with adjoining stem cell laboratories building projects intended to facilitate the 'IVF-stem cell interface' may depend not only on architecture, but also on the part such social and ethical factors play in configuration of embryos as particular kinds of moral work objects. Copyright © 2010

  8. PreImplantation Factor (PIF correlates with early mammalian embryo development-bovine and murine models

    Directory of Open Access Journals (Sweden)

    Coulam Carolyn B

    2011-05-01

    Full Text Available Abstract Background PreImplantation Factor (PIF, a novel peptide secreted by viable embryos is essential for pregnancy: PIF modulates local immunity, promotes decidual pro-adhesion molecules and enhances trophoblast invasion. To determine the role of PIF in post-fertilization embryo development, we measured the peptide's concentration in the culture medium and tested endogenous PIF's potential trophic effects and direct interaction with the embryo. Methods Determine PIF levels in culture medium of multiple mouse and single bovine embryos cultured up to the blastocyst stage using PIF-ELISA. Examine the inhibitory effects of anti-PIF-monoclonal antibody (mAb added to medium on cultured mouse embryos development. Test FITC-PIF uptake by cultured bovine blastocysts using fluorescent microscopy. Results PIF levels in mouse embryo culture medium significantly increased from the morula to the blastocyst stage (ANOVA, P = 0.01. In contrast, atretic embryos medium was similar to the medium only control. Detectable - though low - PIF levels were secreted already by 2-cell stage mouse embryos. In single bovine IVF-derived embryos, PIF levels in medium at day 3 of culture were higher than non-cleaving embryos (control (P = 0.01 and at day 7 were higher than day 3 (P = 0.03. In non-cleaving embryos culture medium was similar to medium alone (control. Anti-PIF-mAb added to mouse embryo cultures lowered blastocyst formation rate 3-fold in a dose-dependent manner (2-way contingency table, multiple groups, X2; P = 0.01 as compared with non-specific mouse mAb, and medium alone, control. FITC-PIF was taken-up by cultured bovine blastocysts, but not by scrambled FITC-PIF (control. Conclusions PIF is an early embryo viability marker that has a direct supportive role on embryo development in culture. PIF-ELISA use to assess IVF embryo quality prior to transfer is warranted. Overall, our data supports PIF's endogenous self sustaining role in embryo development and the

  9. Development of a transient expression assay for detecting environmental oestrogens in zebrafish and medaka embryos

    Directory of Open Access Journals (Sweden)

    Lee Okhyun

    2012-06-01

    Full Text Available Abstract Background Oestrogenic contaminants are widespread in the aquatic environment and have been shown to induce adverse effects in both wildlife (most notably in fish and humans, raising international concern. Available detecting and testing systems are limited in their capacity to elucidate oestrogen signalling pathways and physiological impacts. Here we developed a transient expression assay to investigate the effects of oestrogenic chemicals in fish early life stages and to identify target organs for oestrogenic effects. To enhance the response sensitivity to oestrogen, we adopted the use of multiple tandem oestrogen responsive elements (EREc38 in a Tol2 transposon mediated Gal4ff-UAS system. The plasmid constructed (pTol2_ERE-TATA-Gal4ff, contains three copies of oestrogen response elements (3ERE that on exposure to oestrogen induces expression of Gal4ff which this in turn binds Gal4-responsive Upstream Activated Sequence (UAS elements, driving the expression of a second reporter gene, EGFP (Enhanced Green Fluorescent Protein. Results The response of our construct to oestrogen exposure in zebrafish embryos was examined using a transient expression assay. The two plasmids were injected into 1–2 cell staged zebrafish embryos, and the embryos were exposed to various oestrogens including the natural steroid oestrogen 17ß-oestradiol (E2, the synthetic oestrogen 17α- ethinyloestradiol (EE2, and the relatively weak environmental oestrogen nonylphenol (NP, and GFP expression was examined in the subsequent embryos using fluorescent microscopy. There was no GFP expression detected in unexposed embryos, but specific and mosaic expression of GFP was detected in the liver, heart, somite muscle and some other tissue cells for exposures to steroid oestrogen treatments (EE2; 10 ng/L, E2; 100 ng/L, after 72 h exposures. For the NP exposures, GFP expression was observed at 10 μg NP/L after 72 h (100 μg NP/L was toxic to the fish. We

  10. In vitro culture of mouse embryos in human amniotic fluid | Coetzee ...

    African Journals Online (AJOL)

    Human amniotic fluid was compared with Ham's F-10 culture medium as a possible alternative for use in in vitro fertilisation. The cleavage success of mouse embryos in human amniotic fluid (experimental group) was 92% compared with 86% in Ham's F-10 medium. It is concluded that human amniotic fluid is a viable ...

  11. Correction of β-thalassemia mutant by base editor in human embryos

    Directory of Open Access Journals (Sweden)

    Puping Liang

    2017-09-01

    Full Text Available Abstract β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB −28 (A>G mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor (BE system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB −28 (A>G mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB −28 (A>G homozygous mutation. Data showed that base editor could precisely correct HBB −28 (A>G mutation in the patient’s primary cells. To model homozygous mutation disease embryos, we constructed nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured (IVM oocytes. Notably, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB −28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.

  12. PENAL PROTECTION OF THE HUMAN EMBRYO. NEW CHALLENGES FOR ROMANIA

    OpenAIRE

    Laura STANILA

    2014-01-01

    The rapid developments in the last decades in the fields of biology and medicine raise important issues regarding the need to respect the human being both as an individual and in its membership in the human species. Medical acts performed both in research and in terms of curative work must be subordinated to a goal: the need to respect human dignity. In their desire to discover new treatments or just out of curiosity, biology and medicine can become instruments through an improper use, to end...

  13. Functional Genomics of 5- to 8-Cell Stage Human Embryos by Blastomere Single-Cell cDNA Analysis

    Science.gov (United States)

    Galán, Amparo; Montaner, David; Póo, M. Eugenia; Valbuena, Diana; Ruiz, Verónica; Aguilar, Cristóbal; Dopazo, Joaquín; Simón, Carlos

    2010-01-01

    Blastomere fate and embryonic genome activation (EGA) during human embryonic development are unsolved areas of high scientific and clinical interest. Forty-nine blastomeres from 5- to 8-cell human embryos have been investigated following an efficient single-cell cDNA amplification protocol to provide a template for high-density microarray analysis. The previously described markers, characteristic of Inner Cell Mass (ICM) (n = 120), stemness (n = 190) and Trophectoderm (TE) (n = 45), were analyzed, and a housekeeping pattern of 46 genes was established. All the human blastomeres from the 5- to 8-cell stage embryo displayed a common gene expression pattern corresponding to ICM markers (e.g., DDX3, FOXD3, LEFTY1, MYC, NANOG, POU5F1), stemness (e.g., POU5F1, DNMT3B, GABRB3, SOX2, ZFP42, TERT), and TE markers (e.g., GATA6, EOMES, CDX2, LHCGR). The EGA profile was also investigated between the 5-6- and 8-cell stage embryos, and compared to the blastocyst stage. Known genes (n = 92) such as depleted maternal transcripts (e.g., CCNA1, CCNB1, DPPA2) and embryo-specific activation (e.g., POU5F1, CDH1, DPPA4), as well as novel genes, were confirmed. In summary, the global single-cell cDNA amplification microarray analysis of the 5- to 8-cell stage human embryos reveals that blastomere fate is not committed to ICM or TE. Finally, new EGA features in human embryogenesis are presented. PMID:21049019

  14. The nuclear mitotic apparatus (NuMA) protein: localization and dynamics in human oocytes, fertilization and early embryos.

    Science.gov (United States)

    Alvarez Sedó, Cristian; Schatten, Heide; Combelles, Catherine M; Rawe, Vanesa Y

    2011-06-01

    The oocyte's meiotic spindle is a dynamic structure that relies on microtubule organization and regulation by centrosomes. Disorganization of centrosomal proteins, including the nuclear mitotic apparatus (NuMA) protein and the molecular motor complex dynein/dynactin, can lead to chromosomal instability and developmental abnormalities. The present study reports the distribution and function of these proteins in human oocytes, zygotes and early embryos. A total of 239 oocytes, 90 zygotes and discarded embryos were fixed and analyzed with confocal microscopy for NuMA and dynactin distribution together with microtubules and chromatin. Microtubule-associated dynein-dependent transport functions were explored by inhibiting phosphatase and ATPase activity with sodium-orthovanadate (SOV). At germinal vesicle (GV) stages, NuMA was dispersed across the nucleoplasm. After GV breaks down, NuMA became cytoplasmic before localizing at the spindle poles in metaphase I and II oocytes. Aberrant NuMA localization patterns were found during oocyte in vitro maturation. After fertilization, normal and abnormal pronuclear stage zygotes and embryos displayed translocation of NuMA to interphase nuclei. SOV treatment for up to 2 h induced lower maturation rates with chromosomal scattering and ectopic localization of NuMA. Accurate distribution of NuMA is important for oocyte maturation, zygote and embryo development in humans. Proper assembly of NuMA is likely necessary for bipolar spindle organization and human oocyte developmental competence.

  15. Arabidopsis phosphatidylinositol-phospholipase C2 (PLC2) is required for female gametogenesis and embryo development

    NARCIS (Netherlands)

    Di Fino, L.M.; D'Ambrosio, J.M.; Tejos, R.; van Wijk, R.; Lamattina, Lorenzo; Munnik, T.; Pagnussat, G.C.; Laxalt, A.M.

    2017-01-01

    MAIN conclusion: AtPLC2 is an essential gene in Arabidopsis, since it is required for female gametogenesis and embryo development. AtPLC2 might play a role in cell division during embryo-sac development and early embryogenesis. Phosphoinositide-specific phospholipase C (PI-PLC) plays an important

  16. Embryo dune development drivers: beach morphology, growing season precipitation, and storms

    NARCIS (Netherlands)

    Puijenbroek, van M.E.B.; Limpens, J.; Groot, de Alma; Riksen, M.J.P.M.; Gleichman, J.M.; Slim, P.A.; Dobben, van H.F.; Berendse, F.

    2017-01-01

    For development of embryo dunes on the highly dynamic land–sea boundary, summer growth and the absence of winter erosion are essential. Other than that, however, we know little about the specific conditions that favour embryo dune development. This study explores the boundary conditions for early

  17. Chromosome fragility at FRAXA in human cleavage stage embryos at risk for fragile X syndrome.

    Science.gov (United States)

    Verdyck, Pieter; Berckmoes, Veerle; De Vos, Anick; Verpoest, Willem; Liebaers, Inge; Bonduelle, Maryse; De Rycke, Martine

    2015-10-01

    Fragile X syndrome (FXS), the most common inherited intellectual disability syndrome, is caused by expansion and hypermethylation of the CGG repeat in the 5' UTR of the FMR1 gene. This expanded repeat, also known as the rare fragile site FRAXA, causes X chromosome fragility in cultured cells from patients but only when induced by perturbing pyrimidine synthesis. We performed preimplantation genetic diagnosis (PGD) on 595 blastomeres biopsied from 442 cleavage stage embryos at risk for FXS using short tandem repeat (STR) markers. In six blastomeres, from five embryos an incomplete haplotype was observed with loss of all alleles telomeric to the CGG repeat. In all five embryos, the incomplete haplotype corresponded to the haplotype carrying the CGG repeat expansion. Subsequent analysis of additional blastomeres from three embryos by array comparative genomic hybridization (aCGH) confirmed the presence of a terminal deletion with a breakpoint close to the CGG repeat in two blastomeres from one embryo. A blastomere from another embryo showed the complementary duplication. We conclude that a CGG repeat expansion at FRAXA causes X chromosome fragility in early human IVF embryos at risk for FXS. © 2015 Wiley Periodicals, Inc.

  18. The beginning of human life : status of embryo. Perspectives in Halakha (Jewish Religious Law).

    Science.gov (United States)

    Schenker, Joseph G

    2008-06-01

    The Jewish religion is characterized by a strict association between faith and practical precept. Jewish law has two sections, the written and the oral tradition. The foundation of the written law and the origin of authority is the Torah, the first five books of the Scripture. It is an expression of God's revelation, teaching and guiding humanity. The oral laws interpret, expand, and elucidate the written Torah and behavior patterns regulate new rules and customs. The main parts of the oral law are as follows: the Mishnah, the Talmud, Post-Talmudic Codes and. Responsa Literature. Life is a process that has a beginning and an end. The consensus about the time when human life really begins is still not reached among scientists, philosophers, ethicists, sociologists and theologizes. The scientific data suggested that a single developmental moment marking the beginning of human life does not exist. Current biological perspectives on when human life begins range through fertilization, gastrulation, to birth and even after. The development of a newborn is a smoothly continuous process. Procreation is acknowledged in the Bible to be the gift of God. The (Halachic) Jewish interpretation of when human life begins is extracted predominantly from procreation is acknowledged in the Bible to be the gift of God. The Jewish interpretation of when human life begins is extracted predominantly from The Halachic sources. The Bible does not make any other direct references regarding the beginning of human life. While the Talmud gives the full status of humanness to a child at birth, the rabbinical writings have partially extended the acquisition of humanness to the 13th postnatal day of life for full-term infants. The Babylonian Talmud Yevamot 69b states that: "the embryo is considered to be mere water until the fortieth day." Afterwards, it is considered subhuman until it is born. The issues of abortion, embryo research, multifetal reduction and cloning will be discussed according to

  19. Latrunculin A treatment prevents abnormal chromosome segregation for successful development of cloned embryos.

    Directory of Open Access Journals (Sweden)

    Yukari Terashita

    Full Text Available Somatic cell nuclear transfer to an enucleated oocyte is used for reprogramming somatic cells with the aim of achieving totipotency, but most cloned embryos die in the uterus after transfer. While modifying epigenetic states of cloned embryos can improve their development, the production rate of cloned embryos can also be enhanced by changing other factors. It has already been shown that abnormal chromosome segregation (ACS is a major cause of the developmental failure of cloned embryos and that Latrunculin A (LatA, an actin polymerization inhibitor, improves F-actin formation and birth rate of cloned embryos. Since F-actin is important for chromosome congression in embryos, here we examined the relation between ACS and F-actin in cloned embryos. Using LatA treatment, the occurrence of ACS decreased significantly whereas cloned embryo-specific epigenetic abnormalities such as dimethylation of histone H3 at lysine 9 (H3K9me2 could not be corrected. In contrast, when H3K9me2 was normalized using the G9a histone methyltransferase inhibitor BIX-01294, the Magea2 gene-essential for normal development but never before expressed in cloned embryos-was expressed. However, this did not increase the cloning success rate. Thus, non-epigenetic factors also play an important role in determining the efficiency of mouse cloning.

  20. Human immunodeficiency type-1 virus (HIV-1) infection in serodiscordant couples (SDCs) does not have an impact on embryo quality or intracytoplasmic sperm injection (ICSI) outcome.

    Science.gov (United States)

    Melo, Marco Antonio Barreto; Meseguer, Marcos; Bellver, José; Remohí, José; Pellicer, Antonio; Garrido, Nicolás

    2008-01-01

    To evaluate the embryo quality in our program for human immunodeficiency type-1 virus (HIV-1) serodiscordant couples (SDCs) with the male infected in comparison with a tubal-factor infertility control group. Retrospective case-control study. Instituto Valenciano de Infertilidad, Valencia, Spain. Thirty SDC and 79 control couples without HIV-1 infection attending for intracytoplasmic sperm injection (ICSI). Only first cycles were considered. Controlled ovarian hyperstimulation and ICSI in both groups; sperm wash, nested polymerase chain reaction (PCR) in semen sample, and capacitation by swim-up after thawing the semen sample in the SDC group; and sperm capacitation by swim-up after thawing the semen sample in the control group. ICSI procedure and embryo characteristics (fertilization, cleavage, embryo morphology, and development) and cycle outcome (ongoing pregnancy and miscarriage rates). Fertilization and cleavage rates were similar between the groups. On days 2 and 3 of embryo development, very similar embryo features were found between the groups. There was no difference in mean number of optimal embryos on day 3. When embryos were cultured up to 5-6 days, a significant increase in embryo blockage was found in the SDC group compared with the control group. The mean number of optimal blastocysts on day 6 was comparable in both groups. No difference was found regarding the number of cryopreserved and transferred embryos or implantation, pregnancy, multiple pregnancy, or miscarriage rates between the groups. HIV-1 infection in SDCs with infected males does not appear to have a significantly negative impact on embryo development or ICSI outcome.

  1. Calcium-sensing receptor (CASR) is involved in porcine in vitro fertilisation and early embryo development.

    Science.gov (United States)

    Liu, C; Liu, Y; Larsen, K; Hou, Y P; Callesen, H

    2017-07-17

    It has been demonstrated that extracellular calcium is necessary in fertilisation and embryo development but the mechanism is still not well understood. The present study mainly focussed on the extracellular calcium effector called the calcium-sensing receptor (CASR) and examined its expression in porcine gametes and embryos and its function during fertilisation and early embryo development. By using reverse transcription polymerase chain reaction, CASR was found to be expressed in porcine oocytes, spermatozoa and embryos at different developmental stages. Functionally, medium supplementation with a CASR agonist or an antagonist during in vitro fertilisation (IVF) and in vitro culture (IVC) was tested. During fertilisation, the presence of a CASR agonist increased sperm penetration rate and decreased polyspermy rate leading to an increased normal fertilisation rate. During embryo development, for the IVF embryos, agonist treatment during IVC significantly increased cleavage rate and blastocyst formation rate compared with the control group. Furthermore, parthenogenetically activated embryos showed similar results with lower cleavage and blastocyst formation rates in the antagonist group than in the other groups. It was concluded that CASR, as the effector of extracellular calcium, modulates porcine fertilisation and early embryo development.

  2. Human developmental anatomy: microscopic magnetic resonance imaging (μMRI) of four human embryos (from Carnegie Stage 10 to 20).

    Science.gov (United States)

    Lhuaire, Martin; Martinez, Agathe; Kaplan, Hervé; Nuzillard, Jean-Marc; Renard, Yohann; Tonnelet, Romain; Braun, Marc; Avisse, Claude; Labrousse, Marc

    2014-12-01

    Technological advances in the field of biological imaging now allow multi-modal studies of human embryo anatomy. The aim of this study was to assess the high magnetic field μMRI feasibility in the study of small human embryos (less than 21mm crown-rump) as a new tool for the study of human descriptive embryology and to determine better sequence characteristics to obtain higher spatial resolution and higher signal/noise ratio. Morphological study of four human embryos belonging to the historical collection of the Department of Anatomy in the Faculty of Medicine of Reims was undertaken by μMRI. These embryos had, successively, crown-rump lengths of 3mm (Carnegie Stage, CS 10), 12mm (CS 16), 17mm (CS 18) and 21mm (CS 20). Acquisition of images was performed using a vertical nuclear magnetic resonance spectrometer, a Bruker Avance III, 500MHz, 11.7T equipped for imaging. All images were acquired using 2D (transverse, sagittal and coronal) and 3D sequences, either T1-weighted or T2-weighted. Spatial resolution between 24 and 70μm/pixel allowed clear visualization of all anatomical structures of the embryos. The study of human embryos μMRI has already been reported in the literature and a few atlases exist for educational purposes. However, to our knowledge, descriptive or morphological studies of human developmental anatomy based on data collected these few μMRI studies of human embryos are rare. This morphological noninvasive imaging method coupled with other techniques already reported seems to offer new perspectives to descriptive studies of human embryology.

  3. Proteomics of desiccation tolerance during development and germination of maize embryos.

    Science.gov (United States)

    Huang, Hui; Møller, Ian Max; Song, Song-Quan

    2012-02-02

    Maize seeds were used to identify the key embryo proteins involved in desiccation tolerance during development and germination. Immature maize embryos (28N) during development and mature embryos imbibed for 72 h (72HN) are desiccation sensitive. Mature maize embryos (52N) during development are desiccation tolerant. Thiobarbituric acid reactive substance and hydrogen peroxide contents decreased and increased with acquisition and loss of desiccation tolerance, respectively. A total of 111 protein spots changed significantly (1.5 fold increase/decrease) in desiccation-tolerant and -sensitive embryos before (28N, 52N and 72HN) and after (28D, 52D and 72HD) dehydration. Nine pre-dominantly proteins, 17.4 kDa Class I heat shock protein 3, late embryogenesis abundant protein EMB564, outer membrane protein, globulin 2, TPA:putative cystatin, NBS-LRR resistance-like protein RGC456, stress responsive protein, major allergen Bet v 1.01C and proteasome subunit alpha type 1, accumulated during embryo maturation, decreased during germination and increased in desiccation-tolerant embryos during desiccation. Two proteins, Rhd6-like 2 and low-molecular-weight heat shock protein precursor, showed the inverse pattern. We infer that these eleven proteins are involved in seed desiccation tolerance. We conclude that desiccation-tolerant embryos make more economical use of their resources to accumulate protective molecules and antioxidant systems to deal with maturation drying and desiccation treatment. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Automated microinjection of recombinant BCL-X into mouse zygotes enhances embryo development.

    Science.gov (United States)

    Liu, Xinyu; Fernandes, Roxanne; Gertsenstein, Marina; Perumalsamy, Alagammal; Lai, Ingrid; Chi, Maggie; Moley, Kelle H; Greenblatt, Ellen; Jurisica, Igor; Casper, Robert F; Sun, Yu; Jurisicova, Andrea

    2011-01-01

    Progression of fertilized mammalian oocytes through cleavage, blastocyst formation and implantation depends on successful implementation of the developmental program, which becomes established during oogenesis. The identification of ooplasmic factors, which are responsible for successful embryo development, is thus crucial in designing possible molecular therapies for infertility intervention. However, systematic evaluation of molecular targets has been hampered by the lack of techniques for efficient delivery of molecules into embryos. We have developed an automated robotic microinjection system for delivering cell impermeable compounds into preimplantation embryos with a high post-injection survival rate. In this paper, we report the performance of the system on microinjection of mouse embryos. Furthermore, using this system we provide the first evidence that recombinant BCL-XL (recBCL-XL) protein is effective in preventing early embryo arrest imposed by suboptimal culture environment. We demonstrate that microinjection of recBCL-XL protein into early-stage embryos repairs mitochondrial bioenergetics, prevents reactive oxygen species (ROS) accumulation, and enhances preimplantation embryo development. This approach may lead to a possible treatment option for patients with repeated in vitro fertilization (IVF) failure due to poor embryo quality.

  5. Automated microinjection of recombinant BCL-X into mouse zygotes enhances embryo development.

    Directory of Open Access Journals (Sweden)

    Xinyu Liu

    Full Text Available Progression of fertilized mammalian oocytes through cleavage, blastocyst formation and implantation depends on successful implementation of the developmental program, which becomes established during oogenesis. The identification of ooplasmic factors, which are responsible for successful embryo development, is thus crucial in designing possible molecular therapies for infertility intervention. However, systematic evaluation of molecular targets has been hampered by the lack of techniques for efficient delivery of molecules into embryos. We have developed an automated robotic microinjection system for delivering cell impermeable compounds into preimplantation embryos with a high post-injection survival rate. In this paper, we report the performance of the system on microinjection of mouse embryos. Furthermore, using this system we provide the first evidence that recombinant BCL-XL (recBCL-XL protein is effective in preventing early embryo arrest imposed by suboptimal culture environment. We demonstrate that microinjection of recBCL-XL protein into early-stage embryos repairs mitochondrial bioenergetics, prevents reactive oxygen species (ROS accumulation, and enhances preimplantation embryo development. This approach may lead to a possible treatment option for patients with repeated in vitro fertilization (IVF failure due to poor embryo quality.

  6. [10]-Gingerdiols as the major metabolites of [10]-gingerol in zebrafish embryos and in humans and their hematopoietic effects in zebrafish embryos

    Science.gov (United States)

    Chen, Huadong; Soroka, Dominique N.; Haider, Jamil; Ferri-Lagneau, Karine F.; Leung, TinChung; Sang, Shengmin

    2013-01-01

    Gingerols are a series of major constituents in fresh ginger with the most abundant being [6]-, [8]-, and [10]-gingerols (6G, 8G, and 10G). We previously found that ginger extract and its purified components, especially 10G, potentially stimulate both the primitive and definitive waves of hematopoiesis (blood cell formation) in zebrafish embryos. However, it is still unclear if the metabolites of 10G retain the efficacy of the parent compound towards pathological anemia treatment. In the present study, we first investigated the metabolism of 10G in zebrafish embryos, and then explored the biotransformation of 10G in humans. Our results show that 10G was extensively metabolized in both zebrafish embryos and in humans, in which two major metabolites, (3S,5S)-[10]-gingerdiol and (3R,5S)-[10]-gingerdiol, were identified by analysis of the MSn spectra and comparison to authentic standards that we synthesized. After 24 hours of treatment of zebrafish embryos, 10G was mostly converted to its metabolites. Our results clearly indicate the reductive pathway is a major metabolic route for 10G in both zebrafish embryos and in humans. Furthermore, we investigated the hematopoietic effect of 10G and its two metabolites, which show similar hematopoietic effects as 10G in zebrafish embryos. PMID:23701129

  7. Human amniotic epithelial cells as feeder layer to derive and maintain human embryonic stem cells from poor-quality embryos

    Directory of Open Access Journals (Sweden)

    Daniela Ávila-González

    2015-09-01

    Full Text Available Data from the literature suggest that human embryonic stem cell (hESC lines used in research do not genetically represent all human populations. The derivation of hESC through conventional methods involve the destruction of viable human embryos, as well the use of mouse embryonic fibroblasts as a feeder layer, which has several drawbacks. We obtained the hESC line (Amicqui-1 from poor-quality (PQ embryos derived and maintained on human amniotic epithelial cells (hAEC. This line displays a battery of markers of pluripotency and we demonstrated the capacity of these cells to produce derivates of the three germ layers.

  8. Identification of chromosomal errors in human preimplantation embryos with oligonucleotide DNA microarray.

    Directory of Open Access Journals (Sweden)

    Lifeng Liang

    Full Text Available A previous study comparing the performance of different platforms for DNA microarray found that the oligonucleotide (oligo microarray platform containing 385K isothermal probes had the best performance when evaluating dosage sensitivity, precision, specificity, sensitivity and copy number variations border definition. Although oligo microarray platform has been used in some research fields and clinics, it has not been used for aneuploidy screening in human embryos. The present study was designed to use this new microarray platform for preimplantation genetic screening in the human. A total of 383 blastocysts from 72 infertility patients with either advanced maternal age or with previous miscarriage were analyzed after biopsy and microarray. Euploid blastocysts were transferred to patients and clinical pregnancy and implantation rates were measured. Chromosomes in some aneuploid blastocysts were further analyzed by fluorescence in-situ hybridization (FISH to evaluate accuracy of the results. We found that most (58.1% of the blastocysts had chromosomal abnormalities that included single or multiple gains and/or losses of chromosome(s, partial chromosome deletions and/or duplications in both euploid and aneuploid embryos. Transfer of normal euploid blastocysts in 34 cycles resulted in 58.8% clinical pregnancy and 54.4% implantation rates. Examination of abnormal blastocysts by FISH showed that all embryos had matching results comparing microarray and FISH analysis. The present study indicates that oligo microarray conducted with a higher resolution and a greater number of probes is able to detect not only aneuploidy, but also minor chromosomal abnormalities, such as partial chromosome deletion and/or duplication in human embryos. Preimplantation genetic screening of the aneuploidy by DNA microarray is an advanced technology used to select embryos for transfer and improved embryo implantation can be obtained after transfer of the screened normal

  9. Effect of MEM vitamins and forskolin on embryo development and vitrification tolerance of in vitro-produced pig embryos.

    Science.gov (United States)

    Cuello, C; Gomis, J; Almiñana, C; Maside, C; Sanchez-Osorio, J; Gil, M A; Sánchez, A; Parrilla, I; Vazquez, J M; Roca, J; Martinez, E A

    2013-01-30

    The aims of this study were (1) to determine the effect of in vitro maturation (IVM) medium supplementation with MEM vitamins on in vitro embryo development and sensitivity to vitrification of Day 6 blastocysts and (2) to evaluate whether the addition of forskolin to in vitro culture (IVC) medium enhances blastocyst survival following Super Open Pulled Straw (SOPS) vitrification. Cumulus-oocyte complexes (COCs; n=4000) were matured with 0.0% or 0.05% (v/v) MEM vitamins. After 44h of IVM, the oocytes were in vitro fertilized, and presumptive zygotes were cultured. At Day 5 of IVC, embryos from both experimental groups were cultured for 24h with 0 or 10μM forskolin, achieving a 2×2 factorial design. The blastocyst formation rate was assessed on Day 6, and subsets of samples from the four experimental groups were vitrified (n=469) or kept fresh (n=546). Fresh and vitrified-warmed blastocysts were cultured for 24h prior to embryo survival and total blastocyst cell number assessment. The MEM vitamins increased (Pvitamins improved the blastocyst formation rate, and the addition of 10μM forskolin to the culture medium increased survival in Day 6 in vitro-produced blastocysts after SOPS vitrification. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Optimized ex-ovo culturing of chick embryos to advanced stages of development.

    Science.gov (United States)

    Cloney, Kellie; Franz-Odendaal, Tamara Anne

    2015-01-24

    Research in anatomy, embryology, and developmental biology has largely relied on the use of model organisms. In order to study development in live embryos model organisms, such as the chicken, are often used. The chicken is an excellent model organism due to its low cost and minimal maintenance, however they present observational challenges because they are enclosed in an opaque eggshell. In order to properly view the embryo as it develops, the shell must be windowed or removed. Both windowing and ex ovo techniques have been developed to assist researchers in the study of embryonic development. However, each of the methods has limitations and challenges. Here, we present a simple, optimized ex ovo culture technique for chicken embryos that enables the observation of embryonic development from stage HH 19 into late stages of development (HH 40), when many organs have developed. This technique is easy to adopt in both undergraduate classes and more advanced research laboratories where embryo manipulations are conducted.

  11. Proposed guidelines on the nomenclature and annotation of dynamic human embryo monitoring by a time-lapse user group.

    Science.gov (United States)

    Ciray, H Nadir; Campbell, Alison; Agerholm, Inge Errebo; Aguilar, Jesús; Chamayou, Sandrine; Esbert, Marga; Sayed, Shabana

    2014-12-01

    Can the approach to, and terminology for, time-lapse monitoring of preimplantation embryo development be uniformly defined in order to improve the utilization and impact of this novel technology? The adoption of the proposed guidelines for defining annotation practice and universal nomenclature would help unify time-lapse monitoring practice, allow validation of published embryo selection algorithms and facilitate progress in this field. An increasing quantity of publications and communications relating to time-lapse imaging of in vitro embryo development have demonstrated the added clinical value of morphokinetic data for embryo selection. Several articles have identified similar embryo selection or de-selection variables but have termed them differently. An evidence-based consensus document exists for static embryo grading and selection but, to date, no such reference document is available for time-lapse methodology or dynamic embryo grading and selection. A series of meetings were held between September 2011 and May 2014 involving time-lapse users from seven different European centres. The group reached consensus on commonly identified and novel time-lapse variables. Definitions, calculated variables and additional annotations for the dynamic monitoring of human preimplantation development were all documented. Guidelines are proposed for a standard methodology and terminology for the of use time-lapse monitoring of preimplantation embryo development. The time-lapse variables considered by this group may not be exhaustive. This is a relatively new clinical technology and it is likely that new variables will be introduced in time, requiring revised guidelines. A different group of users from those participating in this process may have yielded subtly different terms or definitions for some of the morphokinetic variables discussed. Due to the technical processes involved in time-lapse monitoring, and acquisition of images at varied intervals through limited focal

  12. Embryo-maternal communication

    DEFF Research Database (Denmark)

    Østrup, Esben; Hyttel, Poul; Østrup, Olga

    2011-01-01

    Communication during early pregnancy is essential for successful reproduction. In this review we address the beginning of the communication between mother and developing embryo; including morphological and transcriptional changes in the endometrium as well as epigenetic regulation mechanisms...... directing the placentation. An increasing knowledge of the embryo-maternal communication might not only help to improve the fertility of our farm animals but also our understanding of human health and reproduction....

  13. Invasiveness of mouse embryos to human ovarian cancer cells HO8910PM and the role of MMP-9

    Directory of Open Access Journals (Sweden)

    Ding Xiaoyan

    2012-06-01

    Full Text Available Abstract Background Our previous work found that mouse embryos could invade malignant cancer cells. In the process of implantation, embryo trophoblast cells express matrix metalloproteinases and the invasive ability of trophoblast cells is proportional to matrix metalloproteinase-9 protein expression. So the purpose of this study is to observe the effects of mouse embryos on human ovarian cancer cells in the co-culture environment in vitro and explore the possible mechanism of matrix metalloproteinase-9. Methods Several groups of human ovarian cancer cells HO8910PM were co-cultured with mouse embryos for different time duration, after which the effects of mouse embryos on morphology and growth behavior of HO8910PM were observed under the light microscope real-time or by H.E staining. Apoptosis was detected under laser confocal microscope by Annexin V-EGFP/PI staining in situ. Invasion ability of tumor cells was studied by transwell experiments. After matrix metalloproteinase 9 (MMP −9 activity was inhibited by MMP-9 Inhibitor I, the interaction between mouse embryos and human ovarian cancer cells HO8910PM was observed. Results Mouse embryos were able to invade co-cultured human ovarian cancer cell layer which extended in the bottom of the culture dish, and gradually pushed away tumor cells to form their own growth space. The number of apoptosis tumor cells surrounding the embryo increased under laser confocal microscope. After co-cultured with mouse embryos, tumor cells invasive ability was lowered compared with the control group. After MMP-9 activity was inhibited, the interaction between mouse embryos and HO8910PM cells had no significant difference compared with the normal MMP-9 activity group. Conclusion Mouse embryos were able to invade human ovarian cancer cells in vitro and form their own growth space, promote apoptosis of human ovarian cancer cells and lower their invasive ability. The mouse embryo was still able to invade human

  14. Effect of different ovule isolation times on the embryo development of Campanula hybrids

    DEFF Research Database (Denmark)

    Röper, Anna Catharina; Lütken, Henrik Vlk; Hegelund, Josefine Nymark

    2012-01-01

    , hybridization between plant species is associated with many challenges to enable survival of the developing embryo. Here we present an optimised technique for embryo rescue via ovule isolation in selected intra- and interspecific Campanula hybrids. A frequent problem in embryo rescue is the malformation......Conventional breeding within natural cross border frames is not always sufficient to increase genetic variability and produce new characteristics such as leaf and flower shape or cold tolerance. Interspecific hybridisation is an approach to obtain new plants with desired features. However...... for 2-3 weeks the number of germinating embryos increased as compared to ovules isolated one week after pollination. Additionally, ovules isolated 2-3 weeks after pollination showed an increased embryo germination rate. Among the Campanula hybrids, produced here from both the intraspecific crosses...

  15. Ultrastructural analyses of somatic embryo initiation, development and polarity establishment from mesophyll cells of Dactylis glomerata

    Science.gov (United States)

    Vasilenko, A.; McDaniel, J. K.; Conger, B. V.

    2000-01-01

    Somatic embryos initiate and develop directly from single mesophyll cells in in vitro-cultured leaf segments of orchardgrass (Dactylis glomerata L.). Embryogenic cells establish themselves in the predivision stage by formation of thicker cell walls and dense cytoplasm. Electron microscopy observations for embryos ranging from the pre-cell-division stage to 20-cell proembryos confirm previous light microscopy studies showing a single cell origin. They also confirm that the first division is predominantly periclinal and that this division plane is important in establishing embryo polarity and in determining the embryo axis. If the first division is anticlinal or if divisions are in random planes after the first division, divisions may not continue to produce an embryo. This result may produce an embryogenic cell mass, callus formation, or no structure at all. Grant numbers: NAGW-3141, NAG10-0221.

  16. Embryo sac development in some South African cultivars of Lantana camara

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    J. J. Spies

    1982-12-01

    Full Text Available Twenty embryo sacs from each of 20 different  Lantana camara L. cultivars naturalized in South Africa were examined. The normal sexual embryo sacs were monosporic 8-nucleated embryo sacs of the polygonum type and were encountered in 55% of the material examined. Several deviations from this pattern were recorded. Occasionally one of the nuclei failed to develop into a synergid, resulting in three polar nuclei. Contrary to published information, the antipodal cells did not increase in size, nor was there an increase in the number of nuclei per cell. Although the occurrence of sexuality is confirmed, no definite evidence exists for the occurrence of apomixis. The occurrence of two embryo sacs per locule might be the result of either apospory or of sexuality whereby two embryo sacs were formed from two megaspores.

  17. Cardio-respiratory development in bird embryos: new insights from a venerable animal model

    Directory of Open Access Journals (Sweden)

    Warren W. Burggren

    Full Text Available ABSTRACT The avian embryo is a time-honored animal model for understanding vertebrate development. A key area of extensive study using bird embryos centers on developmental phenotypic plasticity of the cardio-respiratory system and how its normal development can be affected by abiotic factors such as temperature and oxygen availability. Through the investigation of the plasticity of development, we gain a better understanding of both the regulation of the developmental process and the embryo's capacity for self-repair. Additionally, experiments with abiotic and biotic stressors during development have helped delineate not just critical windows for avian cardio-respiratory development, but the general characteristics (e.g., timing and dose-dependence of critical windows in all developing vertebrates. Avian embryos are useful in exploring fetal programming, in which early developmental experiences have implications (usually negative later in life. The ability to experimentally manipulate the avian embryo without the interference of maternal behavior or physiology makes it particularly useful in future studies of fetal programming. The bird embryo is also a key participant in studies of transgenerational epigenetics, whether by egg provisioning or effects on the germline that are transmitted to the F1 generation (or beyond. Finally, the avian embryo is heavily exploited in toxicology, in which both toxicological testing of potential consumer products as well as the consequences of exposure to anthropogenic pollutants are routinely carried out in the avian embryo. The avian embryo thus proves useful on numerous experimental fronts as an animal model that is concurrently both of adequate complexity and sufficient simplicity for probing vertebrate cardio-respiratory development.

  18. Chromosomal preparations of human triploid zygotes and embryos fertilized in vitro.

    Science.gov (United States)

    Macas, E; Suchanek, E; Grizelj, V; Puharic, I; Simunic, V

    1988-12-01

    Forty-eight zygotes with more than two pronuclei were identified after in vitro fertilization, representing 6.1% of all fertilized oocytes. The chromosome preparations from pronuclear stage to the cleaved human embryos were examined. Prophase was found in eight out of ten zygotes. The spreading of chromosomes allowed an adequate counting in only two cases. Six of the eight preparations displayed a late prophase. In this stage each haploid group of chromosomes can be analysed separately. Kariogamy usually occurred 4 to 5 h after the pronuclei had disappeared, and polyploid number of chromosomes were found in well-spread metaphases. The chromosomal preparations were made for eleven human embryos arising from zygotes with three pronuclei. Out of ten preparations, where the chromosomes could be counted, seven embryos (70%) contained hypodiploidic groups of chromosomes. In two of the cases, however, triploid metaphases were found, and in the last one a triploid/diploid mosaicism.

  19. Early maternal serum ß-human chorionic gonadotropin (ß-hCG) levels and sex-related growth difference of IVF embryos.

    Science.gov (United States)

    Esh-Broder, Efrat; Oron, Galia; Son, Weon-Young; Holzer, Hananel; Tulandi, Togas

    2015-10-01

    Maternal serum ß-human chorionic gonadotropin (ß-hCG) represents the trophoblastic cell mass and is an indirect measurement of embryo development at early implantation stage. Studies in animals and human embryos detected sex-related growth differences (SRGD) in favour of male embryos during the pre-implantation period. The purpose of our study was to correlate SRGD and maternal serum ß-hCG at 16 days after embryo transfer. We retrospectively analysed all (fresh and frozen) non-donor, single embryo transfers (SET), elective and not elective, that were performed between December 2008 and December 2013. We included ß-hCG values from day 16 after oocyte collection of pregnancies resulting in live birth. Neonatal gender was retrieved from patient files. Male and female embryos were further grouped to cleavage and blastocyst stage transfers. Regression analysis for confounding variables included maternal age, maternal body mass index (BMI), use of micromanipulation (ICSI), embryo quality (grade), assisted hatching, day of transfer and fresh or frozen embryo transfer. Seven hundred eighty-six non-donor SETs resulted in live birth. After including only day 16 serum ß-hCG results, 525 SETs were analysed. Neonatal gender was available for 522 cases. Mean maternal serum ß-hCG levels were similar, 347 ± 191 IU/L in the male newborn group and 371 ± 200 IU/L in the female group. The difference between ß-hCG levels remained insignificant after adjusting for confounding variables. Early maternal ß-hCG levels after embryo transfers did not represent SRGD in our study.

  20. Cytoskeleton involvement during human cytomegalovirus replicative cycle in human embryo fibroblasts.

    Science.gov (United States)

    Arcangeletti, M C; Pinardi, F; Medici, M C; Pilotti, E; De Conto, F; Ferraglia, F; Landini, M P; Chezzi, C; Dettori, G

    2000-07-01

    Several studies indicate that viruses can induce different cytoskeletal modifications. The present investigation examines the possible involvement of human embryo fibroblast cytoskeleton in the replication of human cytomegalovirus (HCMV). Significant cytoskeletal modifications occur in infected cells; specifically, microfilament depolymerization is observed very early during the HCMV replicative cycle, whilst microtubules and intermediate filaments do not undergo any change for longer times after infection. Our data focus, in particular, on microfilament depolymerization, which starts within the first hour of the replicative cycle, and on the significance of this event, as a CMV-induced mechanism to modify the post-transcriptional regulation of cellular gene expression for its own benefit. Among the possible mechanisms exploited by HCMV to induce microfilament modifications, one might involve the cellular ADP-ribosylation activity, which is increased by HCMV very early in the infectious cycle. Experiments carried out on HCMV-infected cells, in the presence of ADP-ribosylation inhibitors, seem to confirm this hypothesis.

  1. Promoting Cas9 degradation reduces mosaic mutations in non-human primate embryos.

    Science.gov (United States)

    Tu, Zhuchi; Yang, Weili; Yan, Sen; Yin, An; Gao, Jinquan; Liu, Xudong; Zheng, Yinghui; Zheng, Jiezhao; Li, Zhujun; Yang, Su; Li, Shihua; Guo, Xiangyu; Li, Xiao-Jiang

    2017-02-03

    CRISPR-Cas9 is a powerful new tool for genome editing, but this technique creates mosaic mutations that affect the efficiency and precision of its ability to edit the genome. Reducing mosaic mutations is particularly important for gene therapy and precision genome editing. Although the mechanisms underlying the CRSIPR/Cas9-mediated mosaic mutations remain elusive, the prolonged expression and activity of Cas9 in embryos could contribute to mosaicism in DNA mutations. Here we report that tagging Cas9 with ubiquitin-proteasomal degradation signals can facilitate the degradation of Cas9 in non-human primate embryos. Using embryo-splitting approach, we found that shortening the half-life of Cas9 in fertilized zygotes reduces mosaic mutations and increases its ability to modify genomes in non-human primate embryos. Also, injection of modified Cas9 in one-cell embryos leads to live monkeys with the targeted gene modifications. Our findings suggest that modifying Cas9 activity can be an effective strategy to enhance precision genome editing.

  2. Promoting Cas9 degradation reduces mosaic mutations in non-human primate embryos

    Science.gov (United States)

    Tu, Zhuchi; Yang, Weili; Yan, Sen; Yin, An; Gao, Jinquan; Liu, Xudong; Zheng, Yinghui; Zheng, Jiezhao; Li, Zhujun; Yang, Su; Li, Shihua; Guo, Xiangyu; Li, Xiao-Jiang

    2017-01-01

    CRISPR-Cas9 is a powerful new tool for genome editing, but this technique creates mosaic mutations that affect the efficiency and precision of its ability to edit the genome. Reducing mosaic mutations is particularly important for gene therapy and precision genome editing. Although the mechanisms underlying the CRSIPR/Cas9-mediated mosaic mutations remain elusive, the prolonged expression and activity of Cas9 in embryos could contribute to mosaicism in DNA mutations. Here we report that tagging Cas9 with ubiquitin-proteasomal degradation signals can facilitate the degradation of Cas9 in non-human primate embryos. Using embryo-splitting approach, we found that shortening the half-life of Cas9 in fertilized zygotes reduces mosaic mutations and increases its ability to modify genomes in non-human primate embryos. Also, injection of modified Cas9 in one-cell embryos leads to live monkeys with the targeted gene modifications. Our findings suggest that modifying Cas9 activity can be an effective strategy to enhance precision genome editing. PMID:28155910

  3. Development of HY1 hybrid embryos between a cultivar of ...

    African Journals Online (AJOL)

    However, the P. vulgaris NI637 cultivar and the wild form NI1108 of P. coccineus present abilities for combination. ... explain their possible causes, and describe the expression of abortion symptoms in hybrid embryos during the embryogenesis within the Phaseolus genus, in view of improving the common bean production.

  4. Microspore embryogenesis: reprogramming cell fate from pollen to embryo development

    NARCIS (Netherlands)

    Hui Li,

    2014-01-01

    Microspore embryogenesis is an expression of plant cell totipotency that leads to the production of haploid embryos. Besides being a widely exploited plant breeding tool, microspore embryogenesis is also a fascinating system that can be used to obtain a deeper mechanistic understanding of plant

  5. Growth factors effects on preimplantation development of mouse embryos exposed to tumor necrosis factor alpha.

    Science.gov (United States)

    Głabowski, Wojciech; Kurzawa, Rafał; Wiszniewska, Barbara; Baczkowski, Tomasz; Marchlewicz, Mariola; Brelik, Paweł

    2005-03-01

    The success rates of assisted reproduction techniques are still unsatisfactory. Relatively few in vitro cultured embryos reach the blastocyst stage. The purpose of the study was to evaluate the protective potential of epidermal growth factor (EGF), insulin-like growth factors 1 and 2 (IGF-I, IGF-II) and stem cell factor (SCF) on in vitro development of pre-implantation mouse embryos exposed to tumor necrosis factor alpha (TNFalpha). C3B6F1 female mice were superovulated with 5 IU of pregnant mare serum gonadotropin (PMSG) and 48 h later with 5IU of equine chorionic gonadotropin (eCG). Following the second injection females were mated with DBA males. Two cell embryos were flushed out from the fallopian tubes 40 h after eCG administration. After retrieval, the embryos were divided into control and experimental media and incubated in groups of ten for 96 h (37 degrees C, 5%CO(2), in droplets of 50 microl under mineral oil). In the first part of experiment, the embryo development was tested in media containing EGF, IGF-I, IGF-II, SCF, TNF-alpha (1 to 1000 ng/ml). In the second part of the study, the development of embryos was examined in medium containing 100 ng/ml TNFalpha and one of following factors: IGF-I, IGF-II; EGF or SCF (100 ng/ml). During the culture embryos were examined at 24 hours intervals to assess the embryo development. Blastocyst rate was determined following 96 hours of culture. Evaluation of total blastocyst cell number (TB) and inner cell mass (ICM) was also performed. TNFalpha significantly reduced (p<0.05) the blastocyst rates as well as TB and ICM. The examined growth factors improved the development of embryos exposed to TNFalpha. Thus, in this study, the protective action of IGF-I and II, EGF and SCF against the detrimental influence of TNFalpha was demonstrated.

  6. Pro-apoptotic Effect of Pifithrin-α on Preimplantation Porcine Fertilized Embryo Development

    Directory of Open Access Journals (Sweden)

    Brendan Mulligan

    2012-12-01

    Full Text Available The aim of this study was to investigate the impact of a reported p53 inhibitor, pifithrin-α (PFT-α, on preimplantation porcine in vitro fertilized (IVF embryo development in culture. Treatment of PFT-α was administered at both early (0 to 48 hpi, and later stages (48 to 168 hpi of preimplantation development, and its impact upon the expression of five genes related to apoptosis (p53, bak, bcl-xL, p66Shc and caspase3, was assessed in resulting d 7 blastocysts, using real-time quantitative PCR. Total cell numbers, along with the number of apoptotic nuclei, as detected by the in situ cell death detection assay, were also calculated on d 7 in treated and non-treated control embryos. The results indicate that PFT-α, when administered at both early and later stages of porcine IVF embryo development, increases the incidence of apoptosis in resulting blastocysts. When administered at early cleavage stages, PFT-α treatment was shown to reduce the developmental competence of porcine IVF embryos, as well as reducing the quality of resulting blastocysts in terms of overall cell numbers. In contrast, at later stages, PFT-α administration resulted in marginally increased blastocyst development rates amongst treated embryos, but did not affect cell numbers. However, PFT-α treatment induced apoptosis and apoptotic related gene expression, in all treated embryos, irrespective of the timing of treatment. Our results indicate that PFT-α may severely compromise the developmental potential of porcine IVF embryos, and is a potent apoptotic agent when placed into porcine embryo culture media. Thus, caution should be exercised when using PFT-α as a specific inhibitor of p53 mediated apoptosis, in the context of porcine IVF embryo culture systems.

  7. Expression of plasminogen activators in preimplantation rat embryos developed in vivo and in vitro

    Directory of Open Access Journals (Sweden)

    Har-Vardi Iris

    2005-02-01

    Full Text Available Abstract Background Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs have been implicated in mammalian fertilization, early stages of development and embryo implantation. The invasion of trophoblast cells into the endometrium during the implantation process can be blocked by inhibitors of serine proteases, illustrating the role of these enzymes in the invasion process. As in vitro developing embryos resulted in lower implantation rate than those developed in vivo we assume that a reduced PAs activity may lead to it. There is hardly any information regarding qualitative or quantitative differences in expression of PAs in preimplantation embryos, or comparisons between in vivo and in vitro developed embryos. The purpose of this study was to assess the expression of urokinase type (uPA and tissue type (tPA plasminogen activators in in vivo and in vitro preimplantation development in rat embryos using immunofluorescence confocal microscopy and computerized image analysis. Methods Zygotes, 2-cell, 4-cell, 8-cell, morula and blastocyst stages of development were flushed from the reproductive tract (control groups of Wistar rats. Zygotes were flushed and grown in vitro to the above mentioned developmental stages and comprised the experimental groups. Immunofluorescence microscopy and computerized image analysis were used to evaluate both qualitative (localization and quantitative expression of plasminogen activators. Results uPA and tPA were found to be expressed in rat embryos throughout their preimplantation development, both in vivo and in vitro. While uPA was localized mainly in the cell cytoplasm, the tPA was detected mainly on cell surface and in the perivitelline space. In blastocysts, both in vivo and in vitro, uPA and tPA were localized in the trophectoderm cells. Total uPA content per embryo was higher in the in vivo as compared with the in vitro developed embryos at all stages

  8. AFSC/RACE/SAP/Long: Data from: Embryo development in golden king crab, Lithodes aequispina.

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The data from this study, describes embryo development in Golden king crab, Lithodes aequispinus. Six female multiparous golden king crab were captured from the...

  9. Cardio-respiratory development in bird embryos: new insights from a venerable animal model

    National Research Council Canada - National Science Library

    Burggren, Warren W; Santin, Josele Flores; Antich, Maria Rojas

    2016-01-01

    .... A key area of extensive study using bird embryos centers on developmental phenotypic plasticity of the cardio-respiratory system and how its normal development can be affected by abiotic factors...

  10. AFSC/RACE/SAP/Swiney: Primiparous and multiparous Tanner crab egg extrusion, embryo development and hatching

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This study compares timing of egg extrusion, embryo development, timing and duration of eclosion, and incubation periods of Kodiak, Alaska primiparous and...

  11. HIGHLY METHYL ESTERIFIED SEEDS is a pectin methyl esterase involved in embryo development.

    Science.gov (United States)

    Levesque-Tremblay, Gabriel; Müller, Kerstin; Mansfield, Shawn D; Haughn, George W

    2015-03-01

    Homogalacturonan pectin domains are synthesized in a highly methyl-esterified form that later can be differentially demethyl esterified by pectin methyl esterase (PME) to strengthen or loosen plant cell walls that contain pectin, including seed coat mucilage, a specialized secondary cell wall of seed coat epidermal cells. As a means to identify the active PMEs in seed coat mucilage, we identified seven PMEs expressed during seed coat development. One of these, HIGHLY METHYL ESTERIFIED SEEDS (HMS), is abundant during mucilage secretion, peaking at 7 d postanthesis in both the seed coat and the embryo. We have determined that this gene is required for normal levels of PME activity and homogalacturonan methyl esterification in the seed. The hms-1 mutant displays altered embryo morphology and mucilage extrusion, both of which are a consequence of defects in embryo development. A significant decrease in the size of cells in the embryo suggests that the changes in embryo morphology are a consequence of lack of cell expansion. Progeny from a cross between hms-1 and the previously characterized PME inhibitor5 overexpression line suggest that HMS acts independently from other cell wall-modifying enzymes in the embryo. We propose that HMS is required for cell wall loosening in the embryo to facilitate cell expansion during the accumulation of storage reserves and that its role in the seed coat is masked by redundancy. © 2015 American Society of Plant Biologists. All Rights Reserved.

  12. Optimized Ex-ovo Culturing of Chick Embryos to Advanced Stages of Development

    OpenAIRE

    Cloney, Kellie; Franz-Odendaal, Tamara Anne

    2015-01-01

    Research in anatomy, embryology, and developmental biology has largely relied on the use of model organisms. In order to study development in live embryos model organisms, such as the chicken, are often used. The chicken is an excellent model organism due to its low cost and minimal maintenance, however they present observational challenges because they are enclosed in an opaque eggshell. In order to properly view the embryo as it develops, the shell must be windowed or removed. Both windowin...

  13. A curious abnormally developed embryo of the pill millipede Glomeris marginata (Villers, 1789)

    OpenAIRE

    Janssen, Ralf

    2013-01-01

    This paper reports on an abnormally developed embryo (ADE) of the common pill millipede Glomeris marginata. This ADE represents a modified case of Duplicitas posterior, in which two posterior ends are present, but only one anterior end. While the major posterior germ band of the embryo appears almost normally developed, the minor posterior germ band is heavily malformed, has no clear correlation to the single head, little or no ventral tissue, and a minute amount of yolk. The anterior end of ...

  14. A curious abnormally developed embryo of the pill millipede Glomeris marginata (Villers, 1789)

    OpenAIRE

    Ralf Janssen

    2013-01-01

    Abstract This paper reports on an abnormally developed embryo (ADE) of the common pill millipede Glomeris marginata . This ADE represents a modified case of Duplicitas posterior , in which two posterior ends are present, but only one anterior end. While the major posterior germ band of the embryo appears almost normally developed, the minor posterior germ band is heavily malformed, has no clear correlation to the single head, little or no ventral tissue, and a minute amount of yolk. The anter...

  15. Predictive value of plasma human chorionic gonadotropin measured 14 days after Day-2 single embryo transfer

    DEFF Research Database (Denmark)

    Løssl, Kristine; Oldenburg, Anna; Toftager, Mette

    2017-01-01

    Introduction: Prediction of pregnancy outcome after in vitro fertilization is important for patients and clinicians. Early plasma human chorionic gonadotropin (p-hCG) levels are the best known predictor of pregnancy outcome, but no studies have been restricted to single embryo transfer (SET) of Day......-2 embryos. The aim of the present study was to investigate the predictive value of p-hCG measured exactly 14 days after the most commonly used Day-2 SET on pregnancy, delivery, and perinatal outcome. Material and methods: A retrospective analysis of prospectively collected data on 466 women who had...

  16. Conception and development of the Second Life® Embryo Physics Course.

    Science.gov (United States)

    Gordon, Richard

    2013-06-01

    The study of embryos with the tools and mindset of physics, started by Wilhelm His in the 1880s, has resumed after a hiatus of a century. The Embryo Physics Course convenes online allowing interested researchers and students, who are scattered around the world, to gather weekly in one place, the virtual world of Second Life®. It attracts people from a wide variety of disciplines and walks of life: applied mathematics, artificial life, bioengineering, biophysics, cancer biology, cellular automata, civil engineering, computer science, embryology, electrical engineering, evolution, finite element methods, history of biology, human genetics, mathematics, molecular developmental biology, molecular biology, nanotechnology, philosophy of biology, phycology, physics, self-reproducing systems, stem cells, tensegrity structures, theoretical biology, and tissue engineering. Now in its fifth year, the Embryo Physics Course provides a focus for research on the central question of how an embryo builds itself.

  17. Embryo manipulation and experimentation.

    Science.gov (United States)

    Warren, M A

    1991-09-01

    I have argued that early human embryos are not human beings, and do not have normal rights. Like human sperm and ova, they are both alive and biologically human. However, they lack the physiological development necessary to sustain a capacity for sentience. If Ford is right, then they are not yet individual human organisms. But the more important point is that their lack of a capacity for sentience makes them inappropriate candidates for the ascription of moral rights. Thus, research on human embryos produced in vitro is not a wrong against them--at least so long as experimentally manipulated embryos are not returned to the womb, or artificially gestated to a stage at which they might become sentient. Some of the more difficult issues about embryo experimentation involve the rights of women as experimental subject and donors. The consent of both male and female gamete donors should normally be required for the production or experimental use of IVF embryos. (Possible exceptions might include cases in which one or both progenitors have died, and the survivor or other responsible family member wished to donate the (frozen) IVF embryos for research or other uses.) However, it is women's rights that are most apt to be endangered, for example, if the large scale therapeutic or commercial use of human embryos leads to a demand for large numbers of ova. Thus, it is vital that researchers and policy-makers heed feminist concerns about embryo research and the new biomedical technologies it may yield. Given adequate information and appropriate procedural protections, women are capable of making autonomous decisions about donating ova or embryos for biomedical research. But regulatory safeguards are needed to ensure against their being coerced, deceived, or manipulated into becoming ovum or embryo donors. As Daniel Callahan has detailed, biomedical technology has reached the point where we can no longer afford to provide everyone with all of the innovative therapies that might

  18. An HPLC method for the determination of selected amino acids in human embryo culture medium.

    Science.gov (United States)

    Drábková, Petra; Andrlová, Lenka; Kanďár, Roman

    2017-02-01

    A method for the determination of selected amino acids in culture medium using HPLC with fluorescence detection is described. Twenty hours after intra-cytoplasmic sperm injection, one randomly selected zygote was transferred to the culture medium. After incubation (72 h after fertilization), the culture medium in which the embryo was incubated and blank medium was immediately stored at -80°C. Filtered medium samples were derivatized with ortho-phthalaldehyde (naphthalene-2,3-dicarboxaldehyde), forming highly fluorescent amino acids derivatives. Reverse-phase columns (LichroCART, Purospher STAR RP18e or Ascentis Express C18 ) were used for the separation. The derivatives were analyzed by gradient elution with a mobile phase containing ethanol and sodium dihydrogen phosphate. The analytical performance of this method is satisfactory for all amino acids; the intra-assay coefficients of variation were amino acids before and after human embryo cultivation were observed. After embryo incubation, the levels of all amino acids in the medium were increased, apart from aspartate and asparagine. After the cultivation of some embryos, amino acids which were not part of the medium were detected. Low amino acids turnover was observed in some embryos. Copyright © 2016 John Wiley & Sons, Ltd.

  19. Optimized protocol for cryopreservation of human eggs improves developmental competence and implantation of resulting embryos.

    Science.gov (United States)

    Wang, Cassie T; Liang, Lifeng; Witz, Craig; Williams, Dan; Griffith, Jason; Skorupski, Josh; Haddad, Ghassan; Gill, Jimmy; Wang, Weihua

    2013-02-13

    Successful egg cryopreservation has many potential benefits to a variety of patients. However, a superior standard protocol describing all aspects of oocyte cryopreservation has not yet been identified. Oocyte cryopreservation is still a technical challenge for many infertility clinics. To maintain satisfactory clinical outcomes, there is a need to develop an easy to use, yet efficient laboratory protocol. The present study was designed to examine if human embryos resulting from eggs frozen with an optimized vitrification protocol have similar developmental competence as those from fresh eggs. Twenty recipients received donated eggs vitrified with a protocol in which short exposure time to the vitrification solution was used and 23 recipients received donated eggs and 6 patients had their own eggs vitrified with a modified protocol in which long exposure time to the vitrification solution was used. After warming, egg survival, fertilization, cleavage, blastocyst formation, clinical pregnancy and implantation rates were compared. The developmental competence of eggs vitrified with the optimized protocol was further compared with fresh eggs donated from the same donors. There was no difference in the oocyte survival, fertilization, cleavage, clinical pregnancy or implantation rates between the short and long protocol groups. However, blastocyst formation rate was significantly (P eggs vitrified with long protocol and fresh eggs from the same donors (12) were compared in 39 recipients, no differences were observed in terms of fertilization (86.4 vs 80.1%), blastocyst formation (50.0 vs 59.2%), clinical pregnancy (63.2 vs 60.0%) and implantation (41.7 vs 44.7%) rates. Four out of 6 patients had ongoing pregnancy after transfer of embryos from their own frozen eggs with a 46.2% implantation rate. These results indicate that blastocyst development is an appropriate measure for egg survival after cryopreservation and frozen eggs have similar developmental potential as

  20. Supramolecular delivery of photoactivatable fluorophores in developing embryos

    Science.gov (United States)

    Zhang, Yang; Tang, Sicheng; Sansalone, Lorenzo; Thapaliya, Ek Raj; Baker, James D.; Raymo, Françisco M.

    2017-02-01

    The identification of noninvasive strategies to monitor dynamics within living organisms in real time is essential to elucidate the fundamental factors governing a diversity of biological processes. This study demonstrates that the supramolecular delivery of photoactivatable fluorophores in Drosophila melanogaster embryos allows the real-time tracking of translocating molecules. The designed photoactivatable fluorophores switch from an emissive reactant to an emissive product with spectrally-resolved fluorescence, under moderate blue-light irradiation conditions. These hydrophobic fluorescent probes can be encapsulated within supramolecular hosts and delivered to the cellular blastoderm of the embryos. Thus, the combination of supramolecular delivery and fluorescence photoactivation translates into a noninvasive method to monitor dynamics in vivo and can evolve into a general chemical tool to track motion in biological specimens.

  1. Development block of golden hamster ICSI embryos is associated with decreased expression of HDAC1, HSPA1A and MYC.

    Science.gov (United States)

    Pan, Xiaoyan; Kong, Delong; Liu, Limei; Gao, Fei; Zhang, Xueming; Tang, Bo; Li, Ziyi

    2014-11-01

    We have investigated the mechanism for embryo development block in vitro and to improve the development rate of golden hamster embryos in vitro. Intracytoplasmic sperm injection (ICSI) technique was used to produce golden hamster ICSI embryos. The changes in the histone acetylation and the expression of histone deacetylase and related genes were analyzed by immunocytochemical staining and real-time PCR both in golden hamster in vivo embryos and in ICSI embryos. Aged oocytes significantly increased the oocyte spontaneous activation rate. In vitro cultured ICSI embryos suffered from severe development block in M199TE medium. Expression of histone deacetylase 1 (HDAC1) was significantly decreased in the nuclei of the arrested ICSI 2-cell embryos, and its nuclear and cytoplasmic expression pattern was also markedly altered. The acetylation level of H4K5, however, was not significantly changed between golden hamster in vivo embryos and ICSI embryos. HSPA1A and MYC, the marker genes for zygotic genome activation (ZGA), were transcriptionally decreased in arrested ICSI 2-cell embryos. Transcription of HDAC1 was also downregulated in these embryos, whereas the mRNA expression of the proapoptotic gene, BAX, was not changed. These results indicate that the golden hamster ICSI embryo development block during ZGA is associated with decreased nuclear expression and altered expression of HDAC1. HSPA1A, MYC, and HDAC1 mRNA levels, which decrease, resulting in ZGA failure. © 2014 International Federation for Cell Biology.

  2. TBP dynamics during mouse oocyte meiotic maturation and early embryo development.

    Science.gov (United States)

    Sun, Shao-Chen; Wang, Xu-Guang; Ma, Xue-Shan; Huang, Xian-Ju; Li, Juan; Liu, Hong-Lin

    2013-01-01

    To maintain cell lineage, cells develop a mechanism which can transmit the gene activity information to the daughter cells. In mitosis, TBP (TATA-binding protein), a transcription factor which belongs to TFIID was associated with M phase chromosomes and was proved to be a bookmark for cellular memory. Although previous work showed that TBP was dispensable for mouse oocyte maturation and early embryo development, exogenous TBP protein was detected in the nuclear of oocytes and early embryos. It is still unknown whether exogenous TBP can associate with condensed chromosomes during meiosis and mouse early embryo development. In present study by the injection of GFP-tagged TBP mRNA we for the first time investigated TBP dynamics in mouse early embryos and confirmed its localization pattern in oocytes. The exogenous TBP enriched at germinal vesicle at GV stage but disappeared from the chromosomes after GVBD. Moreover, exogenous TBP was still dispersed from the chromosomes of somatic donor nuclear in oocytes by nuclear transfer (NT), further proving that oocyte has some mechanism to remove TBP. During mouse embryo development, the exogenous TBP was removed from the chromosomes of M phase zygotes, but was found to express weakly at the M phase of 2-cell. Moreover, in the blastocyst TBP was also detected at the M phase chromosomes. Overexpression of TBP caused the failure of oocyte maturation and embryo development. Our results supported the idea that TBP might be a marker for transmitting cellular memory to daughter cells.

  3. Porcine Pluripotent Stem Cells Derived from IVF Embryos Contribute to Chimeric Development In Vivo.

    Directory of Open Access Journals (Sweden)

    Binghua Xue

    Full Text Available Although the pig is considered an important model of human disease and an ideal animal for the preclinical testing of cell transplantation, the utility of this model has been hampered by a lack of genuine porcine embryonic stem cells. Here, we derived a porcine pluripotent stem cell (pPSC line from day 5.5 blastocysts in a newly developed culture system based on MXV medium and a 5% oxygen atmosphere. The pPSCs had been passaged more than 75 times over two years, and the morphology of the colony was similar to that of human embryonic stem cells. Characterization and assessment showed that the pPSCs were alkaline phosphatase (AKP positive, possessed normal karyotypes and expressed classic pluripotent markers, including OCT4, SOX2 and NANOG. In vitro differentiation through embryonic body formation and in vivo differentiation via teratoma formation in nude mice demonstrated that the pPSCs could differentiate into cells of the three germ layers. The pPSCs transfected with fuw-DsRed (pPSC-FDs could be passaged with a stable expression of both DsRed and pluripotent markers. Notably, when pPSC-FDs were used as donor cells for somatic nuclear transfer, 11.52% of the reconstructed embryos developed into blastocysts, which was not significantly different from that of the reconstructed embryos derived from porcine embryonic fibroblasts. When pPSC-FDs were injected into day 4.5 blastocysts, they became involved in the in vitro embryonic development and contributed to the viscera of foetuses at day 50 of pregnancy as well as the developed placenta after the chimeric blastocysts were transferred into recipients. These findings indicated that the pPSCs were porcine pluripotent cells; that this would be a useful cell line for porcine genetic engineering and a valuable cell line for clarifying the molecular mechanism of pluripotency regulation in pigs.

  4. Porcine Pluripotent Stem Cells Derived from IVF Embryos Contribute to Chimeric Development In Vivo.

    Science.gov (United States)

    Xue, Binghua; Li, Yan; He, Yilong; Wei, Renyue; Sun, Ruizhen; Yin, Zhi; Bou, Gerelchimeg; Liu, Zhonghua

    2016-01-01

    Although the pig is considered an important model of human disease and an ideal animal for the preclinical testing of cell transplantation, the utility of this model has been hampered by a lack of genuine porcine embryonic stem cells. Here, we derived a porcine pluripotent stem cell (pPSC) line from day 5.5 blastocysts in a newly developed culture system based on MXV medium and a 5% oxygen atmosphere. The pPSCs had been passaged more than 75 times over two years, and the morphology of the colony was similar to that of human embryonic stem cells. Characterization and assessment showed that the pPSCs were alkaline phosphatase (AKP) positive, possessed normal karyotypes and expressed classic pluripotent markers, including OCT4, SOX2 and NANOG. In vitro differentiation through embryonic body formation and in vivo differentiation via teratoma formation in nude mice demonstrated that the pPSCs could differentiate into cells of the three germ layers. The pPSCs transfected with fuw-DsRed (pPSC-FDs) could be passaged with a stable expression of both DsRed and pluripotent markers. Notably, when pPSC-FDs were used as donor cells for somatic nuclear transfer, 11.52% of the reconstructed embryos developed into blastocysts, which was not significantly different from that of the reconstructed embryos derived from porcine embryonic fibroblasts. When pPSC-FDs were injected into day 4.5 blastocysts, they became involved in the in vitro embryonic development and contributed to the viscera of foetuses at day 50 of pregnancy as well as the developed placenta after the chimeric blastocysts were transferred into recipients. These findings indicated that the pPSCs were porcine pluripotent cells; that this would be a useful cell line for porcine genetic engineering and a valuable cell line for clarifying the molecular mechanism of pluripotency regulation in pigs.

  5. Arabidopsis phosphatidylinositol-phospholipase C2 (PLC2) is required for female gametogenesis and embryo development.

    Science.gov (United States)

    Di Fino, Luciano M; D'Ambrosio, Juan Martín; Tejos, Ricardo; van Wijk, Ringo; Lamattina, Lorenzo; Munnik, Teun; Pagnussat, Gabriela C; Laxalt, Ana M

    2017-04-01

    AtPLC2 is an essential gene in Arabidopsis, since it is required for female gametogenesis and embryo development. AtPLC2 might play a role in cell division during embryo-sac development and early embryogenesis. Phosphoinositide-specific phospholipase C (PI-PLC) plays an important role in signal transduction during plant development and in the response to various biotic- and abiotic stresses. The Arabidopsis PI-PLC gene family is composed of nine members, named PLC1 to PLC9. Here, we report that PLC2 is involved in female gametophyte development and early embryogenesis. Using two Arabidopsis allelic T-DNA insertion lines with different phenotypic penetrations, we observed both female gametophytic defects and aberrant embryos. For the plc2-1 mutant (Ws background), no homozygous plants could be recovered in the offspring from self-pollinated plants. Nonetheless, plc2-1 hemizygous mutants are affected in female gametogenesis, showing embryo sacs arrested at early developmental stages. Allelic hemizygous plc2-2 mutant plants (Col-0 background) present reduced seed set and embryos arrested at the pre-globular stage with abnormal patterns of cell division. A low proportion (0.8%) of plc2-2 homozygous mutants was found to escape lethality and showed morphological defects and disrupted megagametogenesis. PLC2-promoter activity was observed during early megagametogenesis, and after fertilization in the embryo proper. Immunolocalization studies in early stage embryos revealed that PLC2 is restricted to the plasma membrane. Altogether, these results establish a role for PLC2 in both reproductive- and embryo development, presumably by controlling mitosis and/or the formation of cell-division planes.

  6. In Vitro Maturation and Embryo Development to blastocyst Mouse Germinal Vesicle Oocytes after Vitrification

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    M Nikseresht

    2013-05-01

    Full Text Available Abstract Background & aim: Vitrification is a simple and ultra rapid technique for the conservation of fertility. Improving pregnancy rate associate with the use of cryopreserved oocytes would be an important advanced in human assisted reproductive technology (ART. The purpose of this study was to evaluate survival, oocytes maturation and embryo development to the blastocyst stage after vitrification of oocytes germinal vesicle-stage and multi stage Methods: In the present experimental study, germinal vesicle oocytes with or without cumulus cells were transferred to vitrification solution containing 30% (v/v ethylene glycol, 18% (w/v Ficoll-70, and 0.3 M sucrose, either by single step or in a step-wise way. After vitrification and storage in liquid nitrogen, the oocytes were thawed and washed twice in culture medium TCM119, and then subjected to in vitro maturation, fertilization, and culture. Data analysis was performed by using One-way variance and Tukey tests. Results: Oocytes survival, metaphase 2 stage oocyte maturation, fertilization and embryo formed blastocyst in vitrification methods multistage were significantly higher than the single step procedure (P<0/05 Conclusion: The Germinal vesicle stage oocytes vitrified with cumulus cells and stepwise procedure had positive effect on the survival, maturation and developmental rate on blastocyst compared to oocytes without cumulus cell and single step procedure. Key words: Germinal Vesicle Oocyte, Blastocyst, Vitrification, Ethylene glycol

  7. Preimplantation diagnosis of a human beta-globin transgene in biopsied trophectoderm cells and blastomeres of the mouse embryo.

    Science.gov (United States)

    Sheardown, S A; Findlay, I; Turner, A; Greaves, D; Bolton, V N; Mitchell, M; Layton, D M; Muggleton-Harris, A L

    1992-10-01

    The preimplantation diagnosis of a HbSA-globin transgene in biopsied trophectoderm cells and blastomeres in embryos using a transgenic mouse model for the trait of human sickle-cell anaemia has been undertaken. A sensitive procedure was developed for the amplification of the human beta-globin gene sequence flanking the sickle mutation. Polymerase chain reaction (PCR) assays were undertaken on one to five biopsied trophectoderm cells and isolated blastomeres of the preimplantation mouse embryo. After biopsy the blastocysts were cultured whilst the cells were analysed for the presence of the transgene, and a high proportion (82-91%) were viable as assessed by the presence of a blastocoele cavity within a 5-h period. The majority of the biopsied cultured blastocysts were frozen and used to confirm the diagnosis; 90 biopsied cultured blastocysts were transferred to pseudopregnant recipients and 34% established pregnancy. Material from day 13.5 post-coitum fetuses was also used to confirm the original diagnosis. The time (4-5 h) required to carry out the analysis obviates a need for extended culture or cryopreservation of the biopsied embryo. In individual experiments under optimal conditions, the presence of the transgene in biopsied cells was detected with 100% accuracy, and the PCR analysis was sensitive at the 1-cell level. The overall success rate of diagnosis and confirmation of the presence or absence of the human beta-globin sequence in the biopsied embryo was 70%. Over the entire experimental period (14 months) DNA contamination from a variety of sources did occasionally occur; the methods used to overcome this problem are discussed.

  8. Changes in ribosomal proteins in wheat embryos in the course of grain development and maturation

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    Stanisław Weidner

    2014-01-01

    Full Text Available It was found, by comparing the densitometric profiles of ribosomal proteins of wheat embryos in milk and full grain ripeness, that in the process of development and ripening of caryopses the percentual proportion of low molecular weight proteins increases at the cost of those of high molecular weight. This concerns both acidic and basic proteins. In electrophoretic separation of ribosomal proteins from embryos of fully ripe seeds by the method of two-dimensional electrophoresis the appearance of three new low molecular weight proteins - an acidic one and two basic ones - was observed. These proteins were not found in the embryos of caryopses of milk ripeness. These results indicate that with development and ripening of wheat caryopses new low molecular weight ribosomal proteins are built into the ribosomes in the embryo. These changes are both quantitative and qualitative.

  9. In vitro development of canine somatic cell nuclear transfer embryos in different culture media.

    Science.gov (United States)

    Kim, Dong-Hoon; No, Jin-Gu; Choi, Mi-Kyung; Yeom, Dong-Hyeon; Kim, Dong-Kyo; Yang, Byoung-Chul; Yoo, Jae Gyu; Kim, Min Kyu; Kim, Hong-Tea

    2015-01-01

    The objective of the present study was to investigate the effects of three different culture media on the development of canine somatic cell nuclear transfer (SCNT) embryos. Canine cloned embryos were cultured in modified synthetic oviductal fluid (mSOF), porcine zygote medium-3 (PZM-3), or G1/G2 sequential media. Our results showed that the G1/G2 media yielded significantly higher morula and blastocyst development in canine SCNT embryos (26.1% and 7.8%, respectively) compared to PZM-3 (8.5% and 0%or mSOF (2.3% and 0%) media. In conclusion, this study suggests that blastocysts can be produced more efficiently using G1/G2 media to culture canine SCNT embryos.

  10. Topography of the inferior alveolar nerve in human embryos and fetuses. An histomorphological study.

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    Sergey Lvovich Kabak

    2017-11-01

    Full Text Available The aim of this study is to establish the position of the inferior alveolar nerve in relation to the Meckel’s cartilage, the anlage of the mandibular body and primordia of the teeth, and also to trace the change in nerve trunk structure in the human prenatal ontogenesis. Serial sections (20µm from thirty-two 6-12 weeks-old entire human embryos and serial sections (10µm of six mandibles of 13-20 weeks-old human fetuses without developmental abnormalities were studied. Histological sections were impregnated with silver nitrate according to Bilshovsky-Buke and stained with hematoxylin and eosin. During embryonic development, the number of branches of the inferior alveolar nerve increases and its fascicular structure changes. In conclusion, the architecture of intraosseous canals in the body of the mandible, as well as the location of the foramina, is predetermined by the course and pattern of the vessel/nerve branching in the mandibular arch, even before the formation of bony trabeculae. Particularly, the formation of the incisive canal of the mandible can be explained by the presence of the incisive nerve as the extension of the inferior alveolar nerve. It has also been established that Meckel’s cartilage does not participate in mandibular canal morphogenesis.

  11. Regulation of somatic embryo development in Norway spruce (Picea abies). A molecular approach to the characterization of specific developmental stages

    Energy Technology Data Exchange (ETDEWEB)

    Sabala, I. [Swedish Univ. of Agricultural Sciences, Uppsala (Sweden). Dept. of Forest Genetics

    1998-12-31

    Embryo development is a complex process involving a set of strictly regulated events. The regulation of these events is poorly understood especially during the early stages of embryo development. Somatic embryos go through the same developmental stages as zygotic embryos making them an ideal model system for studying the regulation of embryo development. We have used embryogenic cultures of Picea abies to study some aspects of the regulation of embryo development in gymnosperms. The bottle neck during somatic embryogenesis is the switch from the proliferation stage to the maturation stage. This switch is initiated by giving somatic embryos a maturation treatment i.e. the embryos are treated with abscisic acid (ABA). Somatic embryos which respond to ABA by forming mature somatic embryos were stimulated to secret a 70 kDa protein, AF70. The af70 gene was isolated and characterised. The expression of the af70 gene was constitutive in embryos but was highly ABA-induced in seedlings. Moreover, expression of this gene was stimulated during cold acclimation of Picea abies seedlings. A full length Picea abies cDNA clone Pa18, encoding a protein with the characteristics of plant lipid transfer proteins (LTPs), was isolated and characterised. The Pa18 gene is constitutively expressed in embryogenic cultures of Picea abies representing different stages of development as well as in nonembryogenic callus and seedlings. In situ hybridization showed that Pa18 gene is expressed in all embryonic cells of proliferating somatic embryos but the expression of the gene in mature somatic and zygotic embryos is restricted to the outer cell layer. Southern blot analysis at different stringencies was consistent with a single gene. An alteration in expression of Pa18 causes disturbance in the formation of the proper outer cell layer in the maturing somatic embryos. In addition to its influence on embryo development the Pa18 gene product also inhibits growth of Agrobacterium tumefaciens 195

  12. The effects of genetic line (broilers vs. layers) on embryo development.

    Science.gov (United States)

    Druyan, S

    2010-07-01

    Recent decades were characterized by genetic selection of broiler and layer chickens for enhanced growth rate and meat yield or intensified egg production, respectively. It is to be expected that genetic selection for various traits would also influence embryo development and growth patterns that affect metabolism. The objective of the present study was to examine the effects of broiler (Cobb and Ross) and layer (Lohmann) lines and parent flock age (31 and 38 wk) on embryonic development, heart rate, O2 consumption, and blood parameters. For each line, 2 incubation sets, from flocks aged 31 and 38 wk, with 500 eggs per set, were studied. Development patterns differed between layers and broilers: layers hatched 1 d later and their relative embryonic weight at hatch was significantly lower, probably because of their longer period until hatch, although yolk relative weights were similar. Oxygen consumption of layer embryos was lower than that of broilers, and plasma triiodothyronine concentration, hematocrit, and hemoglobin levels were lower in layers than in broilers. However, layer embryo heart rate was higher from embryonic d (E) 15 onward. Differences were found between the Ross and Cobb lines in embryonic development. Oxygen consumption of Ross embryos was slightly higher than that of Cobb from E16 to E19. Heart rate of Ross embryos was significantly higher than that of Cobb. Furthermore, plasma triiodothyronine concentration of Ross embryos was significantly higher on E14, E16, and hatch. These differences suggest that the genetic selection for rapid growth rate in the 2 broiler lines did not cause differences between their embryonic growth patterns, but it did affect their metabolic rate. Oxygen consumption was higher in embryos from the 38-wk-old flock. The results suggest that genetic selection affected not only production traits but also the developmental pattern of the embryo and its metabolic characteristics.

  13. Effects of cigarette smoke residues from textiles on fibroblasts, neurocytes and zebrafish embryos and nicotine permeation through human skin.

    Science.gov (United States)

    Hammer, Timo R; Fischer, Kirsten; Mueller, Marina; Hoefer, Dirk

    2011-09-01

    Toxic substances from cigarette smoke can attach to carpets, curtains, clothes or other surfaces and thus may pose risks to affected persons. The phenomenon itself and the potential hazards are discussed controversially, but scientific data are rare. The objective of this study was to examine the potential of textile-bound nicotine for permeation through human skin and to assess the effects of cigarette smoke extracts from clothes on fibroblasts, neurocytes and zebrafish embryos. Tritiated nicotine from contaminated cotton textiles penetrated through adult human full-thickness skin as well as through a 3D in vitro skin model in diffusion chambers. We also observed a significant concentration-dependent cytotoxicity of textile smoke extracts on fibroblast viability and structure as well as on neurocytes. Early larval tests with zebrafish embryos were used as a valid assay for testing acute vertebrate toxicity. Zebrafish development was delayed and most of the embryos died when exposed to smoke extracts from textiles. Our data show that textiles contaminated with cigarette smoke represent a potential source of nicotine uptake and can provoke adverse health effects. Copyright © 2011 Elsevier GmbH. All rights reserved.

  14. Cryopreservation of human oocytes, zygotes, embryos and blastocysts: A comparison study between slow freezing and ultra rapid (vitrification methods

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    Tahani Al-Azawi

    2013-12-01

    Full Text Available Preservation of female genetics is currently done primarily by means of oocyte and embryo cryopreservation. The field has seen much progress during its four-decade history, progress driven predominantly by research in humans. It can also be done by preservation of ovarian tissue or entire ovary for transplantation, followed by oocyte harvesting or natural fertilization. Two basic cryopreservation techniques rule the field, slow-rate freezing, the first to be developed and vitrification which in recent years, has gained a foothold. The slow-rate freezing method previously reported had low survival and pregnancy rates, along with the high cost of cryopreservation. Although there are some recent data indicating better survival rates, cryopreservation by the slow freezing method has started to discontinue. Vitrification of human embryos, especially at early stages, became a more popular alternative to the slow rate freezing method due to reported comparable clinical and laboratory outcomes. In addition, vitrification is relatively simple, requires no expensive programmable freezing equipment, and uses a small amount of liquid nitrogen for freezing. Moreover, oocyte cryopreservation using vitrification has been proposed as a solution to maintain women’s fertility by serving and freezing their oocytes at the optimal time. The aim of this research is to compare slow freezing and vitrification in cryopreservation of oocytes, zygotes, embryos and blastocysts during the last twelve years. Therefore, due to a lot of controversies in this regard, we tried to achieve an exact idea about the subject and the best technique used.

  15. Rethinking In Vitro Embryo Culture: New Developments in Culture Platforms and Potential to Improve Assisted Reproductive Technologies1

    Science.gov (United States)

    Smith, Gary D.; Takayama, Shuichi; Swain, Jason E.

    2011-01-01

    ABSTRACT The preponderance of research toward improving embryo development in vitro has focused on manipulation of the chemical soluble environment, including altering basic salt composition, energy substrate concentration, amino acid makeup, and the effect of various growth factors or addition or subtraction of other supplements. In contrast, relatively little work has been done examining the physical requirements of preimplantation embryos and the role culture platforms or devices can play in influencing embryo development within the laboratory. The goal of this review is not to reevaluate the soluble composition of past and current embryo culture media, but rather to consider how other controlled and precise factors such as time, space, mechanical interactions, gradient diffusions, cell movement, and surface interactions might influence embryo development. Novel culture platforms are being developed as a result of interdisciplinary collaborations between biologists and biomedical, material, chemical, and mechanical engineers. These approaches are looking beyond the soluble media composition and examining issues such as media volume and embryo spacing. Furthermore, methods that permit precise and regulated dynamic embryo culture with fluid flow and embryo movement are now available, and novel culture surfaces are being developed and tested. While several factors remain to be investigated to optimize the efficiency of embryo production, manipulation of the embryo culture microenvironment through novel devices and platforms may offer a pathway toward improving embryo development within the laboratory of the future. PMID:21998170

  16. Rethinking in vitro embryo culture: new developments in culture platforms and potential to improve assisted reproductive technologies.

    Science.gov (United States)

    Smith, Gary D; Takayama, Shuichi; Swain, Jason E

    2012-03-01

    The preponderance of research toward improving embryo development in vitro has focused on manipulation of the chemical soluble environment, including altering basic salt composition, energy substrate concentration, amino acid makeup, and the effect of various growth factors or addition or subtraction of other supplements. In contrast, relatively little work has been done examining the physical requirements of preimplantation embryos and the role culture platforms or devices can play in influencing embryo development within the laboratory. The goal of this review is not to reevaluate the soluble composition of past and current embryo culture media, but rather to consider how other controlled and precise factors such as time, space, mechanical interactions, gradient diffusions, cell movement, and surface interactions might influence embryo development. Novel culture platforms are being developed as a result of interdisciplinary collaborations between biologists and biomedical, material, chemical, and mechanical engineers. These approaches are looking beyond the soluble media composition and examining issues such as media volume and embryo spacing. Furthermore, methods that permit precise and regulated dynamic embryo culture with fluid flow and embryo movement are now available, and novel culture surfaces are being developed and tested. While several factors remain to be investigated to optimize the efficiency of embryo production, manipulation of the embryo culture microenvironment through novel devices and platforms may offer a pathway toward improving embryo development within the laboratory of the future.

  17. The Well of the Well (WOW) system: an efficient approach to improve embryo development

    DEFF Research Database (Denmark)

    Vajta, G; Korösi, T; Du, Y

    2008-01-01

    Transfer of human embryos at the blastocyst stage may offer considerable benefits including the increased implantation rates and decreased risks of multiple pregnancies, however, it requires an efficient and reliable in vitro embryo culture system. In our study, the effect of the Well of the Well...... (WOW) system consisting of microwells formed on the bottom of the culture dish was tested in three mammalian species including humans. The WOW system has resulted in significant improvement compared the drops for culture of in vitro matured and parthenogenetically activated porcine oocytes or in vivo...

  18. Studying early lethality of 45,XO (Turner's syndrome embryos using human embryonic stem cells.

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    Achia Urbach

    Full Text Available Turner's syndrome (caused by monosomy of chromosome X is one of the most common chromosomal abnormalities in females. Although 3% of all pregnancies start with XO embryos, 99% of these pregnancies terminate spontaneously during the first trimester. The common genetic explanation for the early lethality of monosomy X embryos, as well as the phenotype of surviving individuals is haploinsufficiency of pseudoautosomal genes on the X chromosome. Another possible mechanism is null expression of imprinted genes on the X chromosome due to the loss of the expressed allele. In contrast to humans, XO mice are viable, and fertile. Thus, neither cells from patients nor mouse models can be used in order to study the cause of early lethality in XO embryos. Human embryonic stem cells (HESCs can differentiate in culture into cells from the three embryonic germ layers as well as into extraembryonic cells. These cells have been shown to have great value in modeling human developmental genetic disorders. In order to study the reasons for the early lethality of 45,XO embryos we have isolated HESCs that have spontaneously lost one of their sex chromosomes. To examine the possibility that imprinted genes on the X chromosome play a role in the phenotype of XO embryos, we have identified genes that were no longer expressed in the mutant cells. None of these genes showed a monoallelic expression in XX cells, implying that imprinting is not playing a major role in the phenotype of XO embryos. To suggest an explanation for the embryonic lethality caused by monosomy X, we have differentiated the XO HESCs in vitro an in vivo. DNA microarray analysis of the differentiated cells enabled us to compare the expression of tissue specific genes in XO and XX cells. The tissue that showed the most significant differences between the clones was the placenta. Many placental genes are expressed at much higher levels in XX cells in compare to XO cells. Thus, we suggest that abnormal

  19. Onset of buccal pumping in catshark embryos: how breathing develops in the egg capsule.

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    Taketeru Tomita

    Full Text Available Respiration in fishes involves buccal pumping, which is characterized by the generation of nearly continuous water flow over the gills because of the rhythmic expansion/compression of the pharyngeal cavity. This mechanism is achieved by the functions of the vascular, skeletal, and muscular systems. However, the process by which the embryo establishes the mechanism remains a mystery. Morphological and kinematical observations on captive cloudy catsharks, Scyliorhinus torazame, have suggested that the embryo starts buccal pumping just before the respiratory slits open on the egg capsule. During the pre-opening period, the embryo acquires oxygen mainly via the external gill filaments. After slit opening, respiration of the embryo involves buccal pumping to pass water over the "internal gills." The onset of buccal pumping accompanies four morphological changes: (1 regression of the external gill filaments, (2 development of blood vessels within the "internal gills," (3 completion of the development of hyoid skeletal and muscular elements, and (4 development of the oral valve. A previous study showed that buccal pumping allows the embryo to actively regulate oxygen intake by changing the pumping frequency. Thus, establishment of buccal pumping in the egg capsule is probably important for embryo survival in the unstable oxygen environment of the egg capsule after slit opening.

  20. Effects of the popular food additive sodium benzoate on neural tube development in the chicken embryo.

    Science.gov (United States)

    Emon, Selin Tural; Orakdogen, Metin; Uslu, Serap; Somay, Hakan

    2015-01-01

    Many more additives have been introduced with the development of processed foods. Neural tube defects are congenital malformations of the central nervous system. More than 300 000 children are born with neural tube defects every year and surviving children remain disabled for life. Sodium benzoate is used intensively in our daily lives. We therefore aimed to evaluate the effects of sodium benzoate on neural tube defects in chicken embryos. Fertile, specific pathogen-free eggs were used. The study was conducted on five groups. After 30 hours of incubation, the eggs were opened under 4x optical magnification. The embryonic disc was identified and sodium benzoate solution was injected. Eggs were closed with sterile adhesive strips and incubation was continued till the end of the 72nd hour. All eggs were then reopened and embryos were dissected from embryonic membranes and evaluated histopathologically. We found that the development of all embryos was consistent with the stage. We detected neural tube obstruction in one embryo. Neural tube defects were not detected in any embryos. This study showed that sodium benzoate as one of the widely used food preservatives has no effect to neural tube defect development in chicken embryos even at high doses.

  1. The Effect of Prolonged Culture of Chromosomally Abnormal Human Embryos on The Rate of Diploid Cells

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    Masood Bazrgar

    2016-12-01

    Full Text Available Background: A decrease in aneuploidy rate following a prolonged co-culture of human blastocysts has been reported. As co-culture is not routinely used in assisted reproductive technology, the present study aimed to evaluate the effect of the prolonged single culture on the rate of diploid cells in human embryos with aneuploidies. Materials and Methods: In this cohort study, we used fluorescence in situ hybridization (FISH to reanalyze surplus blastocysts undergoing preimplantation genetic diagnosis (PGD on day 3 postfertilization. They were randomly studied on days 6 or 7 following fertilization. Results: Of the 30 analyzed blastocysts, mosaicism was observed in 26(86.6%, while 2(6.7% were diploid, and 2(6.7% were triploid. Of those with mosaicism, 23(88.5% were determined to be diploid-aneuploid and 3(11.5% were aneuploid mosaic. The total frequency of embryos with more than 50% diploid cells was 33.3% that was lower on day 7 in comparison with the related value on day 6 (P<0.05; however, there were no differences when the embryos were classified according to maternal age, blastocyst developmental stage, total cell number on day 3, and embryo quality. Conclusion: Although mosaicism is frequently observed in blastocysts, the prolonged single culture of blastocysts does not seem to increase the rate of normal cells.

  2. The endo-siRNA pathway is essential for robust development of the Drosophila embryo.

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    Elena M Lucchetta

    2009-10-01

    Full Text Available Robustness to natural temperature fluctuations is critical to proper development in embryos and to cellular functions in adult organisms. However, mechanisms and pathways which govern temperature compensation remain largely unknown beyond circadian rhythms. Pathways which ensure robustness against temperature fluctuations may appear to be nonessential under favorable, uniform environmental conditions used in conventional laboratory experiments where there is little variation for which to compensate. The endo-siRNA pathway, which produces small double-stranded RNAs in Drosophila, appears to be nonessential for robust development of the embryo under ambient uniform temperature and to be necessary only for viral defense. Embryos lacking a functional endo-siRNA pathway develop into phenotypically normal adults. However, we hypothesized that small RNAs may regulate the embryo's response to temperature, as a ribonucleoprotein complex has been previously shown to mediate mammalian cell response to heat shock.Here, we show that the genes DICER-2 and ARGONAUTE2, which code for integral protein components of the endo-siRNA pathway, are essential for robust development and temperature compensation in the Drosophila embryo when exposed to temperature perturbations. The regulatory functions of DICER-2 and ARGONAUTE2 were uncovered by using microfluidics to expose developing Drosophila embryos to a temperature step, in which each half of the embryo develops at a different temperature through developmental cycle 14. Under this temperature perturbation, dicer-2 or argonaute2 embryos displayed abnormal segmentation. The abnormalities in segmentation are presumably due to the inability of the embryo to compensate for temperature-induced differences in rate of development and to coordinate developmental timing in the anterior and posterior halves. A deregulation of the length of nuclear division cycles 10-14 is also observed in dicer-2 embryos at high temperatures

  3. Gluconeogenesis, non-essential amino acid synthesis and substrate partitioning in chicken embryos during later development.

    Science.gov (United States)

    Hu, Q; Agarwal, U; Bequette, B J

    2017-02-01

    We aimed to quantify the rate of gluconeogenesis (GNG), non-essential amino-acid (NEAA) synthesis, and substrate partitioning to the Krebs cycle in embryonic (e) day e14 and e19 chicken embryos. An in ovo continuous tracer infusion approach was employed to test the hypotheses that GNG and NEAA synthesis in developing chicken embryo increases from e14 to e19. [ 13 C 6 ]Glucose or [ 13 C 3 ]glycerol was continuously infused (8 h) into the chorio-allantoic compartment of eggs on e14 and e19. Glucose entry rate, Cori cycling, and GNG were higher (P < 0.05) in e19 compared to e14 embryos, presumably to support higher glycogen deposition in liver and muscle. Whereas de novo synthesis of alanine, aspartate, and glutamate via glycolysis and the Krebs cycle was higher (P < 0.01) in e14 embryos, synthesis of these NEAA from glycerol was higher (P < 0.05) in e19 compared to e14 embryos. These patterns of glucose and glycerol utilization suggest a metabolic shift to conserve glucose for glycogen synthesis and an increased utilization of yolk glycerol (from triacylglyceride) after e14. Although the contribution of glycerol to GNG in e19 embryos was higher (P < 0.05) than that in e14 embryos, the contribution of glycerol to GNG (1.3 to 6.0%) was minor. Based on [ 13 C 6 ]glucose tracer kinetics, the activities of both pyruvate carboxylase (PC) and pyruvate dehydrogenase (PDH) in the liver were higher (P < 0.05) in e19 embryos; whereas the higher (P < 0.01) relative activity of liver PC compared to PDH in e14 embryos suggests a greater anaplerotic flux into the Krebs cycle. In summary, the in ovo continuous tracer infusion approach allowed for a measurement of chicken embryo whole body and liver metabolism over a shorter window of development. This study provided quantitative estimates of the developmental shifts in substrate utilization, GNG, and NEAA synthesis by chicken embryos, as well as qualitative estimates of the activities of enzymes central to the Krebs cycle

  4. Cryopreservation of canine embryos.

    Science.gov (United States)

    Abe, Yasuyuki; Suwa, Yoshinori; Asano, Tomoyoshi; Ueta, Yoshiko Yanagimoto; Kobayashi, Nanae; Ohshima, Natsumi; Shirasuna, Saori; Abdel-Ghani, Mohammed Ali; Oi, Maya; Kobayashi, Yoshiyasu; Miyoshi, Masafumi; Miyahara, Kazuro; Suzuki, Hiroshi

    2011-02-01

    The assisted reproductive techniques (ARTs) such as in vitro fertilization, embryo transfer, and cryopreservation of gametes have contributed considerably to the development of biomedical sciences in addition to improving infertility treatments in humans as well as the breeding of domestic animals. However, ARTs used in canine species have strictly limited utility when compared with other mammalian species, including humans. Although successful somatic cell cloning has been reported, artificial insemination by frozen semen to date is only available for the improved breeding and reproduction for companion and working dogs as well as guide dogs for the blind. We describe here the successful cryopreservation of embryos and subsequent embryo transfer in dogs. Canine embryos were collected from excised reproductive organs after artificial insemination and subsequently cryopreserved by a vitrification method. When the 4-cell to morula stage of cryopreserved embryos were nonsurgically transferred into the uteri of nine recipient bitches using a cystoscope, five recipients became pregnant and four of them delivered a total of seven pups. The cryopreservation of embryos in canine species will facilitate the transportation and storage of genetic materials and will aid in the elimination of vertically transmitted diseases in dogs. In addition, this technique will contribute to the improved breeding of companion and working dogs such as guide dogs, drug-detecting dogs, and quarantine dogs.

  5. Magnetic resonance imaging of iron-oxide labeled SK-Mel 28 human melanoma cells in the chick embryo using a clinical whole body MRI scanner.

    Science.gov (United States)

    Oppitz, M; Pintaske, J; Kehlbach, R; Schick, F; Schriek, G; Busch, C

    2007-02-01

    To evaluate advantages and limitations of magnetic resonance imaging (MRI) to monitor the migration of superparamagnetic iron oxide (SPIO) labeled cells in the chick embryo. Labeled human SK-Mel 28 melanoma cells were injected into the E2 chick embryo neural tube. Embryos were examined with a clinical 3 T MRI whole body system using 3D T*(2)-weighted sequences with isotropic spatial resolutions of 0.3-1.0 mm. MR-measurements of embryos were performed 2 - 16 days after cell injection. MRI findings were verified by dissection and histology. After injection, melanoma cells formed aggregations that were detectable in the neural tube as signal voids in MR images from day 2 after injection. Emigrating cells later left MRI detectable tracks. Aggregates that remained in the neural tube left label that was absorbed by glia cells. In E18 chick embryos, signals of haematopoiesis interfered with signals from cell labeling. It was shown that SK-Mel 28 cells will resume the neural crest pathways after injection into the embryonic micro-environment. SPIO cell labeling allows monitoring of transplanted melanoma cells during embryonic development. MRI using the standard clinical equipment promises to be valuable for high-sensitive monitoring of ex-vivo labeled cells in the chick embryo.

  6. The Effects of Ethanol and Strontium on Growth and Development of Two-Cell Arrested Mouse Embryos

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Darabi

    2012-01-01

    Full Text Available Background: Arresting at a certain stage of development like the two-cell stage could be one of the causes of infertility. The aim of this study is to evaluate the effects of ethanol and strontium on growth and development of mice embryos arrested at the two-cell stage.Materials and Methods: In this experimental study, female mice were coupled with a male following superovulation. Positive vaginal plug mice were sacrificed 48 hours after human chorionic gonadotropin (hCG injection. Two-cell embryos were transferred to M16 medium and divided to four groups. The first control group was incubated without any exposure to low temperatures. Groups 2, 3 and 4 were exposed to 4°C for 24 hours. The second control group was incubated immediately, while the third and fourth groups were exposed to 10 mM strontium for five minutes and 0.1% ethanol for a further five minutes. Growth rate and developmental parameters of embryos were analyzed by one-way ANOVA. The significant difference between the groups was determined by Post Hoc.Results: The data shows that developmental rate is decreased significantly by 4°C exposure. The mean percentage of degenerated embryo was significantly different between groups but the mean cleavage rate was not significantly different. The mean percent of morula, blastocyst and hatched blastocyst formation were significantly different between groups during a 120 hours study post hCG injection.Conclusion: The effect of strontium and ethanol on arrested two-cell embryos had no significant effect on the mean percentage of morula, but ethanol treatment significantly increased the percentage of blastocyst and hatched blastocyst formation compared to strontium.

  7. Effects of steroidal glycoalkaloids from potatoes (Solanum tuberosum) on in vitro bovine embryo development.

    Science.gov (United States)

    Wang, S; Panter, K E; Gaffield, W; Evans, R C; Bunch, T D

    2005-02-01

    alpha-Solanine and alpha-chaconine are two naturally occurring steroidal glycoalkaloids in potatoes (Solanum tuberosum), and solanidine-N-oxide is a corresponding steroidal aglycone. The objective of this research was to screen potential cyto-toxicity of these potato glycoalkaloids using bovine oocyte maturation, in vitro fertilization techniques and subsequent embryonic development as the in vitro model. A randomized complete block design with four in vitro oocyte maturation (IVM) treatments (Experiment 1) and four in vitro embryo culture (IVC) treatments (Experiment 2) was used. In Experiment 1, bovine oocytes (n=2506) were matured in vitro in medium supplemented with 6 microM of alpha-solanine, alpha-chaconine, solanidine-N-oxide or IVM medium only. The in vitro matured oocytes were then subject to routine IVF and IVC procedures. Results indicated that exposure of bovine oocytes to the steroidal glycoalkaloids during in vitro maturation inhibited subsequent pre-implantation embryo development. Potency of the embryo-toxicity varied between these steroidal glycoalkaloids. In Experiment 2, IVM/IVF derived bovine embryos (n=2370) were cultured in vitro in medium supplemented with 6 microM of alpha-solanine, alpha-chaconine, solanidine-N-oxide or IVC medium only. The results showed that the pre-implantation embryo development is inhibited by exposure to these glycoalkaloids. This effect is significant during the later pre-implantation embryo development period as indicated by fewer numbers of expanded and hatched blastocysts produced in the media containing these alkaloids. Therefore, we conclude that in vitro exposure of oocytes and fertilized ova to the steroidal glycoalkaloids from potatoes inhibits pre-implantation embryo development. Furthermore, we suggest that ingestion of Solanum species containing toxic amounts of glycoalkaloids may have negative effects on pre-implantation embryonic survival.

  8. Discarded human spermatozoa, eggs and embryos for personnel training and practice in assisted reproduction.

    Science.gov (United States)

    Heng, Boon Chin

    2007-12-01

    An ethical issue that has been largely overlooked is the use of discarded human gametes and embryos for personnel training in clinical assisted reproduction technology, e.g. intracytoplasmic sperm injection and preimplantation genetic diagnosis. Unlike experimental research for generating peer-reviewed journal publications and intellectual property, there is no similar paper trail or smoking gun if human gametes and embryos are utilized solely for personnel training without the patient's knowledge and consent. For many assisted reproduction laboratories in private practice that are not affiliated with research or academic institutions, there are often difficulties in procuring animal material for personnel training. In contrast, discarded human gametes and embryos are readily available and can be convenient for training inexperienced personnel in assisted reproduction techniques. Very often, only verbal consent is obtained from patients, without written documentation, and this situation can potentially lead to abuse. For example, fertility clinics and laboratories may conduct training courses and workshops for generating additional income and revenue; and there is a possibility of discarded human material being utilized for such profit-making ventures without patients' prior knowledge. Hence, it is the moral duty and obligation of international professional bodies to advocate and draft clearly defined regulatory guidelines and legislative framework for this purpose.

  9. Sildenafil citrate (Viagra) impairs fertilization and early embryo development in mice.

    Science.gov (United States)

    Glenn, David R J; McClure, Neil; Cosby, S Louise; Stevenson, Michael; Lewis, Sheena E M

    2009-03-01

    To determine the effects of sildenafil citrate, a cyclic monophosphate-specific type 5 phosphodiesterase inhibitor known to affect sperm function, on fertilization and early embryo cleavage. This acute mammal study included male and female mice assigned randomly, the females sacrificed after mating and their oocytes/embryos evaluated at four time periods after treatment. Academic research environment. Male and female CBAB(6) mice. Female mice were injected intraperitoneally with 5 IU gonadotropin (hCG) to stimulate follicular growth and induce ovulation. They were each caged with a male that had been gavaged with sildenafil citrate (0.06 mg/0.05 mL) and allowed to mate. After 12, 36, 60, and 84 h, females were killed, their oviducts were dissected out, and retrieved embryos were assessed for blastomere number and quality. Fertilization rates and numbers of embryos were evaluated after treatment. Fertilization rates (day 1) were markedly reduced (-33%) in matings where the male had taken sildenafil citrate. Over days 2-4, the numbers of embryos developing in the treated group were significantly fewer than in the control group. There was also a trend for impaired cleavage rates within those embryos, although this did not reach significance. The impairments to fertility caused by sildenafil citrate have important implications for infertility centers and for couples who are using this drug precoitally while attempting to conceive.

  10. MORPHOLOGICAL CHANGES DURING THE DEVELOPMENT OF SOMATIC EMBRYOS OF SAGO (Metroxylon sagu Rottb.

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    Pauline D. Kasi

    2016-10-01

    Full Text Available Development of somatic embryos of sago (Metroxylon sagu Rottb. on agar-solidified medium are highly varied producing heterogeneous seedlings. Understanding of this phenomenon may help in improving the cultural procedures and conditions of sagosomatic embryogenesis to obtain uniform seedlings in a large scale. This experiment was conducted at the laboratory for plant cell culture and micropropagation, Indonesian Biotechnology Research Institute for Estate Crops from January to March 2006 to examine morphological changes i.e. color and development stages of sago during their somatic embryo development on an agar-solidified medium. Twenty single globular somatic embryos of sago with specific color (yellowish, greenish, and reddish were cultured in a Petri dish supplemented with a solid medium. The medium was a micronutrients-modified MS (MMS with half strength of macronutrients containing 0.01 mg l-1 ABA, 2 mg l-1 kinetin, 20 g l-1 sucrose, 0.5 g l-1 activated charcoal, and 2 g l-1 gelrite. Parameter observed was the percentage of embryo’s number based on color and developmental stage. The result showed that at the end of 6-week culture passage, most originally greenish (80.8% and reddish (95.8% embryos remained unchanged in their colors, whereas almost half of the originally yellowish embryos turned to greenish and only 30%remained yellowish. At the same time, single globular embryos have changed gradually into the next developmental stages, although not all of the embryos were germinated. The initial color of embryo affected the rate of the developmental stage changes. Yellowish and greenish globular embryos developed more rapidly into cotyledon or germinant stages at 58% and 55% respectively, in 6 weeks than the reddish ones (41%. Therefore, the yellowish and greenish embryos are the best sources of material for in vitro mass propagation and synthetic seed production of sago.

  11. Effect of Adding Human Chorionic Gonadotropin to The Endometrial Preparation Protocol in Frozen Embryo Transfer Cycles

    Directory of Open Access Journals (Sweden)

    Maryam Eftekhar

    2012-01-01

    Full Text Available Background: Human chorionic gonadotropin (HCG, one of the initial embryonic signals, isprobably a major regulator of the embryo-endometrial relationship. This study aims to assess theadvantage of HCG supplementation during the secretory phase of hormonally prepared cycles forthe transfer of cryopreserved-thawed embryos.Materials and Methods: This study was a randomized clinical trial. Infertile women who werecandidates for frozen-thawed embryo transfers entered the study and were divided into two groups,HCG and control. The endometrial preparation method was similar in both groups: all women receivedestradiol valerate (6 mg po per day from the second day of the menstrual cycle and progesteronein oil (100 mg intramuscular (I.M. when the endometrial thickness reached 8 mm. Estradiol andprogesterone were continued until the tenth week of gestation. In the HCG group, patients received anHCG 5000 IU injection on the first day of progesterone administration and the day of embryo transfer.Results: In this study, 130 couples participated: 65 in the HCG group and 65 in the control group.There was no statistically significant difference between groups regarding basic characteristics.Implantation rate, chemical pregnancy, clinical pregnancy, ongoing pregnancy, and abortion rateswere similar in both groups.Conclusion: Although HCG has some advantages in assisted reproductive technology (ARTcycles, our study did not show any benefit of HCG supplementation during the secretory phase offrozen cycles (Registration Number: IRCT201107266420N4.

  12. Does Embryo Culture Medium Influence the Health and Development of Children Born after In Vitro Fertilization?

    Science.gov (United States)

    Bouillon, Céline; Léandri, Roger; Desch, Laurent; Ernst, Alexandra; Bruno, Céline; Cerf, Charline; Chiron, Alexandra; Souchay, Céline; Burguet, Antoine; Jimenez, Clément; Sagot, Paul; Fauque, Patricia

    2016-01-01

    In animal studies, extensive data revealed the influence of culture medium on embryonic development, foetal growth and the behaviour of offspring. However, this impact has never been investigated in humans. For the first time, we investigated in depth the effects of embryo culture media on health, growth and development of infants conceived by In Vitro Fertilization until the age of 5 years old. This single-centre cohort study was based on an earlier randomized study. During six months, in vitro fertilization attempts (No. 371) were randomized according to two media (Single Step Medium--SSM group) or Global medium (Global group). This randomized study was stopped prematurely as significantly lower pregnancy and implantation rates were observed in the SSM group. Singletons (No. 73) conceived in the randomized study were included (42 for Global and 31 for SSM). The medical data for gestational, neonatal and early childhood periods were extracted from medical records and parental interviews (256 variables recorded). The developmental profiles of the children in eight domains (social, self-help, gross motor, fine motor, expressive language, language comprehension, letter knowledge and number knowledge--270 items) were compared in relation to the culture medium. The delivery rate was significantly lower in the SSM group than in the Global group (pculture medium had no significant effect on birthweight, risk of malformation (minor and major), growth and the frequency of medical concerns. However, the children of the Global group were less likely than those of the SSM group to show developmental problems (p = 0.002), irrespective of the different domains. In conclusion, our findings showed that the embryo culture medium may have an impact on further development.

  13. Does Embryo Culture Medium Influence the Health and Development of Children Born after In Vitro Fertilization?

    Directory of Open Access Journals (Sweden)

    Céline Bouillon

    Full Text Available In animal studies, extensive data revealed the influence of culture medium on embryonic development, foetal growth and the behaviour of offspring. However, this impact has never been investigated in humans. For the first time, we investigated in depth the effects of embryo culture media on health, growth and development of infants conceived by In Vitro Fertilization until the age of 5 years old. This single-centre cohort study was based on an earlier randomized study. During six months, in vitro fertilization attempts (No. 371 were randomized according to two media (Single Step Medium--SSM group or Global medium (Global group. This randomized study was stopped prematurely as significantly lower pregnancy and implantation rates were observed in the SSM group. Singletons (No. 73 conceived in the randomized study were included (42 for Global and 31 for SSM. The medical data for gestational, neonatal and early childhood periods were extracted from medical records and parental interviews (256 variables recorded. The developmental profiles of the children in eight domains (social, self-help, gross motor, fine motor, expressive language, language comprehension, letter knowledge and number knowledge--270 items were compared in relation to the culture medium. The delivery rate was significantly lower in the SSM group than in the Global group (p<0.05. The culture medium had no significant effect on birthweight, risk of malformation (minor and major, growth and the frequency of medical concerns. However, the children of the Global group were less likely than those of the SSM group to show developmental problems (p = 0.002, irrespective of the different domains. In conclusion, our findings showed that the embryo culture medium may have an impact on further development.

  14. Optimized protocol for cryopreservation of human eggs improves developmental competence and implantation of resulting embryos

    Directory of Open Access Journals (Sweden)

    Wang Cassie T

    2013-02-01

    Full Text Available Abstract Background Successful egg cryopreservation has many potential benefits to a variety of patients. However, a superior standard protocol describing all aspects of oocyte cryopreservation has not yet been identified. Oocyte cryopreservation is still a technical challenge for many infertility clinics. To maintain satisfactory clinical outcomes, there is a need to develop an easy to use, yet efficient laboratory protocol. The present study was designed to examine if human embryos resulting from eggs frozen with an optimized vitrification protocol have similar developmental competence as those from fresh eggs. Methods Twenty recipients received donated eggs vitrified with a protocol in which short exposure time to the vitrification solution was used and 23 recipients received donated eggs and 6 patients had their own eggs vitrified with a modified protocol in which long exposure time to the vitrification solution was used. After warming, egg survival, fertilization, cleavage, blastocyst formation, clinical pregnancy and implantation rates were compared. The developmental competence of eggs vitrified with the optimized protocol was further compared with fresh eggs donated from the same donors. Results There was no difference in the oocyte survival, fertilization, cleavage, clinical pregnancy or implantation rates between the short and long protocol groups. However, blastocyst formation rate was significantly (P Conclusions These results indicate that blastocyst development is an appropriate measure for egg survival after cryopreservation and frozen eggs have similar developmental potential as fresh eggs if they are frozen with an optimized method.

  15. Lipid analysis of developing Camelina sativa seeds and cultured embryos.

    Science.gov (United States)

    Pollard, Mike; Martin, Tina M; Shachar-Hill, Yair

    2015-10-01

    Camelina sativa is a cultivated oilseed rich in triacylglycerols containing oleic, linoleic, α-linolenic and eicosenoic acids. As it holds promise as a model species, its lipid synthesis was characterized in vivo and in culture. Lipid accumulates at a maximum rate of about 26 μg/day/seed (11.5 mg lipid/day/g fresh seed weight), a rate comparable with other oilseeds. Noteworthy is a late stage surge in α-linolenic acid accumulation. Small amounts of unusual epoxy and hydroxy fatty acids are also present in the triacylglycerols. These include 15,16-epoxy- and 15-hydroxy-octadecadienoic acids and homologous series of ω7-hydroxy-alk-ω9-enoic and ω9/10-hydroxy-alkanoic acids. Mid-maturation embryos cultured in vitro have growth and lipid deposition rates and fatty acid compositions that closely match that of seeds, but extended culture periods allow these rates to rise and surpass those observed in planta. Optimized thin layer chromatography systems for characterization of labeled products from acetate or glycerol labeling are described. Glycerol label is only found in acylglycerols, largely as the intact glyceryl backbone, but acetate can label acyl groups and sterols, the latter to a much higher relative specific activity. This presumably occurs because mevalonic acid precursor is derived from the non-plastid pool of acetyl-CoA that is also the source for malonyl-CoA to drive FAE1-dependent chain elongation. Particular attention has been paid to the separation of sterols and diacylglycerols, and to hydrogenation of triacylglycerols to simplify their analysis. These improved methods will allow more accurate analyses of the fluxes of lipid metabolism in cultured plant embryos. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Nrf2 inhibition affects cell cycle progression during early mouse embryo development.

    Science.gov (United States)

    Lin, Ying; Sui, Liu-Cai; Wu, Rong-Hua; Ma, Ru-Jun; Fu, Hai-Yan; Xu, Juan-Juan; Qiu, Xu-Hua; Chen, Li

    2017-12-16

    Brusatol, a quassinoid isolated from the fruit of Bruceajavanica, has recently been shown to inhibit nuclear factor erythroid 2-related factor 2 (Nrf2) via Keap1-dependent ubiquitination and proteasomal degradation or protein synthesis. Nrf2 is a transcription factor that regulates the cellular defense response. Most studies have focused on the effects of Nrf2in tumor development. Here, the critical roles of Nrf2 in mouse early embryonic development were investigated. We found that brusatol treatment at the zygotic stage prevented the early embryo development. Most embryos stayed at the two-cell stage after 5 days of culture (P CRISPR activation plasmid. Thus, brusatol inhibited early embryo development by affecting Nrf2-related cell cycle transition from G2 to M phase that is dependent on cyclin B-CDK1 complex.

  17. Quantitative proteomics of Xenopus laevis embryos: expression kinetics of nearly 4000 proteins during early development

    Science.gov (United States)

    Sun, Liangliang; Bertke, Michelle M.; Champion, Matthew M.; Zhu, Guijie; Huber, Paul W.; Dovichi, Norman J.

    2014-03-01

    While there is a rich literature on transcription dynamics during the development of many organisms, protein data is limited. We used iTRAQ isotopic labeling and mass spectrometry to generate the largest developmental proteomic dataset for any animal. Expression dynamics of nearly 4,000 proteins of Xenopus laevis was generated from fertilized egg to neurula embryo. Expression clusters into groups. The cluster profiles accurately reflect the major events that mark changes in gene expression patterns during early Xenopus development. We observed decline in the expression of ten DNA replication factors after the midblastula transition (MBT), including a marked decline of the licensing factor XCdc6. Ectopic expression of XCdc6 leads to apoptosis; temporal changes in this protein are critical for proper development. Measurement of expression in single embryos provided no evidence for significant protein heterogeneity between embryos at the same stage of development.

  18. Effects of low doses of mifepristone on human embryo implantation process in a three-dimensional human endometrial in vitro co-culture system.

    Science.gov (United States)

    Boggavarapu, N R; Berger, C; von Grothusen, C; Menezes, J; Gemzell-Danielsson, K; Lalitkumar, P G L

    2016-08-01

    We wanted to explore the effects of two different low doses (0.5μM and 0.05μM) of mifepristone, exposed during the receptive period, on the human embryo implantation process, using a well-established three-dimensional in vitro cell culture model, specifically developed to study this process. An in vitro three-dimensional cell culture model was constructed using human endometrial cells isolated from the endometrium of proven fertile women, collected on cycle day LH+4. After 5 days of culture, supernumerary human embryos were added and cultured for another 5 days with mifepristone 0.5μM (n=8) or 0.05μM (n=10) or vehicle as control (n=10). The cultures were checked for embryo attachment and terminated. We studied the expression of 16 reported endometrial receptivity markers in the endometrial constructs using real-time polymerase chain reaction. None of the embryos in 0.5μM of mifepristone attached to the endometrial constructs (p=.004), whereas 4 out of 10 in 0.05μM (p=.3698) and 7 out of 10 embryos in the control group attached to the cultures. We found that most of the studied receptivity markers were significantly altered with mifepristone exposure in a similar direction in both treatment groups. Only IL6 was significantly differentially expressed between the treatment groups (p=.017). We report for the first time that exposure to a low concentration (0.5μM) of mifepristone during the receptive period successfully inhibits human embryo implantation process in vitro. Further, we observed a dose-dependent effect of mifepristone on endometrial receptivity at the functional level. This study contributes new knowledge that low dose of mifepristone during the short period of receptive phase can inhibit endometrial receptivity, which further promotes mifepristone as a contraceptive agent. This could give women a treatment choice to avoid unwanted pregnancy with high efficacy and minimal side effects. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Differences in egg nutrient availability, development, and nutrient metabolism of broiler and layer embryos.

    Science.gov (United States)

    Nangsuay, A; Molenaar, R; Meijerhof, R; van den Anker, I; Heetkamp, M J W; Kemp, B; van den Brand, H

    2015-03-01

    Selection for production traits of broilers and layers leads to physiological differences, which may already be present during incubation. This study aimed to investigate the influence of strain (broiler vs layer) on egg nutrient availability, embryonic development and nutrient metabolism. A total of 480 eggs with an egg weight range of 62.0 to 64.0 g from Lohmann Brown Lite and Ross 308 breeder flocks of 41 or 42 weeks of age were selected in two batches of 120 eggs per batch per strain. For each batch, 30 eggs per strain were used to determine egg composition, including nutrient and energy content, and 90 eggs per strain were separately incubated in one of two climate respiration chambers at an eggshell temperature of 37.8°C. The results showed that broiler eggs had a higher ratio of yolk: albumen with 2.41 g more yolk and 1.48 g less albumen than layers. The yolk energy content of broiler eggs was 46.32 kJ higher than that of layer eggs, whereas total energy content of broiler eggs was 47.85 kJ higher compared to layer eggs. Yolk-free body mass at incubation day 16 and chick weight and length at hatch were higher in broilers compared to layers. Respiration quotient of broiler embryos was higher than layer embryos during incubation day 8 to incubation day 10. A 0.24 g lower residual yolk at the hatch of broiler embryos than for the layer embryos indicated that broiler embryos used more yolk and had a higher energy utilization and energy deposition in yolk-free body mass. Heat production of broiler embryos was higher than that of layer embryos from incubation day 12 to incubation day 18, but efficiency of converting egg energy used by embryos to form yolk-free body mass was similar. In conclusion, broiler and layer embryos have different embryonic development patterns, which affect energy utilization and embryonic heat production. However, the embryos are equal in efficiency of converting the energy used to yolk-free body mass. © 2015 Poultry Science

  20. Nickel affects gill and muscle development in oriental fire-bellied toad (Bombina orientalis) embryos

    Energy Technology Data Exchange (ETDEWEB)

    Park, Chan Jin; Song, Sang Ha; Kim, Dae Han; Gye, Myung Chan, E-mail: mcgye@hanyang.ac.kr

    2017-01-15

    Highlights: • Nickel inhibited the development of external gill in B. orientalis embryos. • The 168 h LC{sub 50} and EC{sub 50} values of nickel were 33.8 and 5.4 μM, respectively, in embryos. • Nickel induced abnormal tail development of embryos. • NF stage 26–31 was the most sensitive window for embryos to nickel exposure. • Nickel affected the calcium-dependent myogenic gene expression in embryos. - Abstract: The developmental toxicity of nickel was examined in the embryos of Bombina orientalis, a common amphibian in Korea. Based on a standard frog embryo teratogenesis assay, the LC{sub 50} and EC{sub 50} for malformation of nickel after 168 h of treatment were 33.8 μM and 5.4 μM, respectively. At a lethal concentration (100 μM), nickel treatment decreased the space between gill filaments and caused epithelial swelling and abnormal fusion of gill filaments. These findings suggest that nickel affects the functional development of gills, leading to embryonic death. At sublethal concentrations (1–10 μM), nickel produced multiple embryonic abnormalities, including bent tail and tail dysplasia. At 10 μM, nickel significantly decreased tail length and tail muscle fiber density in tadpoles, indicating inhibition of myogenic differentiation. Before hatching, the pre-muscular response to muscular response stages (stages 26–31) were the most sensitive period to nickel with respect to tail muscle development. During these stages, MyoD mRNA was upregulated, whereas myogenic regulatory factor 4 mRNA was downregulated by 0.1 μM nickel. Calcium-dependent kinase activities in muscular response stage embryos were significantly decreased by nickel, whereas these activities were restored by exogenous calcium. In tadpoles, 10 μM nickel significantly decreased the expression of the myosin heavy chain and the 12/101 muscle marker protein in the tail. Expression was restored by exogenous calcium. Our results indicate that nickel affects muscle development by

  1. Ethics, politics, and human embryo stem cell research.

    Science.gov (United States)

    Macklin, R

    2000-01-01

    Three reports on ethical aspects of research involving human embryonic stem cells were issued in the final months of 1999. Two were from governmental agencies or commissions: the National Institutes of Health and the National Bioethics Advisory Commission. The third report was issued by the American Association for the Advancement of Science and the Institute for Civil Society. All three reports endorse the use of federal funds for embryonic stem cell research, but other differences distinguish these reports. This article describes the differences and provides an ethical analysis of the main arguments.

  2. Is the creation of admixed embryos "an offense against human dignity"?

    Science.gov (United States)

    Jones, David Albert

    2010-01-01

    The controversy over the creation of admixed human-nonhuman embryos, and specifically of what have been termed "cybrids," involves a range of ethical and political issues. It is not reducible to a single question. This paper focuses on one question raised by that controversy, whether creating admixed human-nonhuman entities is "an offense against human dignity. "In the last decade there has been sustained criticism of the use of the concept of human dignity within bioethics. The concept has been criticized as "vague" and "useless." Nevertheless, the concept continues to be invoked in bioethical discussion and in international instruments. This paper defends a concept of human dignity that is coherent but that is wider than contemporary post-Kantian approaches. "Human dignity" is best regarded as having a set of analogically related meanings, more than one of which is relevant to the field of bioethics. A more subtle understanding of the concept of human dignity can help identify what is ethically problematic in human-nonhuman combinations and so shed light on one aspect of the admixed embryo debate.

  3. Effects of reactive oxygen species levels in prepared culture media on embryo development: a comparison of two media.

    Science.gov (United States)

    Shih, Ying-Fu; Lee, Tsung-Hsien; Liu, Chung-Hsien; Tsao, Hui-Mei; Huang, Chun-Chia; Lee, Maw-Sheng

    2014-12-01

    This study determined the correlation between the levels of reactive oxygen species (ROS) in prepared culture media and the early development of human embryos. This was an autocontrolled comparison study. A total of 159 patients undergoing in vitro fertilization/intracytoplasmic sperm injection treatment were recruited in this study. The pH values, osmolarity pressures, and ROS levels of 15 batches of two culture media were measured. Sibling oocytes or embryos from individual patients were randomly assigned to two culture groups with Quinn's Advantage Cleavage and Blastocyst media (QAC/QAB) or GIII series cleavage and blastocyst media (G1.3/G2.3). The difference between the two culture groups was analyzed using one-sample t test. The QAC/QAB and G1.3/G2.3 media exhibited similar pH values and osmolarity pressures. However, the prepared QAC/QAB media were characterized to contain lower amounts of ROS than the G1.3/G2.3 media. Furthermore, the blastocysts that developed under the QAC/QAB media were morphologically superior to those that developed under the G1.3/G2.3 media. The elevated ROS levels in culture media were associated with poor development of blastocyst-stage embryos. Measurement of ROS levels may be a valuable process for medium selection or modification. Copyright © 2014. Published by Elsevier B.V.

  4. Dynamic Subcellular Localization of Iron during Embryo Development in Brassicaceae Seeds

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    Miguel A. Ibeas

    2017-12-01

    Full Text Available Iron is an essential micronutrient for plants. Little is know about how iron is loaded in embryo during seed development. In this article we used Perls/DAB staining in order to reveal iron localization at the cellular and subcellular levels in different Brassicaceae seed species. In dry seeds of Brassica napus, Nasturtium officinale, Lepidium sativum, Camelina sativa, and Brassica oleracea iron localizes in vacuoles of cells surrounding provasculature in cotyledons and hypocotyl. Using B. napus and N. officinale as model plants we determined where iron localizes during seed development. Our results indicate that iron is not detectable by Perls/DAB staining in heart stage embryo cells. Interestingly, at torpedo development stage iron localizes in nuclei of different cells type, including integument, free cell endosperm and almost all embryo cells. Later, iron is detected in cytoplasmic structures in different embryo cell types. Our results indicate that iron accumulates in nuclei in specific stages of embryo maturation before to be localized in vacuoles of cells surrounding provasculature in mature seeds.

  5. Protein Expression Landscape of Mouse Embryos during Pre-implantation Development

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    Yawei Gao

    2017-12-01

    Full Text Available Pre-implantation embryo development is an intricate and precisely regulated process orchestrated by maternally inherited proteins and newly synthesized proteins following zygotic genome activation. Although genomic and transcriptomic studies have enriched our understanding of the genetic programs underlying this process, the protein expression landscape remains unexplored. Using quantitative mass spectrometry, we identified nearly 5,000 proteins from 8,000 mouse embryos of each stage (zygote, 2-cell, 4-cell, 8-cell, morula, and blastocyst. We found that protein expression in zygotes, morulas, and blastocysts is distinct from 2- to 8-cell embryos. Analysis of protein phosphorylation identified critical kinases and signal transduction pathways. We highlight key factors and their important roles in embryo development. Combined analysis of transcriptomic and proteomic data reveals coordinated control of RNA degradation, transcription, and translation and identifies previously undefined exon-junction-derived peptides. Our study provides an invaluable resource for further mechanistic studies and suggests core factors regulating pre-implantation embryo development.

  6. Localization and expression of histone H2A variants during mouse oogenesis and preimplantation embryo development.

    Science.gov (United States)

    Wu, B J; Dong, F L; Ma, X S; Wang, X G; Lin, F; Liu, H L

    2014-08-07

    Epigenetic modifications of the genome, such as histone H2A variants, ensure appropriate gene activation or silencing during oogenesis and preimplantation embryo development. We examined global localization and expression of the histone H2A variants, including H2A.Bbd, H2A.Z and H2A.X, during mouse oogenesis and preimplantation embryo development. Immunocytochemistry with specific antibodies against various histone H2A variants showed their localization and changes during oogenesis and preimplantation development. H2A.Bbd and H2A.Z were almost absent from nuclei of growing oocytes (except 5-day oocyte), whereas H2A.X was deposited in nuclei throughout oogenesis and in preimplantation embryos. In germinal vesicle (GV) oocyte chromatin, H2A.Bbd was detected as a weak signal, whereas no fluorescent signal was detected in GV breakdown (GVBD) or metaphase II (MII) oocytes; H2A.Z showed intense signals in chromatin of GV, GVBD and MII oocytes. H2A. Bbd showed very weak signals in both pronucleus and 2-cell embryo nuclei, but intense signals were detected in nuclei from 4-cell embryo to blastula. The H2A.Z signal was absent from pronucleus to morula chromatin, whereas a fluorescent signal was detected in blastula nuclei. Our results suggest that histone H2A variants are probably involved in reprogramming of genomes during oocyte meiosis or after fertilization.

  7. Dynamic Subcellular Localization of Iron during Embryo Development in Brassicaceae Seeds.

    Science.gov (United States)

    Ibeas, Miguel A; Grant-Grant, Susana; Navarro, Nathalia; Perez, M F; Roschzttardtz, Hannetz

    2017-01-01

    Iron is an essential micronutrient for plants. Little is know about how iron is loaded in embryo during seed development. In this article we used Perls/DAB staining in order to reveal iron localization at the cellular and subcellular levels in different Brassicaceae seed species. In dry seeds of Brassica napus, Nasturtium officinale, Lepidium sativum, Camelina sativa, and Brassica oleracea iron localizes in vacuoles of cells surrounding provasculature in cotyledons and hypocotyl. Using B. napus and N. officinale as model plants we determined where iron localizes during seed development. Our results indicate that iron is not detectable by Perls/DAB staining in heart stage embryo cells. Interestingly, at torpedo development stage iron localizes in nuclei of different cells type, including integument, free cell endosperm and almost all embryo cells. Later, iron is detected in cytoplasmic structures in different embryo cell types. Our results indicate that iron accumulates in nuclei in specific stages of embryo maturation before to be localized in vacuoles of cells surrounding provasculature in mature seeds.

  8. Effects of griseofulvin on in vitro porcine oocyte maturation and embryo development.

    Science.gov (United States)

    Miao, Yi-Liang; Zhang, Xia; Zhao, Jian-Guo; Spate, Lee; Zhao, Ming-Tao; Murphy, Clifton N; Prather, Randall S; Sun, Qing-Yuan; Schatten, Heide

    2012-08-01

    Griseofulvin is an orally administered antifungal drug that affects microtubule formation in vitro and interferes with microtubule dynamics in vivo as clearly shown for mitotic cells in several cell systems. This article reports the effects of griseofulvin on in vitro maturation of porcine oocytes and subsequent effects on embryo development. Our results revealed a concentration-dependent effect on meiotic spindles with 20-40 μM griseofulvin affecting oocyte maturation, and 40 μM affecting fertilization and embryo development. These concentrations of griseofulvin did not affect mitochondrial and cortical granule distribution that also depend on microtubule and cytoskeletal functions during oocyte maturation. Specific effects on the meiotic spindle included spindle disorganization and aberrant chromosome separation displayed as prominent chromosome clusters in oocytes treated with 40 μM griseofulvin. These results strongly suggested that griseofulvin affected porcine oocyte in vitro maturation and following embryo development by disturbing microtubule dynamics. Copyright © 2012 Wiley Periodicals, Inc.

  9. Volatiles identified from five stages of embryo development separated from a heterogeneous suspension culture of Daucus carota.

    Science.gov (United States)

    Kennedy, A H; Chamberlain, D; Wilson, G; Ryan, M F

    1991-11-01

    Five stages of embryo development were fractionated from a mature culture of Daucus carota (Gelbe Rheinsche), using a series of metal sieves. The composition of the population of embryos in each fraction was determined quantitatively from microscopic investigations. Volatiles from samples of tissue from six stages of development were trapped on activated charcoal cartridges. These volatiles, some of which may play a significant role in the interaction of the plant with the carrot root fly (Psila rosae), were analysed using gas chromatography/mass spectroscopy. The resulting chromatograms are arranged in order of embryo development. The progressive elaboration of the volatile profile reflects the increased biosynthetic capacity of the developing embryo.

  10. Embryo Development of Tree Frog Polypedates leucomystax at Campus of State University of Malang

    Directory of Open Access Journals (Sweden)

    Pearlindah

    2012-12-01

    Full Text Available Tree frogs live in natural places which are unpolluted. Regarding their role as an ecological indicator, the decrease of frogs population in a particular habitat indicates the danger of environment quality decrease. Moreover, this condition can harm the frogs themselves. All kinds of frogs breed in aqueous environment such as ponds, marshes, and farming fields. One of the tree frogs, Polypedates leucomystax, which belongs to Familia Rachophoridae, is widely spread in Indonesia. This frog has yellowish brown skin with black spots or six lines extending from head to the posterior tip of body. A breeding couple of the frog produces foam nests on the water or plants around water body, where they will nest their fertilized eggs. This species produces over a hundred embryos in one spawning season. These embryos require appropriate conditions to develop normally in the nature. Frog embryo development may becomes a reference to understand how the frog population survives. This study focused on P. leucomystax with regards to its decrease in number due to the drying up of the environment and a lot lost of trees in Campus of State University of Malang. The development of P. leucomystax embryos in the reproduction foam was observed until it reached a tadpole stage. The result showed that the embryos developed in the foam until they hatched then they move out of the foam into the water under which they would continue their development. Considering that water body is a critical requirement for the development of P. leucomystax embryos, it is our responsibility to make any efforts to conserve not only the trees but also any type of water bodies including ponds, marshes, and farming fields as well.

  11. In vitro and in vivo effects of ulipristal acetate on fertilization and early embryo development in mice.

    Science.gov (United States)

    Gómez-Elías, Matías D; Munuce, María J; Bahamondes, Luis; Cuasnicú, Patricia S; Cohen, Débora J

    2016-01-01

    vitro embryo culture. Considering the ethical and technical limitations inherent to the use of human gametes for fertilization studies, the mouse model was used as an approach for exploring the potential effects of UPA on in vivo sperm transport and fertilization. Nevertheless, the extrapolation of these results to humans requires further investigation. This study presents new evidence on the lack of effect of UPA on gamete interaction and embryo development, providing new insights into the mechanism of action of UPA as an emergency contraceptive method with potential clinical implications. These new findings could contribute to increase the acceptability and proper use of UPA as an emergency contraceptive method. This study was partially supported by a National Agency of Scientific and Technological Promotion (ANPCyT), Argentina grants PICT 2011-061 to D.J.C. and PICT 2011-2023 to P.S.C. None of the authors has any competing interests to declare. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  12. EVALUATION OF ETHINYLESTRADIOL (EE2 EFFECT ON EMBRYO DEVELOPMENT IN COMMON CARP (CYPRINUS CARPIO

    Directory of Open Access Journals (Sweden)

    GABI DUMITRESCU

    2009-10-01

    Full Text Available Worldwide, the scientific researches performed during the last years are focused on the determination of the negative effects caused by natural and antropogeneous chemical compounds on aquatic species; these species are more exposed to most pollutants than the land species, for the simple reason that the aquatic environment is the last destination for most residues. Our research team proposed to test the toxic effect caused by ethinylestradiol on embryo development in common carp (Cyprinus carpio. Common carp embryos were purchased from the fish farm S.C. Acva Prod S.R.L. Cefa, Bihor County these were obtained by artificial reproduction. After taking and selection, the fecundated spawns were introduced in 10 Nunk culture plates of 45 ml, where we introduced 40 ml water, too. We created 3 batches, with two replications, namely: batch 1 – control, batch 2 – in water, we added ethinylestradiol (EE2 in concentration of 1.5 ng L-1 and batch 3 – we added in water a concentration of 7 ng L-1 EE2. During the incubation, the Nunk plates were kept in breeding aquariums, at a temperature of 24°C. Successive to the supervision of embryos in batch 3, 48 hours post-fecundation, we could observe evolution stagnations, 70% of them being in the stage of 40 somites of the segmentation period. At the same age, 100% of the control batch- embryos entered the stage of advanced faringula, and in batch 2 all embryos were in the stage of incipient faringula. 60-72 hours post-fecundation, all embryos in the batch 3 died, 90% in the 40 somite stage of the segmentation period and 10% in the stage of incipient faringula. 85 hours post-fecundation, all embryos belonging to the control batch were in the larva stage, while in batch 2, 90% were in the larva stage and 10% died in the stage of advanced faringula.

  13. Development of hematological respiratory variables in late chicken embryos: the relative importance of incubation time and embryo mass.

    Science.gov (United States)

    Tazawa, Hiroshi; Andrewartha, Sarah J; Burggren, Warren W

    2011-07-01

    Oxygen demand increases during embryonic development, requiring an increase in red blood cells (RBCs) containing hemoglobin (Hb) to transport O(2) between the respiratory organ and systemic tissues. A thorough ontogenetic understanding of the onset and maturation of the complex regulatory processes for RBC concentration ([RBC]), Hb concentration ([Hb]), hematocrit (Hct), mean corpuscular indices (mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration ([MCHb])) is currently lacking. We hypothesize that during the last half of incubation when the respiratory organ (the chorioallantoic membrane) envelops most of the egg contents, mean corpuscular indices will stabilize. Accordingly, Hct, [RBC] and [Hb] must also all change proportionally across development. Further, we hypothesize that the hematological respiratory variables develop and mature as a function of incubation duration, independently of embryonic growth. As predicted, a similar increase in Hct (from 18.7±0.6% on day 10 (d10) to 34.1±0.5% on d19 of incubation), [RBC] (1.13±0.03×10(6)/μL to 2.50±0.03×10(6)/μL) and [Hb] (6.1±0.2 g% to 11.2±0.1 g%) occurred during d10-19. Both [RBC] and [Hb] demonstrated high linear correlation with Hct, resulting in constant [MCHb] (~33 g% from d10 to d19). The decrease in MCV (from ~165 μ(3) on d10 to ~140 μ(3) on d13) and MCH (~55 pg to ~45 pg) during d10-13, may be attributed to a changeover from larger primary to smaller secondary and adult-type erythrocytes with MCV and MCH remaining constant (~140 μ(3) and ~45 pg respectively) for the rest of the incubation period (d13-19). Hematological respiratory values on a given incubation day were identical between embryos of different masses using either natural mass variation or experimental growth acceleration, indicating that the hematological variables develop as a function of incubation time, irrespective of embryo growth. Copyright © 2011. Published by

  14. Guarding embryo development of zebrafish by shell engineering: a strategy to shield life from ozone depletion.

    Science.gov (United States)

    Wang, Ben; Liu, Peng; Tang, Yanyan; Pan, Haihua; Xu, Xurong; Tang, Ruikang

    2010-04-01

    The reduced concentration of stratospheric ozone results in an increased flux of biologically damaging mid-ultraviolet radiation (UVB, 280 to 320 nm) reaching earth surfaces. Environmentally relevant levels of UVB negatively impact various natural populations of marine organisms, which is ascribed to suppressed embryonic development by increased radiation. Inspired by strategies in the living systems generated by evolution, we induce an extra UVB-adsorbed coat on the chorion (eggshell surrounding embryo) of zebrafish, during the blastula period. Short and long UV exposure experiments show that the artificial mineral-shell reduces the UV radiation effectively and the enclosed embryos become more robust. In contrast, the uncoated embryos cannot survive under the enhanced UVB condition. We suggest that an engineered shell of functional materials onto biological units can be developed as a strategy to shield lives to counteract negative changes of global environment, or to provide extra protection for the living units in biological research.

  15. Kinetics data from bovine sex-specific embryo development from three different bulls

    Directory of Open Access Journals (Sweden)

    C.S. Oliveira

    2016-06-01

    Full Text Available Here we present kinetics data from bovine sex-specific embryo development. Embryos were originated using sex-sorted semen from three different Nelore bulls, and semen from the same batch was used for X-and Y-chromosome spermatozoa sorting. Data was obtained for six time points (24, 48, 96, 120, and 144 h.p.i.. Analyses for each bull׳s embryos (1, 2 and 3 is presented for female and male groups separately. Also, grouped data analysis, considering bull and sex interaction, is shown. For further interpretation and discussion, see "Cell death is involved in sexual dimorphism during preimplantation development" (Oliveira et al., 2015 [1].

  16. Development of Au nanowire injector system to deliver plasmid into mouse embryo

    Directory of Open Access Journals (Sweden)

    Kkotchorong Park

    2017-10-01

    Full Text Available In this data article, we developed a Au nanowire injector (Au NWI for directly delivering plasmid into the 1-cell stage of the mouse embryos designed to successfully attach and detach the plasmid on the Au NWI, highly minimizing physical and chemical damage on the embryos. This data presents that a Au NWI system does not induce detrimental damages on development of embryos and efficiently express the green fluorescence protein in vitro. The data provided herein is in association with the research article related to reduce the occurrence of mosaicism by a Au NWI,” Suppressing Mosaicism by Au Nanowire Injector-driven Direct Delivery of Plasmids into Mouse Embryos” (Park et al., 2017 [1].

  17. In vitro development of cloned embryos derived from miniature pig somatic cells after activation by ultrasound stimulation.

    Science.gov (United States)

    Miyoshi, Kazuchika; Sato, Keisuke; Yoshida, Mitsutoshi

    2006-01-01

    The present study was carried out to examine the activation and development of cloned embryos produced by transferring miniature pig somatic cells into enucleated farm pig oocytes after exposing to ultrasound. The rates of the pronucleus-like structure formation and polar body-like structure extrusion in embryos exposed to ultrasound did not differ from those applied electric pulses. Although there was no significant difference in the blastocyst formation rates between different activation methods, the mean number of cells in the blastocysts developed from embryos activated by exposing to ultrasound was significantly (p ultrasound stimulation can induce the activation and in vitro development of cloned embryos derived from miniature pig somatic cells.

  18. Effect of nanoparticles of silver and gold on metabolic rate and development of broiler and layer embryos

    DEFF Research Database (Denmark)

    Pineda, L; Sawosz, E; Hotowy, A

    2012-01-01

    -air-circuit respiration unit, and HP was calculated for 10, 13, 16 and 19-day-old embryos. Relative chick and muscle weights were used as a measure of growth rate and development. AgNano but not AuNano increased the rates of O(2) consumption and HP of the layer embryos. The metabolic rate of broiler embryos...... was not affected by either of the treatments, but it was significantly higher compared to the layer embryos. Neither of the nanoparticles promoted nor depressed growth and development of the embryos, irrespective of breed. Although the metabolic rate of AgNano-injected layer embryos was significantly increased......, their BW and muscle weights at hatching were similar to those of the control group, which suggests that the concentration of AgNano used was adequate for increasing the metabolic rate but not enough to affect growth and development. The results show that AgNano could be a potential metabolic modifier...

  19. Extracellular Vesicles from BOEC in In Vitro Embryo Development and Quality.

    Directory of Open Access Journals (Sweden)

    Ricaurte Lopera-Vásquez

    Full Text Available To evaluate the effect of conditioned media (CM and Extracellular Vesicles (EVs derived from bovine oviduct epithelial cell (BOEC lines on the developmental capacity of bovine zygotes and the quality of embryos produced in vitro, presumptive zygotes were cultured under specific conditions. In experiment 1, zygotes were cultured either on monolayers from BOEC extended culture (E, together with fresh BOEC suspension cells, or with BOEC-CM from fresh or E-monolayers. In experiment 2, EVs were isolated from BOEC-CM and characterized (150-200 nm by Nanosight® and electron microscopy. Zygotes were cultured in the presence of 3x10(5 EVs/mL, 1.5x10(5 EVs/mL or 7.5x10(4 EVs/mL of fresh or frozen BOEC-EVs. In experiment 3, zygotes were cultured in absence of FCS but with EVs from BOEC-E that had been cultured in different culture media. In experiment 4, zygotes were cultured in SOF+5% normal-FCS, or EV-depleted-FCS. In all cases, cleavage rate (Day 2 and blastocyst development (Day 7-9 was assessed. Blastocysts on Days 7/8 were used for quality evaluation through differential cell count, cryotolerance and gene expression patterns. No differences were found among all FCS-containing groups in cleavage rate or blastocyst yield. However, embryos derived from BOEC-CM had more trophectoderm cells, while embryos derived from BOEC-EVs, both fresh and frozen, has more trophectoderm and total cells. More embryos survived vitrification in the BOEC-CM and BOEC-EV groups. In contrast, more embryos survived in the EV-depleted-FCS than in normal-FCS group. Gene expression patterns were modified for PAG1 for embryos cultured with EVs in the presence of FCS and for IFN-T, PLAC8, PAG1, CX43, and GAPDH in the absence of FCS. In conclusion, EVs from FCS have a deleterious effect on embryo quality. BOEC-CM and EVs during in vitro culture had a positive effect on the quality of in vitro produced bovine embryos, suggesting that EVs have functional communication between the

  20. Extracellular Vesicles from BOEC in In Vitro Embryo Development and Quality.

    Science.gov (United States)

    Lopera-Vásquez, Ricaurte; Hamdi, Meriem; Fernandez-Fuertes, Beatriz; Maillo, Verónica; Beltrán-Breña, Paula; Calle, Alexandra; Redruello, Alberto; López-Martín, Soraya; Gutierrez-Adán, Alfonso; Yañez-Mó, María; Ramirez, Miguel Ángel; Rizos, Dimitrios

    2016-01-01

    To evaluate the effect of conditioned media (CM) and Extracellular Vesicles (EVs) derived from bovine oviduct epithelial cell (BOEC) lines on the developmental capacity of bovine zygotes and the quality of embryos produced in vitro, presumptive zygotes were cultured under specific conditions. In experiment 1, zygotes were cultured either on monolayers from BOEC extended culture (E), together with fresh BOEC suspension cells, or with BOEC-CM from fresh or E-monolayers. In experiment 2, EVs were isolated from BOEC-CM and characterized (150-200 nm) by Nanosight® and electron microscopy. Zygotes were cultured in the presence of 3x10(5) EVs/mL, 1.5x10(5) EVs/mL or 7.5x10(4) EVs/mL of fresh or frozen BOEC-EVs. In experiment 3, zygotes were cultured in absence of FCS but with EVs from BOEC-E that had been cultured in different culture media. In experiment 4, zygotes were cultured in SOF+5% normal-FCS, or EV-depleted-FCS. In all cases, cleavage rate (Day 2) and blastocyst development (Day 7-9) was assessed. Blastocysts on Days 7/8 were used for quality evaluation through differential cell count, cryotolerance and gene expression patterns. No differences were found among all FCS-containing groups in cleavage rate or blastocyst yield. However, embryos derived from BOEC-CM had more trophectoderm cells, while embryos derived from BOEC-EVs, both fresh and frozen, has more trophectoderm and total cells. More embryos survived vitrification in the BOEC-CM and BOEC-EV groups. In contrast, more embryos survived in the EV-depleted-FCS than in normal-FCS group. Gene expression patterns were modified for PAG1 for embryos cultured with EVs in the presence of FCS and for IFN-T, PLAC8, PAG1, CX43, and GAPDH in the absence of FCS. In conclusion, EVs from FCS have a deleterious effect on embryo quality. BOEC-CM and EVs during in vitro culture had a positive effect on the quality of in vitro produced bovine embryos, suggesting that EVs have functional communication between the oviduct and

  1. PHO1 Exports Phosphate from the Chalazal Seed Coat to the Embryo in Developing Arabidopsis Seeds.

    Science.gov (United States)

    Vogiatzaki, Evangelia; Baroux, Célia; Jung, Ji-Yul; Poirier, Yves

    2017-10-09

    Seed production requires the transfer of nutrients from the maternal seed coat to the filial endosperm and embryo. Because seed coat and filial tissues are symplasmically isolated, nutrients arriving in the seed coat via the phloem must be exported to the apoplast before reaching the embryo. Proteins implicated in the transfer of inorganic phosphate (Pi) from the seed coat to the embryo are unknown despite seed P content being an important agronomic trait. Here we show that the Arabidopsis Pi exporters PHO1 and PHOH1 are expressed in the chalazal seed coat (CZSC) of developing seeds. PHO1 is additionally expressed in developing ovules. Phosphorus (P) content and Pi flux between the seed coat and embryo were analyzed in seeds from grafts between WT roots and scions from either pho1, phoh1, or the pho1 phoh1 double mutant. Whereas P content and distribution between the seed coat and embryo in fully mature dry seeds of these mutants are similar to the WT, at the mature green stage of seed development the seed coat of the pho1 and pho1 phoh1 mutants, but not of the phoh1 mutant, retains approximately 2-fold more P than its WT control. Expression of PHO1 under a CZSC-specific promoter complemented the seed P distribution phenotype of the pho1 phoh1 double mutant. CZSC-specific down-expression of PHO1 also recapitulated the seed P distribution phenotype of pho1. Together, these experiments show that PHO1 expression in the CZSC is important for the transfer of P from the seed coat to the embryo in developing seeds. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. [Prolonged culture of human embryos: comparison of coculture on human tubal epithelium and culture in synthetic media].

    Science.gov (United States)

    Ventruba, P; Záková, J; Adler, J; Trávník, P; Komárková, J; Nĕmcová, S

    1996-12-01

    The aim of this work was to evaluate prolongation of cultivation time of human embryos. The prolongation above the standard limit was implemented (1) by coculture of embryos on a monolayer of human epithelial cells and (2) by using a synthetic medium. Ovarian stimulation, oocyte recovery, insemination and cultivation up to the pronuclear stage were done in our centre in the usual way. Group I: 104 women, prolonged culture by the coculture method. The zygotes were placed on the monolayer and cultivated for an other 24 or 48 hours. Group II: 249 women, prolonged culture in a synthetic medium. The zygotes were cultivated up to the 2- to 4-cell stage in standard IVF medium, and then put into M3 medium for the next 24 or 48 hours. A transfer of a maximum of 4 embryos was done. In group I in 104 women from 341 zygotes 181 embryos (53.1%) reached the eight- and more than eight-cell stage, and 76 transfers were done. 15 pregnancies were achieved (19.4% pregnancy rate). In group II in 249 patients from 672 embryos 49.7% of them reached 8- and more than 8-cell stage. 51 pregnancies were achieved (22.6 pregnancy rate). In the control group of 250 IVF after 48 hours cultivation 165 transfers (66.0%) were done, and 16.4% pregnancies were achieved, i.e. 6.2% less compared to synthetic medium and 3.3% less than in cocultivation. There is evidence of better IVF/ET results in case of prolonged culture time. The experience in our centre has shown the need of reevaluation of the coculture method. The exacting character of its preparation and manipulation will have to be replaced by synthetic media in spite of their high price.

  3. Semi-viviparous embryo development and dehydrin expression in the mangrove Rhizophora mucronata Lam.

    Science.gov (United States)

    Ismail, Flora Abdulrahman; Nitsch, Lisette M C; Wolters-Arts, Mieke M C; Mariani, Celestina; Derksen, Jan W M

    2010-06-01

    Rhizophora mucronata Lam. is a tropical mangrove with semi-viviparous (cotyledon body protrusion before shedding), non-quiescent and non-desiccating (recalcitrant) seeds. As recalcitrance has been thought to relate to the absence of desiccation-related proteins such as dehydrins, we for the first time systematically described and classified embryogenesis in R. mucronata and assessed the presence of dehydrin-like proteins. Embryogenesis largely follows the classic pattern till stage eight, the torpedo stage, with the formation of a cotyledonary body. Ovule and embryo express radical adaptations to semi-vivipary in the saline environment: (1) A large, highly vacuolated and persistent endosperm without noticeable food reserves that envelopes the developing embryo. (2) Absence of vascular tissue connections between embryo and maternal tissue, but, instead, transfer layers in between endosperm and integument and endosperm and embryo. Dehydrin-like proteins (55-65 kDa) were detected by the Western analysis, in the ovules till stage 10 when the integuments are dehisced. An additional 50 kDa band was detected at stages 6-8. Together these results suggest a continuous flow of water with nutrients from the integument via the endosperm to the embryo, circumventing the vascular route and probably suppressing the initially induced dehydrin expression.

  4. Sperm selection based on motility in polyvinylpyrrolidone is associated with successful pregnancy and embryo development.

    Science.gov (United States)

    Irez, T; Ocal, P; Guralp, O; Kaleli, S; Ocer, F; Sahmay, S

    2013-08-01

    The aim of this study was to investigate whether spermatozoon motility in polyvinylpyrrolidone (PVP) is associated with better embryo development and pregnancy rates in ICSI cycles. A total of 123 primary ICSI treatment cycles were included in this study. Semen samples were tested for motility before ICSI procedure in PVP. Within 3 min, the presence or absence of motility was recorded. Sperm functions were examined by the aniline blue (AB) chromatin condensation test and the hypoosmotic swelling test, and the chromatin stability was evaluated by inducing its decondensation with sodium dodecyl sulphate and ethylenediaminetetraacetic acid (EDTA). Fertilisation and embryo scoring were evaluated. Fifty (64%) of 78 women conceived in the PVP (+) group; and 12 (26%) of 45 women conceived in the PVP (-) group; the pregnancy rate was significantly higher in the PVP (+) group (P = 0.003). Semen parameters were observed to be similar in both groups. The mean number of total embryos obtained in ICSI procedure and transferred grade 1 embryos were significantly higher in PVP (+) group (P = 0.01 and P = 0.003 respectively). The presence of sperm motility in PVP is associated with increased pregnancy rate, higher percentage of good quality embryos, sperm chromatin condensation and decondensation. © 2012 Blackwell Verlag GmbH.

  5. The Teratogenic Effects of Antiepileptic Drug, Topiramate, on the Development of Chick Embryos

    Directory of Open Access Journals (Sweden)

    Jantima Roongruangchai

    2017-05-01

    Full Text Available Background: Anti-epileptic drugs are known to be the risk of teratogenicity. Topiramate (TPM is a new kind of such drug, for which no research has confirmed the incidence of producing congenital abnormalities. Objective: This study was conducted to study the teratogenic effects of TPM by using chick embryos as an animal model and the results can be compared to the human embryo of the same stage. Methods: Fertilized Leghorn hen eggs were injected in ovo with two concentrations of TPM, which were 10mg, and 20mg, in NSS at a volume of 0.1 ml into the yolk sac at 21 hrs of incubation and repeated injections at 72 hrs at a volume of 0.05 ml. The chick embryos on day 3, 6 and 11 of incubation were sacrificed and all living embryos were processed for total mount and serial section. Results: The mortality rate increased corresponding to the concentrations of TPM, and the embryonic stage. The total mount of day 3 showed major abnormalities of the eye and heart, such as microphthalmia and looser of heart looping. The serial section of day 3 showed opening of the anterior neuropore, ectopia viscerae and multiple malformations of the eye and heart. Day 6 chick embryos showed ectopia cordis and ectopia viscerae. Moreover, there were retardation and abnormalities of several organs such as eye, heart, liver, mesonephros and gonads. Day 11 chick embryos showed ectopia viscerae and several growth retardations, retardation of ossification of both limb bones and skull bones. Conclusion: This study showed that TPM might cause embryonic death, growth retardation and abnormalities of the eye, heart, an opening of the anterior neuropore and ectopia viscerae. This might indicate abnormalities to the baby born from mother with gestational epilepsy who was taking this drug continuously, and it might lead to spontaneous abortion or congenital anomalies of the fetus.

  6. Effects of fluoride on morphology, growth, development, and thyroid hormone of Chinese toad (Bufo gargarizans) embryos.

    Science.gov (United States)

    Zhang, Yuhui; Xie, Lei; Li, Xinyi; Chai, Lihong; Chen, Mengxing; Kong, Xiaojing; Wang, Qingqing; Liu, Jingfei; Zhi, Lijuan; Yang, Chang; Wang, Hongyuan

    2017-10-12

    Excessive fluoride in natural water ecosystem has the potential to detrimentally affect amphibians, but little is known of such effects or underlying mechanisms in Bufo gargarizans embryos. In the present study, the effects of fluoride exposure on B. gargarizans embryos were investigated. First, fluoride teratogenic experiment showed that the 9 days EC50 of fluoride on B. gargarizans embryos was 177.62 mg/L. Then, we studied the sublethal effects of fluoride on B. gargarizans embryos at control, 0.7, 4.1, 19.6, 41.9, and 62.7 mg/L fluoride concentration. Malformation, growth, and development of embryos were monitored, and type 2 and 3 iodothyronine deiodinase (Dio2 and Dio3), thyroid hormone receptors (TRα and TRβ) mRNA levels were measured. Our results showed the morphological malformations, such as tail curvature (lordosis), edema, cuticularized ciliated cells, and hyperplasia were occurred during fluoride exposure. Growth and development were all inhibited at 19.5, 41.9, and 62.7 mg/L fluoride-treated groups after 9 days' exposure. According to real-time PCR results, exposure to fluoride upregulated Dio3 and TRβ mRNA expression and downregulated Dio2 and TRα mRNA level. All above indicated that excessive fluoride could induce morphology malformations, inhibit embryonic growth and development, and disrupt the normal function of maternal thyroid hormone in B. gargarizans embryos. Environ. Mol. Mutagen., 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  7. Imprinting disorder in donor cells is detrimental to the development of cloned embryos in pigs

    Science.gov (United States)

    Sun, Yueping; Li, Huatao; Gao, Shansong; Zhang, Liping; Xue, Binghua; Zhao, Guimin; Li, Jingyu; Liu, Zhonghua; He, Hongbin; Huan, Yanjun

    2017-01-01

    Imprinting disorder during somatic cell nuclear transfer usually leads to the abnormality of cloned animals and low cloning efficiency. However, little is known about the role of donor cell imprinting in the development of cloned embryos. Here, we demonstrated that the imprinting (H19/Igf2) in porcine fetus fibroblasts derived from the morphologically abnormal cloned fetuses (the abnormal imprinting group) was more hypomethylated, and accordingly, significantly higher H19 transcription and lower Igf2 expression occurred in comparison with those in fibroblasts derived from morphologically normal cloned fetuses (the normal imprinting group) or donor fetus fibroblasts (the control group). When these fibroblasts were used as donor cells, the abnormal imprinting group displayed an even lower imprinting methylation level, in correspondence to the significantly downregulated expression of Dnmt1, Dnmt3a and Zfp57, and a markedly reduced blastocyst rate, while the normal imprinting group took on the similar patterns of imprinting, gene expression and embryo development to the control group. When 5-aza-dC was applied to reduce the fibroblasts imprinting methylation level in the normal imprinting group, cloned embryos displayed the more severely impaired imprinting and significantly lower blastocyst rate. While the upregulated H19 transcription in the abnormal imprinting group was knocked down, the imprinting statuses were partly rescued, and the cleavage and blastocyst rates significantly increased in cloned embryos. In all, donor cell imprinting disorder reduced the developmental efficiency of cloned embryos. This work provides a new insight into understanding the molecular mechanism of donor cells regulating the cloned embryo development. PMID:29069793

  8. Vitamin D receptor signaling is required for heart development in zebrafish embryo

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Hye-Joo, E-mail: hjkwon@pnu.edu.sa [Biology Department, Texas A& M University, College Station, TX77843-3258 (United States); Biology Department, Princess Nourah University, Riyadh 11671 (Saudi Arabia)

    2016-02-12

    Vitamin D has been found to be associated with cardiovascular diseases. However, the role of vitamin D in heart development during embryonic period is largely unknown. Vitamin D induces its genomic effects through its nuclear receptor, the vitamin D receptor (VDR). The present study investigated the role of VDR on heart development by antisense-mediated knockdown approaches in zebrafish model system. In zebrafish embryos, two distinct VDR genes (vdra and vdrb) have been identified. Knockdown of vdra has little effect on heart development, whereas disrupting vdrb gene causes various cardiac phenotypes, characterized by pericardial edema, slower heart rate and laterality defects. Depletion of both vdra and vdrb (vdra/b) produce additive, but not synergistic effects. To determine whether atrioventricular (AV) cardiomyocytes are properly organized in these embryos, the expression of bmp4, which marks the developing AV boundary at 48 h post-fertilization, was examined. Notably, vdra/b-deficient embryos display ectopic expression of bmp4 towards the ventricle or throughout atrial and ventricular chambers. Taken together, these results suggest that VDR signaling plays an essential role in heart development. - Highlights: • VDR signaling is involved in embryonic heart development. • Knockdown of vdrb, but not vdra, causes decreased heart rate in zebrafish embryo. • Loss of vdr results in cardiac laterality defects. • Loss of vdra/b alters atrioventricular boundary formation. • Loss of vdra/b causes abnormal cardiac looping.

  9. Carbon monoxide and the embryo.

    Science.gov (United States)

    Robkin, M A

    1997-04-01

    Mammals are homeotherms and expend considerable energy maintaining their body temperatures. The temperature of a mammalian embryo on the other hand is maintained by the mother and the embryo can devote its metabolic energy to growth and development. The mammalian embryo is acting as a poikilotherm and its energy needs are thus considerably less than if it were a comparably sized homeotherm. The energy requirements of the preimplantation rat embryo are generated by anaerobic metabolism. As it grows, aerobic metabolism develops. In culture, the addition of carbon monoxide to the perfusing gas for early rat embryos has a much smaller effect than decreasing the oxygen concentration. Carbon monoxide appears to be a relatively mild toxicant until the embryo is much larger, is depending much more on transport of oxygen by red blood cells, and the fraction of required metabolic energy produced by anaerobic metabolism has become quite small. The effect from smoking during gestation may be either by the concomitant reduction in food intake or a more direct toxic effect from some components in the smoke. Carbon monoxide does not seem to be the culprit. The possible mitigating effect of a compensatory increase in fetal hematocrit in response to any hypoxia must also be considered. Humans have no yolk sac placenta as rodents do, but if the switch from anaerobic to aerobic metabolism is correlated with the stage of development, then carbon monoxide exposure should not represent any significant risk to the human embryo until later in gestation.

  10. Cryotop vitrification for in vitro produced bovine and buffalo (Bubalus bubalis embryos at different stages of development

    Directory of Open Access Journals (Sweden)

    B. Gasparrini

    2010-02-01

    Full Text Available The aim of this study was to evaluate the possibility to vitrify in vitro produced (IVP buffalo and bovine embryos at different stages of development by an advanced version of the “minimal volume approaches”: the Cryotop method. In both experiments, the embryos were vitrified at the tight morula (TM, early blastocyst (eBl, blastocyst (Bl, expanded blastocyst (xBl and, only for buffalo, at the hatched blastocyst (hBl stage. After warming, the embryos were cultured in vitro for 24 hours. Stage of development affected the freezability of IVP embryos of both species with the highest embryo survival rates at advanced stages (xBl=76% and hBl=75% for buffalos and xBl=75% for bovine. These results suggest that Cryotop vitrification is an efficient method for buffalo and bovine IVP embryo cryopreservation.

  11. ADAM10 Is Involved in Cell Junction Assembly in Early Porcine Embryo Development.

    Directory of Open Access Journals (Sweden)

    Jeongwoo Kwon

    Full Text Available ADAM10 (A Disintegrin and Metalloprotease domain-containing protein 10 is a cell surface protein with a unique structure possessing both potential adhesion and protease domains. However, the role of ADAM10 in preimplantation stage embryos is not clear. In this study, we examined the expression patterns and functional roles of ADAM10 in porcine parthenotes during preimplantation development. The transcription level of ADAM10 dramatically increased from the morula stage onward. Immunostaining revealed that ADAM10 was present in both the nucleus and cytoplasm in early cleavage stage embryos, and localized to the apical region of the outer cells in morula and blastocyst embryos. Knockdown (KD of ADAM10 using double strand RNA did not alter preimplantation embryo development until morula stage, but resulted in significantly reduced development to blastocyst stage. Moreover, the KD blastocyst showed a decrease in gene expression of adherens and tight junction (AJ/TJ, and an increase in trophectoderm TJ permeability by disrupting TJ assembly. Treatment with an ADAM10 specific chemical inhibitor, GI254023X, at the morula stage also inhibited blastocyst development and led to disruption of TJ assembly. An in situ proximity ligation assay demonstrated direct interaction of ADAM10 with coxsackie virus and adenovirus receptor (CXADR, supporting the involvement of ADAM10 in TJ assembly. In conclusion, our findings strongly suggest that ADADM10 is important for blastocyst formation rather than compaction, particularly for TJ assembly and stabilization in preimplantation porcine parthenogenetic development.

  12. Developing Xenopus Embryos Recover by Compacting and Expelling Single-Wall Carbon Nanotubes

    Science.gov (United States)

    Holt, Brian D.; Shawky, Joseph H.; Dahl, Kris Noel; Davidson, Lance A.; Islam, Mohammad F.

    2015-01-01

    Single-wall carbon nanotubes are high aspect ratio nanomaterials that are being developed for use in materials, technological and biological applications due to their high mechanical stiffness, optical properties, and chemical inertness. Because of their prevalence, it is inevitable that biological systems will be exposed to nanotubes, yet studies of the effects of nanotubes on developing embryos have been inconclusive and are lacking for single-wall carbon nanotubes exposed to the widely studied model organism Xenopus laevis (African clawed frog). Microinjection of experimental substances into the Xenopus embryo is a standard technique for toxicology studies and cellular lineage tracing. Here we report the surprising finding that superficial (12.5 ± 7.5 μm below the membrane) microinjection of nanotubes dispersed with Pluronic F127 into one-to-two cell Xenopus embryos resulted in the formation and expulsion of compacted, nanotube-filled, punctate masses, at the blastula to mid-gastrula developmental stages, which we call “boluses”. Such expulsion of microinjected materials by Xenopus embryos has not been reported before and is dramatically different from the typical distribution of the materials throughout the progeny of the microinjected cells. Previous studies of microinjections of nanomaterials such as nanodiamonds, quantum dots or spherical nanoparticles report that nanomaterials often induce toxicity and remain localized within the embryos. In contrast, our results demonstrate an active recovery pathway for embryos after exposure to Pluronic F127-coated nanotubes, which we speculate is due to a combined effect of the membrane activity of the dispersing agent, Pluronic F127, and the large aspect ratio of nanotubes. PMID:26153061

  13. [Embryo development in two single-step media: Analysis of 2059 sibling oocytes].

    Science.gov (United States)

    Durand, M; Sermondade, N; Herbemont, C; Benard, J; Gronier, H; Boujenah, J; Cédrin-Durnerin, I; Poncelet, C; Grynberg, M; Sifer, C

    2016-03-01

    The aim of this study was to compare embryo development cultured in two single-step media commercially available: Fert/Sage One Step® (Origio) and Continuous Single Culture® (CSC) (Irvine Scientific). A prospective auto-controlled study of sibling oocytes from women undergoing conventional in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) was performed in our center from February to June 2015. After fertilization, for every patient, half of oocytes were cultured in the single-step Fert/Sage One Step® (serie SAGE) and the other half in the single-step CSC®(serie CSC). Fertilization and embryo morphology rates were assessed by day 1 to day 5-6 if needed. Embryo presentingtwo attempts of IVF and 133 of ICSI were analyzed, corresponding to 2059 inseminated or micro-injected oocytes. Fertilization rate were not different between the 2 series, respectively SAGE vs CSC (IVF: 73.4% vs 68.3% [P=0.49]; ICSI: 58.9% vs 63.8% [P=0.12]). No difference was found for embryo morphology, respectively SAGE vs CSC, at day 2 (top quality embryo at day 2 IVF: 34.4% vs 33% [P=0.98]; ICSI: 42.4% vs 44.9% [P=0.37]; and good quality embryo at day 2 IVF: 44% vs 50.2% [P=0.07]; ICSI: 64% vs 71% [P=0.35]); no difference at day 3 (top quality embryo at day 3 IVF: 19.4% vs 21.3% [P=0.61]; ICSI: 28.7% vs 27.4% [P=0.54]; and good quality embryo at day 3 IVF: 40.4% vs 50.2% [P=0.91]; ICSI: 51% vs 47.6% [P=0.47]). Blastocyst development rate were not different, respectively SAGE vs CSC (IVF: 39.9% vs 41.5% [P=0.63] with 42.9% vs 42.2% of good quality blastocyst [P=0.70]; ICSI: 41.1% vs 37.8% [P=0.18] with 32.9% vs 40.8% of good quality blastocyst [P=0.13]). No difference was found in the useful embryo rate in the 2 series SAGE vs CSC (IVF: 52.8% vs 55.2% [P=0.83]; ICSI: 62.4% vs 61.7% [P=0.70]). Embryo development and rate of useful embryos, transferred or frozen, were not different according to the embryo culture in single-step media Fert/Sage One Step® vs single

  14. [A computer-assisted workplace for anatomical pathology investigations on human embryos].

    Science.gov (United States)

    Fahr, M S; Müller, A M; Müntefering, H; Coerdt, W

    2008-07-01

    Contrary to chromosomal aberrations, which can be recognized by cytogenetic procedures alone, monogenic inherited diseases are determined exclusively by evidence from anatomical-pathological investigations. We present a computer-assisted optical system providing not only efficient dissections of embryos, but also diagnosis of congenital defects, such as congenital heart deformities, neural tube defects and skeletal malformations. A stereomicroscope with an integrated camera as well as two cold light sources creates a three-dimensional image of the human embryo (size: e.g., 2.5 mm=23.-25.d), hence facilitating handling of the autopsy. Scenes of interest are photodocumented by a multifocusing camera. Its technique is based on serial pictures of predefined levels of the embryo, consecutively adding up to one photograph with minimized areas out of focus. The sequences, the rapid as well as exact calibration of the screened objects and digital archiving of the obtained photographs allow efficient diagnostic procedures. As the depth of field is broadened, the computer-assisted workplace improves the diagnosis as well as documentation, providing a base for genetic counseling.

  15. Nerve growth factor regulates axial rotation during early stages of chick embryo development.

    Science.gov (United States)

    Manca, Annalisa; Capsoni, Simona; Di Luzio, Anna; Vignone, Domenico; Malerba, Francesca; Paoletti, Francesca; Brandi, Rossella; Arisi, Ivan; Cattaneo, Antonino; Levi-Montalcini, Rita

    2012-02-07

    Nerve growth factor (NGF) was discovered because of its neurotrophic actions on sympathetic and sensory neurons in the developing chicken embryo. NGF was subsequently found to influence and regulate the function of many neuronal and non neuronal cells in adult organisms. Little is known, however, about the possible actions of NGF during early embryonic stages. However, mRNAs encoding for NGF and its receptors TrkA and p75(NTR) are expressed at very early stages of avian embryo development, before the nervous system is formed. The question, therefore, arises as to what might be the functions of NGF in early chicken embryo development, before its well-established actions on the developing sympathetic and sensory neurons. To investigate possible roles of NGF in the earliest stages of development, stage HH 11-12 chicken embryos were injected with an anti-NGF antibody (mAb αD11) that binds mature NGF with high affinity. Treatment with anti-NGF, but not with a control antibody, led to a dose-dependent inversion of the direction of axial rotation. This effect of altered rotation after anti NGF injection was associated with an increased cell death in somites. Concurrently, a microarray mRNA expression analysis revealed that NGF neutralization affects the expression of genes linked to the regulation of development or cell proliferation. These results reveal a role for NGF in early chicken embryo development and, in particular, in the regulation of somite survival and axial rotation, a crucial developmental process linked to left-right asymmetry specification.

  16. Potential teratogenicity of methimazole: exposure of zebrafish embryos to methimazole causes similar developmental anomalies to human methimazole embryopathy.

    Science.gov (United States)

    Komoike, Yuta; Matsuoka, Masato; Kosaki, Kenjiro

    2013-06-01

    While methimazole (MMI) is widely used in the therapy for hyperthyroidism, several groups have reported that maternal exposure to MMI results in a variety of congenital anomalies, including choanal and esophageal atresia, iridic and retinal coloboma, and delayed neurodevelopment. Thus, adverse effects of maternal exposure to MMI on fetal development have long been suggested; however, direct evidence for the teratogenicity of MMI has not been presented. Therefore, we studied the effects of MMI on early development by using zebrafish as a model organism. The fertilized eggs of zebrafish were collected immediately after spawning and grown in egg culture water containing MMI at various concentrations. External observation of the embryos revealed that exposure to high concentrations of MMI resulted in loss of pigmentation, hypoplastic hindbrain, turbid tissue in the forebrain, swelling of the notochord, and curly trunk. Furthermore, these effects occurred in a dose-dependent manner. Precise observation of the serial cross-sections of MMI-exposed embryos elucidated delayed development and hypoplasia of the whole brain and spinal cord, narrowing of the pharynx and esophagus, severe disruption of the retina, and aberrant structure of the notochord. These neuronal, pharyngeal, esophageal, and retinal anomalous morphologies have a direct analogy to the congenital anomalies observed in children exposed to MMI in utero. Here, we show the teratogenic effects of MMI on the development of zebrafish and provide the first experimental evidence for the connection between exposure to MMI and human MMI embryopathy. © 2013 Wiley Periodicals, Inc.

  17. A technique for sexing fully developed embryos and early-instar larvae of the gypsy moth

    Science.gov (United States)

    Gilbert Levesque

    1963-01-01

    Because variation in sex ratio is an important factor in the population dynamics of the gypsy moth (Porthetria dispar), it is necessary to have some means of determining the ratio of males to females in a population at the beginning of the larval period as well as in the later stages. For determining the sex of fully developed embryos and early-...

  18. Comparing carbohydrate status during norway spruce seed development and somatic embryo formation

    NARCIS (Netherlands)

    Gösslová, M.; Svobodová, H.; Lipavská, H.; Albrechtová, J.; Vreugdenhil, D.

    2001-01-01

    The carbohydrate status of developing seeds of Picea abies was examined in order to provide a frame of reference for the evaluation of changes in carbohydrate content in maturing somatic embryos of the same species. Samples were taken at weekly intervals from 12 May 1998 (estimated time of

  19. GSM 900 MHz microwave radiation affects embryo development of Japanese quails.

    Science.gov (United States)

    Tsybulin, Olexandr; Sidorik, Evgeniy; Kyrylenko, Sergiy; Henshel, Diane; Yakymenko, Igor

    2012-03-01

    A wide range of non thermal biological effects of microwave radiation (MW) was revealed during the last decades. A number of reports showed evident hazardous effects of MW on embryo development in chicken. In this study, we aimed at elucidating the effects of MW emitted by a commercial model of GSM 900 MHz cell phone on embryo development in quails (Coturnix coturnix japonica) during both short and prolonged exposure. For that, fresh fertilized eggs were irradiated during the first 38 h or 14 days of incubation by a cell phone in "connecting" mode activated continuously through a computer system. Maximum intensity of incident radiation on the egg's surface was 0.2 μW/cm2.The irradiation led to a significant (pGSM 900 MHz cell phone on developing quail embryos signify a possibility for non-thermal impact of MW on embryogenesis. We suggest that the facilitating effect of low doses of irradiation on embryo development can be explained by a hormesis effect induced by reactive oxygen species (ROS). Future studies need to be done to clarify this assumption.

  20. Acquisition of desiccation tolerance in developing wheat embryos correlates with appearance of a fluid phase in membranes

    NARCIS (Netherlands)

    Golovina, E.A.; Hoekstra, F.A.

    2003-01-01

    Membrane behaviour in developing wheat (Triticum aestivum cv Priokskaya) embryos was studied in relation to the acquisition of desiccation tolerance, using spin probe techniques. Fresh embryos were able to develop into seedlings at day 15 after anthesis, but it took 18 d before fast-dried, isolated

  1. A comparison of anterior-posterior development in the porcine versus the chicken embryo, using goosecoid expression as a marker

    NARCIS (Netherlands)

    Pavert, van de S.A.; Schipper, H.; Wit, de A.A.C.; Soede, N.M.; Hurk, van den R.; Taverne, M.A.M.; Boerjan, M.L.; Stroband, H.W.J.

    2001-01-01

    During early embryonic development, pig and chicken embryos share striking morphological similarities. In the present study, the timing and location of expression of mRNA for goosecoid (gsc), a gene classically expressed in the nodal region of developing embryos, was examined and compared in

  2. Transcriptional Innate Immune Response of the Developing Chicken Embryo to Newcastle Disease Virus Infection

    Directory of Open Access Journals (Sweden)

    Megan A. Schilling

    2018-02-01

    Full Text Available Traditional approaches to assess the immune response of chickens to infection are through animal trials, which are expensive, require enhanced biosecurity, compromise welfare, and are frequently influenced by confounding variables. Since the chicken embryo becomes immunocompetent prior to hatch, we here characterized the transcriptional response of selected innate immune genes to Newcastle disease virus (NDV infection in chicken embryos at days 10, 14, and 18 of embryonic development. The results suggest that the innate immune response 72 h after challenge of 18-day chicken embryo is both consistent and robust. The expression of CCL5, Mx1, and TLR3 in lung tissues of NDV challenged chicken embryos from the outbred Kuroiler and Tanzanian local ecotype lines showed that their expression was several orders of magnitude higher in the Kuroiler than in the local ecotypes. Next, the expression patterns of three additional innate-immunity related genes, IL-8, IRF-1, and STAT1, were examined in the highly congenic Fayoumi (M5.1 and M15.2 and Leghorn (Ghs6 and Ghs13 sublines that differ only at the microchromosome bearing the major histocompatibility locus. The results show that the Ghs13 Leghorn subline had a consistently higher expression of all genes except IL-8 and expression seemed to be subline-dependent rather than breed-dependent, suggesting that the innate immune response of chicken embryos to NDV infection may be genetically controlled by the MHC-locus. Taken together, the results suggest that the chicken embryo may represent a promising model to studying the patterns and sources of variation of the avian innate immune response to infection with NDV and related pathogens.

  3. KIF20A regulates porcine oocyte maturation and early embryo development.

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    Full Text Available KIF20A (Kinesin-like family member 20A, also called mitotic kinesin-like proteins 2 (MKLP2, is a mammalian mitotic kinesin-like motor protein of the Kinesin superfamily proteins (KIFs, which was originally involved in Golgi apparatus dynamics and thought to essential for cell cycle regulation during successful cytokinesis. In the present study, we investigated whether KIF20A has roles on porcine oocyte meiotic maturation and subsequent early embryo development. By immunofluorescence staining, KIF20A was found to exhibit a dynamic localization pattern during meiosis. KIF20A was restricted to centromeres after germinal vesicle breakdown (GVBD, transferred to the midbody at telophase I (TI, and again associated with centromeres at metaphase II (MII. Inhibition of endogenous KIF20A via a specific inhibitor, Paprotrain, resulted in failure of polar body extrusion. Further cell cycle analysis showed that the percentage of oocytes that arrested at early metaphase I (MI stage increased after KIF20A activity inhibition; however, the proportion of oocytes at anaphase/telophase I (ATI and MII stages decreased significantly. Our results also showed that KIF20A inhibition did not affect spindle morphology. In addition, KIF20A was localized at the nucleus of early embryos, and KIF20A inhibition resulted in failure of early parthenogenetic embryo development. These results demonstrated that KIF20A is critical for porcine oocyte meiotic maturation and subsequent early embryo development.

  4. Inclusion of bovine lipoproteins and the vitamin E analogue, Trolox, during in vitro culture of bovine embryos changes both embryo and fetal development.

    Science.gov (United States)

    Rooke, J A; Watt, R G; Ashworth, C J; McEvoy, T G

    2012-01-01

    This experiment investigated effects of lipoproteins and Trolox (vitamin E analogue) on bovine embryo and fetal development. The treatments were: in vitro culture (IVC) in synthetic oviducal fluid alone (SOF); with bovine lipoproteins (2% v/v; SOFLP); with Trolox (100μM; SOFT); and with lipoproteins and Trolox (SOFLPT). In vitro culture with lipoproteins increased fatty acid content of blastocysts (PTrolox had no effect (P>0.05). Whereas lipoproteins reduced zygote development to blastocysts (P=0.03), Trolox facilitated increased development (PTrolox (P>0.05) had no effect on blastocyst morphological grade. Pregnancy rates resulting from synchronous transfer of IVP embryos were not affected by IVC treatment. At Day 70 of pregnancy, compared with SOF, fetal weight was lower in SOFLP but not SOFLPT (interaction, PTrolox. Placentome numbers were greater in SOF and SOFLPT compared with SOFLP and SOFT (interaction, P=0.002); superior embryo grades were also associated with increased numbers of placentomes (P=0.024). In conclusion, the interactive effects of lipoprotein and Trolox inclusion on in vitro embryo development were also evident in fetal development at Day 70.

  5. In vitro development of embryos from experimentally Kerack-addicted Mice

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    Elham Mohammadzadeh

    2017-08-01

    Full Text Available Background: Prenatal drug exposure, as a common public health concern, is associated with an increased risk of adverse effects on early embryo development. Objective: To investigate the in vitro development of - embryo from experimentally Kerack-addicted mice. Materials and Methods: Twenty-five female mice were studied in five groups: control, vehicle, and three experimental groups of Kerack-dependent mice (I, II, and III which received different doses of Kerack for 14 days. After the establishment of addiction model (7 days, experimental groups I, II, and III were given Kerack intraperitoneally at the doses of 5, 35, and 70 mg/kg, twice a day for a period of 7 days, respectively. The vehicle group received normal saline and lemon juice whilst the control group just received water and food. Morulae were obtained through oviduct flashing. The survived embryos were cultured in T6+ 5mg/ml bovine serum albumin. The developmental rates up to hatched stage daily and embryo quality (differential staining and Tunnel staining were also assessed Results: The developmental potential of embryos obtained from the addicted mother was significantly decreased in comparison with control group. There was a significant reduction in the rate of blastocyst formation in the high dose Kerack dependent group. However, in addicted mice there was reduction in the total cell number (40.92% vs. 65.08% in control and, inner cell mass percentage (17.17% vs. 26.15% in control while apoptotic cells numbers were increased (7.17 vs. 1.46 in control (p<0.05. Conclusion: The Kerack addiction during pregnancy retards preimplantation development and induces apoptosis.

  6. Changes in the dielectric properties of medaka fish embryos during development, studied by electrorotation

    Energy Technology Data Exchange (ETDEWEB)

    Shirakashi, Ryo, E-mail: aa21150@iis.u-tokyo.ac.jp [Institute of Industrial Science, The University of Tokyo, Tokyo 153-8505 (Japan); Mischke, Miriam [Lehrstuhl fuer Biotechnologie und Biophysik, Biozentrum, Universitaet Wuerzburg, Wuerzburg (Germany); Fischer, Peter [Physiologische Chemie, Biozentrum, Universitaet Wuerzburg, Wuerzburg (Germany); Memmel, Simon [Lehrstuhl fuer Biotechnologie und Biophysik, Biozentrum, Universitaet Wuerzburg, Wuerzburg (Germany); Krohne, Georg [Abteilung fuer Elektronenmikroskopie, Biozentrum, Universitaet Wuerzburg, Wuerzburg (Germany); Fuhr, Guenter R. [Lehrstuhl fuer Biotechnologie und Medizintechnik, Universitaet des Saarlandes, Saarbruecken (Germany); Zimmermann, Heiko [Lehrstuhl fuer Molekulare und Zellulaere Biotechnologie, Universitaet des Saarlandes, Saarbruecken (Germany); Sukhorukov, Vladimir L., E-mail: sukhorukov@biozentrum.uni-wuerzburg.de [Lehrstuhl fuer Biotechnologie und Biophysik, Biozentrum, Universitaet Wuerzburg, Wuerzburg (Germany)

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer Electrorotation offers a non-invasive tool for dielectric analysis of fish embryos. Black-Right-Pointing-Pointer The three-shell dielectric model matches the rotation spectra of medaka eggs. Black-Right-Pointing-Pointer The capacitance value suggests a double-membrane structure of yolk envelope. -- Abstract: The Japanese medaka fish, Oryzias latipes, has become a powerful vertebrate model organism in developmental biology and genetics. The present study explores the dielectric properties of medaka embryos during pre-hatching development by means of the electrorotation (ROT) technique. Due to their layered structure, medaka eggs exhibited up to three ROT peaks in the kHz-MHz frequency range. During development from blastula to early somite stage, ROT spectra varied only slightly. But as the embryo progressed to the late-somite stage, the ROT peaks underwent significant changes in frequency and amplitude. Using morphological data obtained by light and electron microscopy, we analyzed the ROT spectra with a three-shell dielectric model that accounted for the major embryonic compartments. The analysis yielded a very high value for the ionic conductivity of the egg shell (chorion), which was confirmed by independent osmotic experiments. A relatively low capacitance of the yolk envelope was consistent with its double-membrane structure revealed by transmission electron microscopy. Yolk-free dead eggs exhibited only one co-field ROT peak, shifted markedly to lower frequencies with respect to the corresponding peak of live embryos. The dielectric data may be useful for monitoring the development and changes in fish embryos' viability/conditions in basic research and industrial aquaculture.

  7. Are human embryos Kantian persons?: Kantian considerations in favor of embryonic stem cell research.

    Science.gov (United States)

    Manninen, Bertha Alvarez

    2008-01-31

    One argument used by detractors of human embryonic stem cell research (hESCR) invokes Kant's formula of humanity, which proscribes treating persons solely as a means to an end, rather than as ends in themselves. According to Fuat S. Oduncu, for example, adhering to this imperative entails that human embryos should not be disaggregated to obtain pluripotent stem cells for hESCR. Given that human embryos are Kantian persons from the time of their conception, killing them to obtain their cells for research fails to treat them as ends in themselves. This argument assumes two points that are rather contentious given a Kantian framework. First, the argument assumes that when Kant maintains that humanity must be treated as an end in itself, he means to argue that all members of the species Homo sapiens must be treated as ends in themselves; that is, that Kant regards personhood as co-extensive with belonging to the species Homo sapiens. Second, the argument assumes that the event of conception is causally responsible for the genesis of a Kantian person and that, therefore, an embryo is a Kantian person from the time of its conception. In this paper, I will present challenges against these two assumptions by engaging in an exegetical study of some of Kant's works. First, I will illustrate that Kant did not use the term "humanity" to denote a biological species, but rather the capacity to set ends according to reason. Second, I will illustrate that it is difficult given a Kantian framework to denote conception (indeed any biological event) as causally responsible for the creation of a person. Kant ascribed to a dualistic view of human agency, and personhood, according to him, was derived from the supersensible capacity for reason. To argue that a Kantian person is generated due to the event of conception ignores Kant's insistence in various aspects of his work that it is not possible to understand the generation of a person qua a physical operation. Finally, I will end the

  8. Are human embryos Kantian persons?: Kantian considerations in favor of embryonic stem cell research

    Directory of Open Access Journals (Sweden)

    Manninen Bertha

    2008-01-01

    Full Text Available Abstract One argument used by detractors of human embryonic stem cell research (hESCR invokes Kant's formula of humanity, which proscribes treating persons solely as a means to an end, rather than as ends in themselves. According to Fuat S. Oduncu, for example, adhering to this imperative entails that human embryos should not be disaggregated to obtain pluripotent stem cells for hESCR. Given that human embryos are Kantian persons from the time of their conception, killing them to obtain their cells for research fails to treat them as ends in themselves. This argument assumes two points that are rather contentious given a Kantian framework. First, the argument assumes that when Kant maintains that humanity must be treated as an end in itself, he means to argue that all members of the species Homo sapiens must be treated as ends in themselves; that is, that Kant regards personhood as co-extensive with belonging to the species Homo sapiens. Second, the argument assumes that the event of conception is causally responsible for the genesis of a Kantian person and that, therefore, an embryo is a Kantian person from the time of its conception. In this paper, I will present challenges against these two assumptions by engaging in an exegetical study of some of Kant's works. First, I will illustrate that Kant did not use the term "humanity" to denote a biological species, but rather the capacity to set ends according to reason. Second, I will illustrate that it is difficult given a Kantian framework to denote conception (indeed any biological event as causally responsible for the creation of a person. Kant ascribed to a dualistic view of human agency, and personhood, according to him, was derived from the supersensible capacity for reason. To argue that a Kantian person is generated due to the event of conception ignores Kant's insistence in various aspects of his work that it is not possible to understand the generation of a person qua a physical

  9. Three-Dimensional High-Frequency Ultrasonography for Early Detection and Characterization of Embryo Implantation Site Development in the Mouse.

    Directory of Open Access Journals (Sweden)

    Mary C Peavey

    Full Text Available Ultrasonography is a powerful tool to non-invasively monitor in real time the development of the human fetus in utero. Although genetically engineered mice have served as valuable in vivo models to study both embryo implantation and pregnancy progression, such studies usually require sacrifice of parous mice for subsequent phenotypic analysis. To address this issue, we used three-dimensional (3-D reconstruction in silico of high-frequency ultrasound (HFUS imaging data for early detection and characterization of murine embryo implantation sites and their development in utero. With HFUS imaging followed by 3-D reconstruction, we were able to precisely quantify embryo implantation site number and embryonic developmental progression in pregnant C57BL6J/129S mice from as early as 5.5 days post coitus (d.p.c. through to 9.5 d.p.c. using a VisualSonics Vevo 2100 (MS550S transducer. In addition to measurements of implantation site number, location, volume and spacing, embryo viability via cardiac activity monitoring was also achieved. A total of 12 dams were imaged with HFUS with approximately 100 embryos examined per embryonic day. For the post-implantation period (5.5 to 8.5 d.p.c., 3-D reconstruction of the gravid uterus in mesh or solid overlay format enabled visual representation in silico of implantation site location, number, spacing distances, and site volume within each uterine horn. Therefore, this short technical report describes the feasibility of using 3-D HFUS imaging for early detection and analysis of post-implantation events in the pregnant mouse with the ability to longitudinally monitor the development of these early pregnancy events in a non-invasive manner. As genetically engineered mice continue to be used to characterize female reproductive phenotypes, we believe this reliable and non-invasive method to detect, quantify, and characterize early implantation events will prove to be an invaluable investigative tool for the study of

  10. Pollen Viability, Pistil Receptivity, and Embryo Development in Hybridization of Nelumbo nucifera Gaertn

    Directory of Open Access Journals (Sweden)

    Yan-Li Wang

    2012-01-01

    Full Text Available Seed set is usually low and differs for different crosses of flower lotus (Nelumbo nucifera Gaertn.. The reasons remain unknown, and this has a negative impact on lotus breeding. To determine the causes, we carried out two crosses of flower lotus, that is, “Jinsenianhua” × “Qinhuaihuadeng” and “Qinhuaihuadeng” × “Jinsenianhua” and pollen viability, pistil receptivity, and embryo development were investigated. The pollen grains collected at 05:00-06:00 hrs had the highest viability, and the viabilities of “Jinsenianhua” and “Qinhuaihuadeng” were 20.6 and 15.7%, respectively. At 4 h after artificial pollination, the number of pollen grains germinating on each stigma reached a peak: 63.0 and 17.2 per stigma in “Jinsenianhua” × “Qinhuaihuadeng” and “Qinhuaihuadeng” × “Jinsenianhua”, respectively. At 1 d after artificial pollination, the percentages of normal embryos in the two crosses were 55.0 and 21.9%, respectively; however, at 11 d after pollination, the corresponding percentages were 20.8 and 11.2%. Seed sets of the two crosses were 17.9 and 8.0%, respectively. The results suggested that low pistil receptivity and embryo abortion caused low seed set in “Qinhuaihuadeng” × “Jinsenianhua”, whereas low fecundity of “Jinsenianhua” × “Qinhuaihuadeng” was mainly attributable to embryo abortion.

  11. Bone morphogenetic protein 2 signaling negatively modulates lymphatic development in vertebrate embryos

    DEFF Research Database (Denmark)

    Dunworth, William P; Cardona-Costa, Jose; Bozkulak, Esra Cagavi

    2014-01-01

    RATIONALE: The emergence of lymphatic endothelial cells (LECs) seems to be highly regulated during development. Although several factors that promote the differentiation of LECs in embryonic development have been identified, those that negatively regulate this process are largely unknown. OBJECTIVE...... signaling in zebrafish embryos and mouse embryonic stem cell-derived embryoid bodies substantially decrease the emergence of LECs. Mechanistically, BMP2 signaling induces expression of miR-31 and miR-181a in a SMAD-dependent mechanism, which in turn results in attenuated expression of prospero homeobox......: Our aim was to delineate the role of bone morphogenetic protein (BMP) 2 signaling in lymphatic development. METHODS AND RESULTS: BMP2 signaling negatively regulates the formation of LECs. Developing LECs lack any detectable BMP signaling activity in both zebrafish and mouse embryos, and excess BMP2...

  12. Expression profiling identifies novel Hh/Gli-regulated genes in developing zebrafish embryos.

    Science.gov (United States)

    Bergeron, Sadie A; Milla, Luis A; Villegas, Rosario; Shen, Meng-Chieh; Burgess, Shawn M; Allende, Miguel L; Karlstrom, Rolf O; Palma, Verónica

    2008-02-01

    The Hedgehog (Hh) signaling pathway plays critical instructional roles during embryonic development. Misregulation of Hh/Gli signaling is a major causative factor in human congenital disorders and in a variety of cancers. The zebrafish is a powerful genetic model for the study of Hh signaling during embryogenesis, as a large number of mutants that affect different components of the Hh/Gli signaling system have been identified. By performing global profiling of gene expression in different Hh/Gli gain- and loss-of-function scenarios we identified known (e.g., ptc1 and nkx2.2a) and novel Hh-regulated genes that are differentially expressed in embryos with altered Hh/Gli signaling function. By uncovering changes in tissue-specific gene expression, we revealed new embryological processes that are influenced by Hh signaling. We thus provide a comprehensive survey of Hh/Gli-regulated genes during embryogenesis and we identify new Hh-regulated genes that may be targets of misregulation during tumorigenesis.

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

    Science.gov (United States)

    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.

  14. Putrescine induces somatic embryo development and proteomic changes in embryogenic callus of sugarcane.

    Science.gov (United States)

    Reis, Ricardo Souza; Vale, Ellen de Moura; Heringer, Angelo Schuabb; Santa-Catarina, Claudete; Silveira, Vanildo

    2016-01-01

    Somatic embryogenesis, an important biotechnological technique, has great potential for application in sugarcane breeding and micropropagation. Polyamines have been associated with the regulation of several physiological processes, including the acquisition of embryogenic competence and somatic embryogenesis. In this study, we used a proteomic approach to evaluate the effects of exogenous polyamine on sugarcane somatic embryo development to better understand this process. Embryogenic cultures were treated with different concentrations of putrescine, spermidine, and spermine. Proteomic analyses combined the shotgun method and the nanoESI-HDMS(E) technology. Among polyamines, 500 μM putrescine gave rise to the highest number of somatic embryos; however, no differences in the amount of fresh matter were observed between polyamines and control. Differences in protein abundance profiles resulting from the effect of 500 μM putrescine on sugarcane somatic embryo maturation were observed. Proteomic analyses of putrescine and control treatment showed differences in the abundances of proteins related to somatic embryogenesis, such as arabinogalactan proteins, peroxidases, heat shock proteins, glutathione s-transferases, late embryogenesis abundant proteins, and 14-3-3 proteins. These results show that putrescine and the identified proteins play important roles in protecting the cells against an in vitro stress environment, contributing to the formation of somatic embryos during the maturation treatment. Despite all studies with somatic embryogenesis, the molecular mechanisms controlling the process have not been completely understood. In this study, we highlighted the effects of the polyamine putrescine on somatic embryogenesis of sugarcane and the differentially abundant proteins related to somatic embryo development. We identified six groups of important stress related proteins that are involved in the adaptation of cells to the stress environment of in vitro culture and

  15. Contrasting Storage Protein Synthesis and Messenger RNA Accumulation during Development of Zygotic and Somatic Embryos of Alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Krochko, J E; Pramanik, S K; Bewley, J D

    1992-05-01

    During development on hormone-free media, somatic embryos pass through distinct morphological stages that superficially resemble those of zygotic embryo development (globular, heart, torpedo, cotyledonary stages). Despite these similarities, they differ from zygotic embryos in the extent of cotyledonary development and the patterns of synthesis and quantitative expression of seed-specific storage proteins (7S, 11S, and 2S proteins). Alfin (7S) is the first storage protein synthesized in developing zygotic embryos (stage IV). The 11S (medicagin) and 2S (Low Molecular Weight, LMW) storage proteins are not detectable until the following stage of development (stage V), although all three are present before the completion of embryo enlargement. Likewise, the 7S storage protein is the first to be synthesized in developing somatic embryos (day 5). Medicagin is evident by day 7 and the LMW protein by day 10. In contrast to zygotic embryos, alfin remains the predominant storage protein in somatic embryos throughout development. Not only are the relative amounts of medicagin and the LMW protein reduced in somatic embryos but the LMW protein is accumulated much later than the other proteins. Quantification of the storage protein mRNAs (7S, 11S, and 2S) by northern blot analysis confirms that there are substantial differences in the patterns of message accumulation in zygotic and somatic embryos of alfalfa (Medicago sativa). In zygotic embryos, the 7S, 11S, and 2S storage protein mRNAs are abundant during maturation and, in particular, during the stages of maximum protein synthesis (alfin, stages VI and VII; medicagin, stage VII; LMW, stage VII). In somatic embryos, the predominance of the 7S storage protein is correlated with increased accumulation of its mRNA, whereas the limited synthesis of the 11S storage protein is associated with much lower steady-state levels of its message. The mRNA for the LMW protein is present already by 3 days after transfer to hormone-free media

  16. Imidacloprid Exposure Suppresses Neural Crest Cells Generation during Early Chick Embryo Development.

    Science.gov (United States)

    Wang, Chao-Jie; Wang, Guang; Wang, Xiao-Yu; Liu, Meng; Chuai, Manli; Lee, Kenneth Ka Ho; He, Xiao-Song; Lu, Da-Xiang; Yang, Xuesong

    2016-06-15

    Imidacloprid is a neonicotinoid pesticide that is widely used in the control pests found on crops and fleas on pets. However, it is still unclear whether imidacloprid exposure could affect early embryo development-despite some studies having been conducted on the gametes. In this study, we demonstrated that imidacloprid exposure could lead to abnormal craniofacial osteogenesis in the developing chick embryo. Cranial neural crest cells (NCCs) are the progenitor cells of the chick cranial skull. We found that the imidacloprid exposure retards the development of gastrulating chick embryos. HNK-1, PAX7, and Ap-2α immunohistological stainings indicated that cranial NCCs generation was inhibited after imidacloprid exposure. Double immunofluorescent staining (Ap-2α and PHIS3 or PAX7 and c-Caspase3) revealed that imidacloprid exposure inhibited both NCC proliferation and apoptosis. In addition, it inhibited NCCs production by repressing Msx1 and BMP4 expression in the developing neural tube and by altering expression of EMT-related adhesion molecules (Cad6B, E-Cadherin, and N-cadherin) in the developing neural crests. We also determined that imidacloprid exposure suppressed cranial NCCs migration and their ability to differentiate. In sum, we have provided experimental evidence that imidacloprid exposure during embryogenesis disrupts NCCs development, which in turn causes defective cranial bone development.

  17. Regulation of Cardiovascular Development by Adenosine and Adenosine-Mediated Embryo Protection

    OpenAIRE

    Rivkees, Scott A; Wendler, Christopher C.

    2012-01-01

    Few signaling molecules have the potential to influence the developing mammal as the nucleoside adenosine. Adenosine levels increase rapidly with tissue hypoxia and inflammation. Adenosine antagonists include the methlyxanthines caffeine and theophylline. The receptors that transduce adenosine action are the A1, A2a, A2b, and A3 adenosine receptors (ARs). We examined how adenosine acts via A1ARs to influence embryo development.

  18. Plasma membrane block to polyspermy in human oocytes and preimplantation embryos.

    Science.gov (United States)

    Sengoku, K; Tamate, K; Horikawa, M; Takaoka, Y; Ishikawa, M; Dukelow, W R

    1995-09-01

    The existence and time course of the human plasma membrane block to polyspermy were investigated by an in vitro fertilization assay using zona pellucida-free unfertilized oocytes, pronuclear oocytes and embryos. In the time course study using a high sperm concentration (10(5) spermatozoa ml-1), the number of penetrating spermatozoa at 30 and 60 min after insemination were 1.3 +/- 0.3 and 2.9 +/- 0.4, respectively. By 2 h after insemination, spermatozoa penetration reached a maximum. A lower maximum number of penetrating spermatozoa was observed at a low sperm concentration (10(4) spermatozoa ml-1), but the number of penetrating spermatozoa still reached a maximum by 2 h after insemination. A reinsemination experiment demonstrated that the number of penetrating spermatozoa was not significantly different between control and reinseminated oocytes, while sperm penetration was not observed in the oocyte beyond the two-cell stage. Furthermore, the number of binding spermatozoa decreased after fertilization and most of the four-cell stage embryos displayed no sperm binding. These results suggest that the plasma membrane block plays an important role in the prevention of polyspermy in the human oocyte, and that the plasma membrane block may involve permanent changes in the binding or fusion ability of spermatozoa in the oolemma after fertilization.

  19. Changes in water status and proline and abscisic acid concentrations in developing somatic embryos of pedunculate oak (Quercus robur) during maturation and germination.

    Science.gov (United States)

    Prewein, Christine; Vagner, Martin; Wilhelm, Eva

    2004-11-01

    Somatic embryos of oak (Quercus robur L.) were matured on P24 media differing in gel strength (0.8, 0.9 and 1.0% (w/v) agar). Viscosity and osmotic potential (Psipi,medium) of the media were determined. Developing cotyledonary embryos were analyzed at maturity Stages I-III for water content, osmotic potential (Psipi,embryo) and concentrations of abscisic acid (ABA) and proline. Proliferation of embryogenic tissue, germination rates and the number of embryos formed were also determined in order to relate embryo quality to physiological parameters. Viscosity increased with agar concentration, a phenomenon apparently related to water availability. Many Stage III embryos with high germination potentials were obtained on P24 medium containing 1.0% agar. Embryo water content decreased progressively from 94 to 80% during embryo maturation. Stage I and II embryos that matured on media containing 0.8 or 0.9% agar had similar values of Psipi,embryo, whereas Psipi,embryo of Stage III embryos that matured on medium containing 1.0% agar was significantly lower, although Psipi,medium was unaffected by gel strength. Stage III embryos showed a nearly 16-fold increase in proline concentration and a 50% decrease in ABA concentration compared with Stage I embryos. We conclude that tissue water status and a complex relationship between ABA and proline concentrations, modulated by medium gel strength, are important factors in the maturation process and the quality of oak somatic embryos.

  20. Carbon conversion efficiency and central metabolic fluxes in developing sunflower (Helianthus annuus L.) embryos.

    Science.gov (United States)

    Alonso, Ana P; Goffman, Fernando D; Ohlrogge, John B; Shachar-Hill, Yair

    2007-10-01

    The efficiency with which developing sunflower embryos convert substrates into seed storage reserves was determined by labeling embryos with [U-(14)C6]glucose or [U-(14)C5]glutamine and measuring their conversion to CO2, oil, protein and other biomass compounds. The average carbon conversion efficiency was 50%, which contrasts with a value of over 80% previously observed in Brassica napus embryos (Goffman et al., 2005), in which light and the RuBisCO bypass pathway allow more efficient conversion of hexose to oil. Labeling levels after incubating sunflower embryos with [U-(14)C4]malate indicated that some carbon from malate enters the plastidic compartment and contributes to oil synthesis. To test this and to map the underlying pattern of metabolic fluxes, separate experiments were carried out in which embryos were labeled to isotopic steady state using [1-(13)C1]glucose, [2-(13)C1]glucose, or [U-(13)C5]glutamine. The resultant labeling in sugars, starch, fatty acids and amino acids was analyzed by NMR and GC-MS. The fluxes through intermediary metabolism were then quantified by computer-aided modeling. The resulting flux map accounted well for the labeling data, was in good agreement with the observed carbon efficiency, and was further validated by testing for agreement with gas exchange measurements. The map shows that the influx of malate into oil is low and that flux through futile cycles (wasting ATP) is low, which contrasts with the high rates previously determined for growing root tips and heterotrophic cell cultures.

  1. A comparison of electronic and traditional cigarette butt leachate on the development of Xenopus laevis embryos

    Directory of Open Access Journals (Sweden)

    Tatiana Tatum Parker

    Full Text Available Potential developmental toxicities of three different cigarette butt leachates were evaluated using the frog embryo teratogenesis assay–Xenopus (FETAX. Xenopus laevis embryos were exposed to regular cigarette butt (RCB, menthol (MCB and electronic (ECB in concentrations ranging from 0 to 4 butts/l for RCB and MCB and 0–10 butts/l for ECB. The embryos were from stage 8 to 11 and were exposed for a 96-h period in static renewal test conditions. Median lethal concentration (LC50, malformation (EC50, non-observed adverse effect concentration (NOAEC, and lowest observed adverse effect concentration (LOAEC were calculated. Results from these studies suggest that each tested leachate is teratogenic for X. laevis embryos. The lowest LC50 was determined for ECB exposure at 17.9 cigarette butts/L. The LC50 value was the highest with RCB and MCB having LC50 s of approximately 1 cigarette butt/L. There were notable EC50 differences with RCB having the highest and ECB the lowest. The NOAEC and LOAEC levels for RCB and MCB were below 1 cigarette butt/L for both mortality and malformations; over 8 butts/L for ECB mortality and over 4 butts/L for malformations. From these results, we conclude that RCB leachate is the most toxic compound, while MCB leachate has the higher teratogenicity. ECB leachate has the lowest toxic and teratogenic effects on embryos but there were still noticeable effects. The results confirmed that the FETAX assay can be useful in an integrated biological hazard assessment for the preliminary screening for ecological risks of cigarette butts, and electronic cigarettes, in aquatic environment. Keywords: Cigarette butt leachate, Xenopus laevis, Development

  2. Primordial germ cells and amnion development in the avian embryo

    OpenAIRE

    de Melo Bernardo, Ana

    2016-01-01

    Primordial germ cells (PGCs) are the progenitors of the gametes, responsible for transmitting genetic information from generation to generation. Although there is a long history of gamete biology research, there is still a lot to be learned about many of the mechanisms underlying germ cell development. This dissertation describes and discusses the dynamics of PGCs in the chicken, with a focus on their migration to the gonads and meiosis that takes place when PGCs are already settled there. We...

  3. Incubation media affect the survival, pathway and time of embryo development in Neotropical annual fish Austrolebias nigrofasciatus (Rivulidae).

    Science.gov (United States)

    da Fonseca, A P; Volcan, M V; Robaldo, R B

    2018-01-01

    To analyse the survival, pathway and time of embryo development in the annual fish Austrolebias nigrofasciatus eggs were monitored in four liquid media and two damp media under experimental conditions for 130 days until their development was complete. Eggs kept in the same breeding water from oviposition remained in diapause I (DI) during all experiments. In constrast, up to the stage prior to entering diapause II (DII), the other media had no influence on development. Embryos at this stage (DII), however, show longer development time when treated in medium with water and powdered coconut shell so that about 80% of embryos remained in DII at 100 days. In contrast, all other treatments had a significantly lower proportion of embryos remaining in DII. When treated with Yamamoto's solution in humid media, embryos showed the fastest development. The first fully developed embryos (DIII) were seen at 27 days after oviposition. It took an average of 46-58 days for 50% of eggs in each treatment to reach DIII. Compared with other studies, survival in all incubation media was high at between 70 and 98%. Taken together, it can be concluded that all incubation media were found to be viable for maintaining embryos. Altering developmental trajectories through the manipulation of diapauses in different media makes this species a potential model organism for laboratory studies. © 2017 The Fisheries Society of the British Isles.

  4. Human Development Report 2000: Human Rights and Human Development

    OpenAIRE

    United Nations Development Programme, UNDP

    2000-01-01

    The Human Development Report 2000 looks at human rights as an intrinsic part of development—and at development as a means to realizing human rights. It shows how human rights bring principles of accountability and social justice to the process of human development.

  5. The fate of the vitelline and umbilical veins during the development of the human liver

    NARCIS (Netherlands)

    Hikspoors, Jill P. J. M.; Peeters, Mathijs M. J. P.; Mekonen, Hayelom K.; Kruepunga, Nutmethee; Mommen, Greet M. C.; Cornillie, Pieter; Köhler, S. Eleonore; Lamers, Wouter H.

    2017-01-01

    Differentiation of endodermal cells into hepatoblasts is well studied, but the remodeling of the vitelline and umbilical veins during liver development is less well understood. We compared human embryos between 3 and 10 weeks of development with pig and mouse embryos at comparable stages, and used

  6. Zona pellucida damage to human embryos after cryopreservation and the consequences for their blastomere survival and in-vitro viability.

    Science.gov (United States)

    Van Den Abbeel, E; Van Steirteghem, A

    2000-02-01

    The study objective was to quantify zona pellucida (ZP) damage in cryopreserved human embryos. The influence of two different freezing containers was investigated, and the influence of freezing damage on the survival and viability of the embryos evaluated. ZP damage did not differ according to whether embryos originated from in-vitro fertilization (IVF) cycles or from IVF cycles in association with intracytoplasmic sperm injection (ICSI). The freezing container, however, significantly influenced the occurrence of ZP damage after cryopreservation. More damage was observed when the embryos were frozen-thawed using plastic cryovials than using plastic mini-straws (16.6% versus 2.3%; P plastic mini-straws. The further cleavage of frozen-thawed embryos suitable for transfer was not different whether there was ZP damage or not; however, it was higher when there was 100% blastomere survival as compared with when some blastomeres were damaged (79.0% versus 43.7%; P plastic mini-straws. In conclusion, the aim of a cryopreservation programme should be to have as many fully intact embryos as possible after thawing. Increased ZP damage might indicate a suboptimal cryopreservation procedure.

  7. Xenopus embryos lacking specific isoforms of the corepressor SMRT develop abnormal heads.

    Science.gov (United States)

    Malartre, Marianne; Short, Stephen; Sharpe, Colin

    2006-04-15

    The corepressor SMRT acts on a range of transcription factors, including the retinoid and thyroid hormone nuclear receptors. The carboxy-terminal region of SMRT contains CoRNR box motifs that mediate these interactions. We have shown, in Xenopus, that SMRT can exist as isoforms containing either two or three CoRNR boxes depending on the alternative splicing of exon 37b. The number of SMRT transcript isoforms expressed increases during development until all sixteen possible isoforms are identified in the swimming tadpole. To eliminate specific SMRT isoforms, we have developed a process that uses an antisense morpholino oligonucleotide in Xenopus to dictate the outcome of alternative splicing at a defined exon and used this to inhibit the formation of transcripts containing exon 37b. These embryos are therefore limited to the expression of SMRT isoforms that contain two rather than three CoRNR boxes. Analysis of responsive genes in these embryos shows that targets of thyroid hormone, but not retinoid signaling are affected by the elimination of exon 37b. Morpholino-injected embryos have swimming abnormalities and develop altered head morphology, an expanded olfactory epithelium and disorganized peripheral axons. These experiments indicate a critical role for the alternative splicing of SMRT in development.

  8. Overexpression of aromatase alone is sufficient for ovarian development in genetically male chicken embryos.

    Directory of Open Access Journals (Sweden)

    Luke S Lambeth

    Full Text Available Estrogens play a key role in sexual differentiation of both the gonads and external traits in birds. The production of estrogen occurs via a well-characterised steroidogenic pathway, which is a multi-step process involving several enzymes, including cytochrome P450 aromatase. In chicken embryos, the aromatase gene (CYP19A1 is expressed female-specifically from the time of gonadal sex differentiation. To further explore the role of aromatase in sex determination, we ectopically delivered this enzyme using the retroviral vector RCASBP in ovo. Aromatase overexpression in male chicken embryos induced gonadal sex-reversal characterised by an enlargement of the left gonad and development of ovarian structures such as a thickened outer cortex and medulla with lacunae. In addition, the expression of key male gonad developmental genes (DMRT1, SOX9 and Anti-Müllerian hormone (AMH was suppressed, and the distribution of germ cells in sex-reversed males followed the female pattern. The detection of SCP3 protein in late stage sex-reversed male embryonic gonads indicated that these genetically male germ cells had entered meiosis, a process that normally only occurs in female embryonic germ cells. This work shows for the first time that the addition of aromatase into a developing male embryo is sufficient to direct ovarian development, suggesting that male gonads have the complete capacity to develop as ovaries if provided with aromatase.

  9. Development and quality of porcine parthenogenetically activated embryos after removal of zona pellucida

    DEFF Research Database (Denmark)

    Li, Rong; Liu, Ying; Pedersen, Hanne Skovsgaard

    2013-01-01

    and 2 combined) blastocysts (69.5 ± 2.0 vs. 55.7 ± 2.4, respectively). However, the process showed a significant decrease in apoptosis indicating an increased embryonic quality. Still, the survival percentage of porcine PA embryos after vitrification was dramatically decreased after ZP removal at all...... observation times (P speed of embryonic development and decreasing the number of apoptotic cells in blastocysts even though developmental percentages and embryonic robustness might be decreased....

  10. Loss of Bmal1 decreases oocyte fertilization, early embryo development and implantation potential in female mice.

    Science.gov (United States)

    Xu, Jian; Li, Yan; Wang, Yizi; Xu, Yanwen; Zhou, Canquan

    2016-10-01

    Biological clock genes expressed in reproductive tissues play important roles in maintaining the normal functions of reproductive system. However, disruption of female circadian rhythm on oocyte fertilization, preimplantation embryo development and blastocyst implantation potential is still unclear. In this study, ovulation, in vivo and in vitro oocyte fertilization, embryo development, implantation and intracellular reactive oxygen species (ROS) levels in ovary and oviduct were studied in female Bmal1+/+ and Bmal1-/- mice. The number of naturally ovulated oocyte in Bmal1-/- mice decreased (5.2 ± 0.8 vs 7.8 ± 0.8, P fertilization rate and obtained blastocyst number were observed in Bmal1-/- female mice either mated with wild-type in vivo or fertilized by sperm from wild-type male mice in vitro (all P fertilization rate of oocytes derived from Bmal1-/- increased significantly compared with in vivo study (P fertilization rate, early embryo development and implantation potential in female mice, and these may be possibly caused by excess ROS levels generated in ovary and oviduct.

  11. RAPID COMMUNICATION: Nerve growth factor influences cleavage rate and embryo development in sheep.

    Science.gov (United States)

    Crispo, M; Dos Santos-Neto, P C; Vilariño, M; Mulet, A P; de León, A; Barbeito, L; Menchaca, A

    2016-10-01

    Recent information about Nerve growth factor (NGF), a protein traditionally associated to the nervous system that regulates survival and maturation of developing neurons, suggests that it may exert action also on different levels in the reproductive system. The aim of this study was to evaluate the effect of NGF added during in vitro oocyte maturation, fertilization or in vitro embryo development in sheep. Nerve growth factor was supplemented to the culture medium at 0, 100, or 1,000 ng/mL, during either in vitro maturation (Exp. 1), in vitro fertilization (Exp. 2), or in vitro culture (Exp. 3). In addition, NGF mRNA expression was determined in cumulus cells and oocytes. Nerve growth factor induced early cleavage when added during oocyte maturation or fertilization, improved embryo development when added during fertilization, and had no significant effect when added during embryo culture. In general, the effect was more evident with 100 rather than 1,000 ng/mL (P growth factor on oocyte maturation and mainly on the fertilization process.

  12. Protein deubiquitination during oocyte maturation influences sperm function during fertilisation, antipolyspermy defense and embryo development.

    Science.gov (United States)

    Yi, Young-Joo; Sutovsky, Miriam; Song, Won-Hee; Sutovsky, Peter

    2015-11-01

    Ubiquitination is a covalent post-translational modification of proteins by the chaperone protein ubiquitin. Upon docking to the 26S proteasome, ubiquitin is released from the substrate protein by deubiquitinating enzymes (DUBs). We hypothesised that specific inhibitors of two closely related oocyte DUBs, namely inhibitors of the ubiquitin C-terminal hydrolases (UCH) UCHL1 (L1 inhibitor) and UCHL3 (L3 inhibitor), would alter porcine oocyte maturation and influence sperm function and embryo development. Aberrant cortical granule (CG) migration and meiotic spindle defects were observed in oocytes matured with the L1 or L3 inhibitor. Embryo development was delayed or blocked in oocytes matured with the general DUB inhibitor PR-619. Aggresomes, the cellular stress-inducible aggregates of ubiquitinated proteins, formed in oocytes matured with L1 inhibitor or PR-619, a likely consequence of impaired protein turnover. Proteomic analysis identified the major vault protein (MVP) as the most prominent protein accumulated in oocytes matured with PR-619, suggesting that the inhibition of deubiquitination altered the turnover of MVP. The mitophagy/autophagy of sperm-contributed mitochondria inside the fertilised oocytes was hindered by DUB inhibitors. It is concluded that DUB inhibitors alter porcine oocyte maturation, fertilisation and preimplantation embryo development. By regulating the turnover of oocyte proteins and mono-ubiquitin regeneration, the DUBs may promote the acquisition of developmental competence during oocyte maturation.

  13. Noninferiority, randomized, controlled trial comparing embryo development using media developed for sequential or undisturbed culture in a time-lapse setup.

    Science.gov (United States)

    Hardarson, Thorir; Bungum, Mona; Conaghan, Joe; Meintjes, Marius; Chantilis, Samuel J; Molnar, Laszlo; Gunnarsson, Kristina; Wikland, Matts

    2015-12-01

    To study whether a culture medium that allows undisturbed culture supports human embryo development to the blastocyst stage equivalently to a well-established sequential media. Randomized, double-blinded sibling trial. Independent in vitro fertilization (IVF) clinics. One hundred twenty-eight patients, with 1,356 zygotes randomized into two study arms. Embryos randomly allocated into two study arms to compare embryo development on a time-lapse system using a single-step medium or sequential media. Percentage of good-quality blastocysts on day 5. Percentage of day 5 good-quality blastocysts was 21.1% (standard deviation [SD] ± 21.6%) and 22.2% (SD ± 22.1%) in the single-step time-lapse medium (G-TL) and the sequential media (G-1/G-2) groups, respectively. The mean difference (-1.2; 95% CI, -6.0; 3.6) between the two media systems for the primary end point was less than the noninferiority margin of -8%. There was a statistically significantly lower number of good-quality embryos on day 3 in the G-TL group [50.7% (SD ± 30.6%) vs. 60.8% (SD ± 30.7%)]. Four out of the 11 measured morphokinetic parameters were statistically significantly different for the two media used. The mean levels of ammonium concentration in the media at the end of the culture period was statistically significantly lower in the G-TL group as compared with the G-2 group. We have shown that a single-step culture medium supports blastocyst development equivalently to established sequential media. The ammonium concentrations were lower in the single-step media, and the measured morphokinetic parameters were modified somewhat. NCT01939626. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  14. Embryo development in the lady's slipper orchid, Paphiopedilum delenatii, with emphasis on the ultrastructure of the suspensor.

    Science.gov (United States)

    Lee, Yung-I; Yeung, Edward C; Lee, Nean; Chung, Mei-Chu

    2006-12-01

    Owing to large-scale collecting, the lady's slipper orchid, Paphiopedilum delenatii, is under threat of extinction. Asymbiotic germination provides a useful way to re-establish plants in the wild and for commercial propagation. A detailed study of embryo development would provide information on subsequent germination events and aid in the propagation of the species. Developing capsules were collected for histochemical and ultrastructural studies by using both light and transmission electron microscopy. The suspensor of this species consists of three vacuolated cells. During the early globular stage of embryo development, structural differentiation occurs, revealing an abundance of smooth endoplasmic reticulum cisternae and wall ingrowths within the suspensor cells. These features are not present in cells of the embryo proper. Furthermore, the results of Nile red staining demonstrate that a cuticular layer is present only in the embryo proper, but absent from the suspensor. Cuticular material is also present in the inner walls of the seed coat, and persists through seed maturation. The morphological features of the transfer cell and the absence of cuticular material in the suspensor cell wall corroborate the hypothesis that the suspensor is the major nutrient uptake site for the developing embryo in the lady's slipper orchid. The absence of an endosperm and presence of cuticular material in the inner walls of the seed coat enclosing the embryo proper further support the notion that nutrient uptake by the embryo is confined to the micropylar end of the seed through the suspensor.

  15. Effect of bovine oviductal extracellular vesicles on embryo development and quality in vitro.

    Science.gov (United States)

    Lopera-Vasquez, Ricaurte; Hamdi, Meriem; Maillo, Veronica; Gutierrez-Adan, Alfonso; Bermejo-Alvarez, Pablo; Ramírez, Miguel Ángel; Yáñez-Mó, María; Rizos, Dimitrios

    2017-04-01

    The aim of this study was to evaluate the effect of extracellular vesicles (EV) from oviductal fluid (OF), either from the ampulla or isthmus, on the development and quality of in vitro-cultured bovine embryos. Zygotes were cultured in synthetic oviduct fluid (SOF + 3 mg/mL BSA) without calf serum (C- group), in the presence of 3 × 105 EV/mL from ampullary or isthmic OF at either 1 × 104 g (10 K) or 1 × 105 g (100 K), and compared with SOF + 5% FCS (C+ group). OF-EV size and concentration were assessed by electron microscopy and nanotracking analysis system. Embryo development was recorded on Days 7-9, and blastocyst quality was assessed through cryotolerance and gene expression analysis. Lower blastocyst yield was observed on Day 7 in the C- and OF-EV groups (12.0-14.3%) compared with C+ (20.6%); however, these differences were compensated at Days 8 and 9 (Day 9: 28.5-30.8%). Importantly, the survival rate of blastocysts produced with isthmic 100 K OF-EV was higher than that of C+ and C- group at 72 h after vitrification and warming (80.1 vs 34.5 and 50.5% respectively, P produced in the presence of 100 K isthmic OF-EV upregulated the water channel AQP3 and DNMT3A and SNRPN transcripts compared with the C+, with the expression in C- being intermediate. The lipid receptor LDLR was downregulated in C+ compared with all other groups. In conclusion, the addition of oviductal fluid extracellular vesicles from isthmus, to in vitro culture of bovine embryos in the absence of serum improves the development and quality of the embryos produced. © 2017 Society for Reproduction and Fertility.

  16. Endometrial stromal cells of women with recurrent miscarriage fail to discriminate between high- and low-quality human embryos.

    Directory of Open Access Journals (Sweden)

    Charlotte H E Weimar

    Full Text Available BACKGROUND: The aetiology of recurrent miscarriage (RM remains largely unexplained. Women with RM have a shorter time to pregnancy interval than normally fertile women, which may be due to more frequent implantation of non-viable embryos. We hypothesized that human endometrial stromal cells (H-EnSCs of women with RM discriminate less effectively between high-and low-quality human embryos and migrate more readily towards trophoblast spheroids than H-EnSCs of normally fertile women. METHODOLOGY/PRINCIPAL FINDINGS: Monolayers of decidualized H-EnSCs were generated from endometrial biopsies of 6 women with RM and 6 fertile controls. Cell-free migration zones were created and the effect of the presence of a high-quality (day 5 blastocyst, n = 13, a low-quality (day 5 blastocyst with three pronuclei or underdeveloped embryo, n = 12 or AC-1M88 trophoblast cell line spheroid on H-ESC migratory activity was analyzed after 18 hours. In the absence of a spheroid or embryo, migration of H-EnSCs from fertile or RM women was similar. In the presence of a low-quality embryo in the zone, the migration of H-EnSCs of control women was inhibited compared to the basal migration in the absence of an embryo (P<0.05 and compared to the migration in the presence of high-quality embryo (p<0.01. Interestingly, the migratory response H-EnSCs of women with RM did not differ between high- and low-quality embryos. Furthermore, in the presence of a spheroid their migration was enhanced compared to the H-EnSCs of controls (p<0.001. CONCLUSIONS: H-EnSCs of fertile women discriminate between high- and low-quality embryos whereas H-EnSCs of women with RM fail to do so. H-EnSCs of RM women have a higher migratory response to trophoblast spheroids. Future studies will focus on the mechanisms by which low-quality embryos inhibit the migration of H-EnSCs and how this is deregulated in women with RM.

  17. RESEARCHES REGARDING THE INFLUENCE OF RECOVERY MEDIA ON THE IN VITRO DEVELOPMENT CAPACITY OF THE PREIMPLANTATIONAL MOUSE EMBRYO

    Directory of Open Access Journals (Sweden)

    ADA CEAN

    2009-05-01

    Full Text Available Phosphate Bufered Saline with 0.4% BSA and M2 medium are one of the most common media used in embryorecovery. The aim of our paper was to investigate if the recovery media used for the recovery of the mouseembryo is influencing in vitro developmental capacity. As biological material we used 10 used were mousefemales, age 2 months superovulated with 5UI PMSG (Pregnant Mare Serum Gonadotropine and 5 UI hCG(human Corionic Gonadotropine. The embryos used were recovered, by oviduct flushing, at 24 hours from theidentification of the vaginal plug. The majority of the embryos (78.3% were in two cells stage. A total of 123, 2cells embryos were cultivated in M16 medium. The evolution of the embryos was examined at 24, 48 and 72hours interval. The proportion of hatched blastocyst was higher at the embryos recovered with M2 (53.7%compared with the embryos recovered with PBS 0.4% BSA. The difference is statistically very significant(p<0.001. Embryos recovered in M2 media have a higher in vitro developmental capacity compared with theembryos recovered in PBS media supplemented with 0,4% BSA, possibly because of the sodium bicarbonate andlactate used in M2 media for pH regulation.

  18. Nucleolar development and allocation of key nucleolar proteins require de novo transcription in bovine embryos

    DEFF Research Database (Denmark)

    Svarcova, Olga; Laurincik, Jozef; Avery, Birthe

    2007-01-01

    The goal of the present study was to investigate whether key nucleolar proteins involved in ribosomal RNA (rRNA) transcription and processing are transcribed de novo or from maternally inherited messenger RNAs (mRNA) in bovine embryos, and to which extent de novo transcription of these proteins mRNA...... embryos cultured from the 2-cell stage with or without (control groups) a-amanitin, which blocks the RNA plymerases II and III transcription and, thus the synthesis of mRNA. In the control groups, weak autoradiographic labelling was initially observed in the periphery of few nuclei at the 4-cell...... is required for the development of functional nucleoli during the major activation of the embryonic genome. Immunofluorescence for localization of key nucleolar proteins, autoradiography for detection of transcriptional activity, and transmission electron microscopy were applied to in vitro produc ed bovine...

  19. Evaluation of the effects of mobile phones on the neural tube development of chick embryos.

    Science.gov (United States)

    Umur, Ahmet Sukru; Yaldiz, Can; Bursali, Adem; Umur, Nurcan; Kara, Burcu; Barutcuoglu, Mustafa; Vatansever, Seda; Selcuki, Deniz; Selcuki, Mehmet

    2013-01-01

    The objective of this study is to examine the effects of radiation of mobile phones on developing neural tissue of chick embryos. There were 4 study groups. All Groups were placed in equal distance, from the mobile phones. Serial sections were taken from each Group to study the neural tube segments. The TUNEL results were statistically significant (p negative in the 48 and 72 hours in the Control Group, had moderate activity in the third Group 3, weak activity in the 48 hour, and was negative in the 72 hour in other groups. Caspase-9 immunoreactivity was weak in Group 1, 2 and 3 at 30 hours and was negative in Group 1 and 4 at 48 and 72 hours. Caspase-9 activity in the third Group was weak in all three stages. Electromagnetic radiation emitted by mobile phones caused developmental delay in chick embryos in early period. This finding suggests that the use of mobile phones by pregnant women may pose risks.

  20. Expression of the bone morphogenetic protein-2 (BMP2 in the human cumulus cells as a biomarker of oocytes and embryo quality

    Directory of Open Access Journals (Sweden)

    Sirin B Demiray

    2017-01-01

    Full Text Available Background: The members of the transforming growth factor-B superfamily, as the bone morphogenetic proteins (BMPs subfamily and anti-Müllerian hormone (AMH, play a role during follicular development, and the bone morphogenetic protein-2 (BMP2, AMH, and THY1 are expressed in ovaries. Aim: This study was designed to define whether or not the expressions of these proteins in human cumulus cells (CCs can be used as predictors of the oocyte and embryo competence. Settings and Design: The study included nine female patients who were diagnosed as idiopathic infertility, aged 25–33 years (median 30 years and underwent Assisted Reproductive Technologies. Materials and Methods: The CCs from 60 oocyte–cumulus complexes obtained from the nine patients were evaluated with immunofluorescence staining in respect of BMPs, AMH and THY1 markers. The CCs surrounding the same oocytes were evaluated separately according to the oocyte and embryo quality. Statistical Analysis: Quantitative data were statistically analyzed for differences using the two-sided Mann–Whitney U test (P < 0.05. Results and Conclusions: Significant differences in immunofluorescence staining were observed in oocyte quality and embryo quality for the BMP2 only (P < 0.05. No significant differences were observed for AMH or CD90/THY1. Conclusion: These results demonstrated that there is a significant difference in the expression of BMP2 in the CCs of good quality oocytes and subsequently a good embryo.

  1. Study of the nervous tissue development in rainbow trout (Oncorhynchus mykiss) embryos treated with oxytetracycline.

    Science.gov (United States)

    Arias, P; Pinochet, L F; Disi, A

    2002-06-01

    The purpose of this study was to discover whether the use of different doses of oxytetracycline causes any alteration in the development of nervous tissue in rainbow trout embryos. Five thousands eggs of females rainbow trout were divided into five groups. One group acted as control and the other four were administered with one of four doses of oxytetracycline, 0.025, 0.050, 0.100, or 0.201 microM, at the moment of fertilization. The eggs were incubated under pisciculture conditions to just before being ready to spring off. From the 10th day, 10-egg samples were taken regularly and fixed. Five were processed for histological techniques and stained with haematoxylin and eosin, cresyl fast violet and silver, the other five were homogenized for antibiotic detection. Histological alterations appeared in 37-day-old embryos, with an abnormal migration of the neuroblasts to the marginal layer of the neural cord, and alterations in the development of the lens and eye layers. Some embryos showed abnormal curvature of the spinal cord but these changes were not statistically significant.

  2. Development of glutathione-deficient embryos in Arabidopsis is influenced by the maternal level of glutathione.

    Science.gov (United States)

    Lim, B; Meyer, A J; Cobbett, C S

    2011-07-01

    Glutathione (GSH) biosynthesis-deficient gsh1 and gsh2 null mutants of Arabidopsis thaliana have late embryonic-lethal and early seedling-lethal phenotypes, respectively, when segregating from a phenotypically wild-type parent plant, indicating that GSH is required for seed maturation and during germination. In this study, we show that gsh2 embryos generated in a partially GSH-deficient parent plant, homozygous for either the cad2 mutation in the GSH1 gene or homozygous for mutations in CLT1, CLT2 and CLT3 encoding plastid thiol transporters, abort early in embryogenesis. In contrast, individuals homozygous for the same combinations of mutations but segregating from heterozygous, phenotypically wild-type parents exhibit the parental gsh2 seedling-lethal phenotype. Similarly, homozygous gsh1 embryos generated in a gsh1/cad2 partially GSH-deficient parent plant abort early in development. These observations indicate that the development of gsh1 and gsh2 embryos to a late stage is dependent on the level of GSH in the maternal plant. © 2011 German Botanical Society and The Royal Botanical Society of the Netherlands.

  3. Guarding embryo development of zebrafish by shell engineering: a strategy to shield life from ozone depletion.

    Directory of Open Access Journals (Sweden)

    Ben Wang

    Full Text Available BACKGROUND: The reduced concentration of stratospheric ozone results in an increased flux of biologically damaging mid-ultraviolet radiation (UVB, 280 to 320 nm reaching earth surfaces. Environmentally relevant levels of UVB negatively impact various natural populations of marine organisms, which is ascribed to suppressed embryonic development by increased radiation. METHODOLOGY/PRINCIPAL FINDINGS: Inspired by strategies in the living systems generated by evolution, we induce an extra UVB-adsorbed coat on the chorion (eggshell surrounding embryo of zebrafish, during the blastula period. Short and long UV exposure experiments show that the artificial mineral-shell reduces the UV radiation effectively and the enclosed embryos become more robust. In contrast, the uncoated embryos cannot survive under the enhanced UVB condition. CONCLUSIONS: We suggest that an engineered shell of functional materials onto biological units can be developed as a strategy to shield lives to counteract negative changes of global environment, or to provide extra protection for the living units in biological research.

  4. Polymethoxy-1-alkenes from Aphanizomenon ovalisporum Inhibit Vertebrate Development in the Zebrafish (Danio rerio Embryo Model

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    John P. Berry

    2012-10-01

    Full Text Available Cyanobacteria are recognized producers of a wide array of toxic or otherwise bioactive secondary metabolites. The present study utilized the zebrafish (Danio rerio embryo as an aquatic animal model of vertebrate development to identify, purify and characterize lipophilic inhibitors of development (i.e., developmental toxins from an isolate of the freshwater cyanobacterial species, Aphanizomenon ovalisporum. Bioassay-guided fractionation led to the purification, and subsequent chemical characterization, of an apparent homologous series of isotactic polymethoxy-1-alkenes (1–6, including three congeners (4–6 previously identified from the strain, and two variants previously identified from other species (2 and 3, as well as one apparently novel member of the series (1. Five of the PMAs in the series (1–5 were purified in sufficient quantity for comparative toxicological characterization, and toxicity in the zebrafish embryo model was found to generally correlate with relative chain length and/or methoxylation. Moreover, exposure of embryos to a combination of variants indicates an apparent synergistic interaction between the congeners. Although PMAs have been identified previously in cyanobacteria, this is the first report of their apparent toxicity. These results, along with the previously reported presence of the PMAs from several cyanobacterial species, suggest a possibly widespread distribution of the PMAs as toxic secondary metabolites and warrants further chemical and toxicological investigation.

  5. Indole Alkaloids from Fischerella Inhibit Vertebrate Development in the Zebrafish (Danio rerio Embryo Model

    Directory of Open Access Journals (Sweden)

    Katherine Walton

    2014-12-01

    Full Text Available Cyanobacteria are recognized producers of toxic or otherwise bioactive metabolite associated, in particular, with so-called “harmful algal blooms” (HABs and eutrophication of freshwater systems. In the present study, two apparently teratogenic indole alkaloids from a freshwater strain of the widespread cyanobacterial genus, Fischerella (Stigonemataceae, were isolated by bioassay-guided fractionation, specifically using the zebrafish (Danio rerio embryo, as a model of vertebrate development. The two alkaloids include the previously known 12-epi-hapalindole H isonitrile (1, and a new nitrile-containing variant, 12-epi-ambiguine B nitrile (2. Although both compounds were toxic to developing embryos, the former compound was shown to be relatively more potent, and to correlate best with the observed embryo toxicity. Related indole alkaloids from Fischerella, and other genera in the Stigonemataceae, have been widely reported as antimicrobial compounds, specifically in association with apparent allelopathy. However, this is the first report of their vertebrate toxicity, and the observed teratogenicity of these alkaloids supports a possible contribution to the toxicity of this widespread cyanobacterial family, particularly in relation to freshwater HABs and eutrophication.

  6. Development of embryos in superovulated guinea pigs following active immunization against the inhibin alpha-subunit.

    Science.gov (United States)

    Shi, F; Mochida, K; Suzuki, O; Matsuda, J; Ogura, A; Tsonis, C G; Watanabe, G; Suzuki, A K; Taya, K

    2000-08-01

    Embryo recovery and subsequent embryonic development from guinea pigs treated with or without inhibin vaccines were compared to determine the effect of active immunization against the inhibin alpha-subunit. Twenty female guinea pigs of the Hartley strain were injected 3 times either with 1 ml inhibin vaccine (recombinant ovine inhibin a-subunit in oil emulsion: 50 microg/ml, inhibin-immunized group), or 1 ml placebo (saline in oil emulsion; control group) at 4 week intervals. After one estrous cycle following the last injection, females were naturally mated and embryos were collected at 11:00 hr of day 6 of pregnancy (Day 1: sperm in the vaginal smear) for culture in vitro. Active immunization increased the number of corpora lutea (12.6+/-3.0 vs. 4.6+/-0.2, P0.05). During subsequent 8 day culture in vitro, most of the recovered embryos formed trophoblast outgrowth; 100% (14/14) and 88.2% (15/17) in control and immunized groups, respectively. High levels of inhibin antibody titers were sustained in the inhibin-immunized guinea pigs at least for 5 months after the last injection while no antibody titer was detected in the control animals. These results indicate that active immunization against the inhibin a-subunit is a long-acting and efficient method to induce superovulation with normal embryonic development in the guinea pig.

  7. The effect of hepatocyte growth factor on mouse oocyte in vitro maturation and subsequent fertilization and embryo development

    Directory of Open Access Journals (Sweden)

    Mohammad H. Bahadori

    2011-05-01

    Full Text Available Background: Oocyte invitro maturation is an enormously promising technology for the treatment of infertility, yet its clinical application remains limited owing to poor success rates. Therefore, this study was devised to evaluate the effect of hepatocyte growth factor (HGF on in vitro maturation of immature mouse oocytes and resulting embryos development. Materials and Method: Cumulus – oocyte complex and germinal vesicle were obtained from eighteen 6-8 weeks-old female NMRI mice 46-48 hours after administration of an injection of 5 IU PMSG (Pregnant Mares’ Serum Gonadotrophin. Oocytes were culture in TCM199 (Tissue culture medium-199 supplemented with dosages of 0, 10, 20, 50 and 100 ng/ml of HGF. After 24 hours, metaphase ІІ oocytes were co-incubated with sperms for 4-6 hours in T6 medium. Following isolation of two pronucleus embryos, cleavage of embryos was assessed in the same medium till blastocyst stage. The number of oocytes and embryos was recorded under an invert microscope and the rate of oocyte maturation, fertilization and embryos cleavage until blastocyst stage compared using of student χ2 test. Results: In all compared groups, oocytes growth and embryos development rate in the 20 ng/ml of HGF treatment group was significantly higher (p<0.05 than the control group (p<0.05.Conclusion: 20 ng/ml of HGF improved the nuclear maturation and embryo development up to blastocyst stage during culture condition

  8. Development of rotating-mesh basket type bioreactor for carrot embryo production in immobilized callus system

    Energy Technology Data Exchange (ETDEWEB)

    Suehara, K.; Nagamori, E.; Honda, H.; Uozumi, N.; Kobayashi, T. [Nagoya Univ. (Japan)

    1998-08-01

    Mass-production of regenerated plantlets using alginate bead encapsulating carrot callus was studied. When gel beads were cultivated in a flask containing regeneration medium, somatic embryos were released at high frequency from the gel beads and developed into plantlets. When a callus less than 320 {mu}m in size was immobilized in the beads, a larger number of embryo was released from the beads. At 0.11 ml-pcv/ml-gel of inoculum size, the maximum number of embryo was obtained. In order to develop a suitable bioreactor for the immobilized callus system, three types of bioreactors, namely air-lift, horizontally shaking vessel and rotating mesh-basket types, were tested. In the case of the rotating mesh-basket type, a larger amount of released cells was obtained. Plantlet formation from the released cells reached a level of 115% compared with that of the flask culture. To achieve continuous recovery of released cells and perform long term culture, a cyclone type cell separator was designed and used in conjunction with a rotating mesh-basket type bioreactor. When 110 ml/min of medium flow was passed through the separator, 90% of suspended cells were collected after 40 min of operation time. Long term cultures at 50, 100, and 150 rpm of rotational speed were carried out. In the experiment at 100 rpm, almost constant values of released cell volume and number of embryos were obtained during 12 d. They were 3 ml-pcv/1-medium/d and 7000 plantlets/g-released cells, respectively. 5 refs., 7 figs.

  9. 17{beta}-Estradiol inhibits chondrogenesis in the skull development of zebrafish embryos

    Energy Technology Data Exchange (ETDEWEB)

    Fushimi, Shigeko, E-mail: fushimi@med.kawasaki-m.ac.jp [Department of Public Health, Kawasaki Medical School, 577 Matsushima, Kurashiki 701-0192 (Japan); Wada, Naoyuki, E-mail: wada@med.kawasaki-m.ac.jp [Department of Molecular and Developmental Biology, Kawasaki Medical School, 577 Matsushima, Kurashiki 701-0192 (Japan); Nohno, Tsutomu, E-mail: nohno@bcc.kawasaki-m.ac.jp [Department of Molecular and Developmental Biology, Kawasaki Medical School, 577 Matsushima, Kurashiki 701-0192 (Japan); Tomita, Masafumi, E-mail: toxicology@med.kawasaki-m.ac.jp [Department of Medical Toxicology, Kawasaki Medical School, 577 Matsushima, Kurashiki 701-0192 (Japan); Saijoh, Kiyofumi, E-mail: saijohk@med.kanazawa-u.ac.jp [Department of Hygiene, Kanazawa University School of Medicine, 13-1 Takaramachi, Kanazawa 920-8564 (Japan); Sunami, Shigeo, E-mail: ssunami@med.kawasaki-m.ac.jp [Department of Clinical Nutrition, Kawasaki University of Medical Welfare, 288 Matsushima, Kurashiki 701-0193 (Japan); Katsuyama, Hironobu, E-mail: katsu@med.kawasaki-m.ac.jp [Department of Public Health, Kawasaki Medical School, 577 Matsushima, Kurashiki 701-0192 (Japan)

    2009-12-13

    17{beta}-Estradiol (E2) plays important roles in the development and differentiation of the gonad and central nervous systems, but little is known regarding the effects of exogenous E2 on chondrogenesis in skeletal development. In the present study, we found that treatment with E2 1-5 days post-fertilization (dpf) at concentrations above 1.5 x 10{sup -5} M increased the mortality rate in zebrafish embryos. Morphological analysis showed that treatment with E2 1-5 dpf caused abnormal cartilage formation in a dose-dependent manner at concentrations above 5 x 10{sup -6} M. E2 1-5 dpf at 1.5 x 10{sup -5} M caused defects of the ethmoid plate, parallel cleft of the trabecular cartilage, and hypoplasia of Meckel's cartilage and the ceratohyal cartilage. The sensitivity of embryos to E2 depended on the developmental stage. In early chondrogenesis (1-2 dpf), the embryos were highly sensitive to E2, leading to hypoplasia of the cartilage. In situ hybridization studies showed that expression levels of patched1 (ptc1) and patched2 (ptc2) receptor mRNAs were markedly decreased by exposure to 2 x 10{sup -5} M E2 1-2 dpf. However, the expression levels of sonic hedgehog (shh) and tiggywinkle hedgehog (twhh) mRNAs were constant in the E2-treated embryos. In addition, the estrogen receptor antagonist ICI 182,780 did not completely abolish the effects of E2, suggesting that E2 may not inhibit chondrogenesis through its nuclear estrogen receptor. These results suggest that exposure to exogenous E2 possibly inhibits chondrogenesis via inhibition of the hedgehog (Hh) signal transduction system.

  10. POST-HARVEST EMBRYO DEVELOPMENT IN GINSENG SEEDS INCREASES DESICCATION SENSITIVITY AND NARROWS THE HYDRATION WINDOW FOR CRYOPRESERVATION.

    Science.gov (United States)

    Han, E; Popova, E; Cho, G; Park, S; Lee, S; Pritchard, H W; Kim, H H

    Despite its self-pollinating characteristics, Korean ginseng germplasm is mainly maintained in clonal gene banks as there is no defined approach to the long-term conservation of its seed, including the most appropriate stage of embryo development for storage. The aim of this study was to reveal the effect of embryo development on desiccation tolerance and cryopreservation success in ginseng seeds. Seeds of Korean ginseng (Panax ginseng C.A. Meyer) at three post-harvest stages (immediately after harvesting and following treatments to enable internal growth of the embryo) were desiccated and cryopreserved. The hydration window for the >80% dehiscence and germination of cryopreserved ginseng seeds varied with embryo developmental stage: 3-9% moisture content (MC) for both unpulped and undehisced seeds when the embryo was 0.1 the length of the endosperm, 7-10% MC for dehisced seeds (0.5 embryo:endosperm) and 9-11% MC for seeds with fully developed embryos (0.9 embryo:endosperm). Whilst dried (4-8% moisture content) and undehisced seeds within fruits (unpulped seeds) lost more than half their viability during 1 year's storage at room temperature, cryopreservation enabled germination levels of c. 90%. Overall, 432 accessions of Korean ginseng landraces have been cryopreserved using undehisced seeds with or without fruits. Post-harvest treatment of Korean ginseng seeds to enable embryo development decreases tolerance of very low MCs, and thus narrows the hydration window for cryopreservation. Fresh-harvested and unpulped seeds that have been dried to c. 5% MC are recommended for long-term cryogenic storage.

  11. Vertical transmission and successive location of symbiotic bacteria during embryo development and larva formation in Corticium candelabrum (Porifera: Demospongiae)

    NARCIS (Netherlands)

    Caralt Bosch, de S.; Uriz, M.J.; Wijffels, R.H.

    2007-01-01

    This study reports on the transfer of heterotrophic bacteria from parental tissue to oocytes in the Mediterranean bacteriosponge Corticium candelabrum (Homosclerophorida) and the description of the successive locations of the microsymbionts during embryo development through transmission and scanning

  12. Rotenone Decreases Hatching Success in Brine Shrimp Embryos by Blocking Development: Implications for Zooplankton Egg Banks.

    Directory of Open Access Journals (Sweden)

    Joseph A Covi

    Full Text Available While many zooplankton species recover quickly after the treatment of water resources with the piscicide, rotenone, some fail to reach pretreatment population density or, in rare cases, do not reappear at all. The variable impact of rotenone on zooplankton populations could stem from differences in the capacity of species to switch entirely to anaerobic catabolic pathways in the presence of rotenone, which blocks mitochondrial electron transport. Alternatively, variable responses among species could originate from differences in permeability of dormant life-stages to lipophilic chemicals like rotenone. The purpose of the present study was to determine the effects of rotenone on development, emergence and hatching of zooplankton embryos that lack both the anaerobic capacity to develop in the presence of rotenone and a permeability barrier to prevent the entry of rotenone during dormancy. Post-diapause embryos of the brine shrimp, Artemia franciscana, were employed as a model system, because they are permeable to lipophilic compounds when dechorionated and require aerobic conditions to support development. Early development in this species is also well characterized in the literature. Brine shrimp embryos were exposed to rotenone while development was either slowed by chilling or suspended by anoxia. Development, emergence and hatching were then observed in rotenone-free artificial seawater. The data presented demonstrate that rotenone freely diffuses across the embryonic cuticle in a matter of hours, and prevents development and emergence after brief exposures to ecologically relevant concentrations (0.025-0.5 mg L-1 of the piscicide. Neither the removal of rotenone from the environment, nor the removal of embryonic water with a hypertonic solution, are sufficient to reverse this block on development and emergence. These data indicate that rotenone could impair recruitment from egg banks for species of zooplankton that lack both an embryonic

  13. Rotenone Decreases Hatching Success in Brine Shrimp Embryos by Blocking Development: Implications for Zooplankton Egg Banks.

    Science.gov (United States)

    Covi, Joseph A; Hutchison, Evan R; Neumeyer, Courtney H; Gunderson, Matthew D

    While many zooplankton species recover quickly after the treatment of water resources with the piscicide, rotenone, some fail to reach pretreatment population density or, in rare cases, do not reappear at all. The variable impact of rotenone on zooplankton populations could stem from differences in the capacity of species to switch entirely to anaerobic catabolic pathways in the presence of rotenone, which blocks mitochondrial electron transport. Alternatively, variable responses among species could originate from differences in permeability of dormant life-stages to lipophilic chemicals like rotenone. The purpose of the present study was to determine the effects of rotenone on development, emergence and hatching of zooplankton embryos that lack both the anaerobic capacity to develop in the presence of rotenone and a permeability barrier to prevent the entry of rotenone during dormancy. Post-diapause embryos of the brine shrimp, Artemia franciscana, were employed as a model system, because they are permeable to lipophilic compounds when dechorionated and require aerobic conditions to support development. Early development in this species is also well characterized in the literature. Brine shrimp embryos were exposed to rotenone while development was either slowed by chilling or suspended by anoxia. Development, emergence and hatching were then observed in rotenone-free artificial seawater. The data presented demonstrate that rotenone freely diffuses across the embryonic cuticle in a matter of hours, and prevents development and emergence after brief exposures to ecologically relevant concentrations (0.025-0.5 mg L-1) of the piscicide. Neither the removal of rotenone from the environment, nor the removal of embryonic water with a hypertonic solution, are sufficient to reverse this block on development and emergence. These data indicate that rotenone could impair recruitment from egg banks for species of zooplankton that lack both an embryonic barrier to the entry of

  14. Trefoil factor family peptides TFF1 and TFF3 in the nervous tissues of developing mouse embryo

    Directory of Open Access Journals (Sweden)

    Tatjana Belovari

    2015-02-01

    Full Text Available Trefoil factor family peptides (TFF1, TFF2, and TFF3 are predominantly found in mucous epithelia of various organs. However, they have also been reported in the nervous tissue, particularly mouse, rat, porcine, and human brain. The aim of this research was to determine the presence of TFF1 and TFF3 in the nervous system of developing mouse embryo. Mouse embryos, at the stages E15 to E17 were isolated, fixed in 4% paraformaldehyde and embedded in paraffin blocks. Sagittal 6µm sections were made, processed for immunohistochemistry, and incubated with anti-TFF1 or anti-TFF3 primary polyclonal rabbit antibodies. Labeled streptavidin-biotin method was used for TFF detection. TFF1 and 3 were found in the cytoplasm of ganglion cell somata, while TFF3 staining was also visible in the cytoplasm of neurons in different areas and nuclei of brain and medulla oblongata. Neurons in the gray matter of spinal cord were also TFF1 and TFF3 positive, and signal for both peptides was found in the choroid plexus. TFF peptides might be involved in the complex processes of nervous system development and differentiation and brain plasticity.

  15. Application of next-generation sequencing for 24-chromosome aneuploidy screening of human preimplantation embryos.

    Science.gov (United States)

    Zheng, Haiyan; Jin, Hua; Liu, Lian; Liu, Jianqiao; Wang, Wei-Hua

    2015-01-01

    Aneuploidy is a leading cause of repeat implantation failure and recurrent miscarriages. Preimplantation genetic screening (PGS) enables the assessment of the numeral and structural chromosomal errors of embryos before transfer in patients undergoing in vitro fertilization. Array comparative genomic hybridization (aCGH) has been demonstrated to be an accurate PGS method and in present thought to be the gold standard, but new technologies, such as next-generation sequencing (NGS), continue to emerge. Validation of the new comprehensive NGS-based 24-chromosome aneuploidy screening technology is still needed to determine the preclinical accuracy before it might be considered as an alternative method for human PGS. In the present study, 43 human trophectoderm (TE) biopsy samples and 5 cytogenetically characterized cell lines (Coriell Cell Repositories) were tested. The same whole genome amplified product of each sample was blindly assessed with Veriseq NGS and Agilent aCGH to identify the aneuploidy status. The result showed that the NGS identified all abnormalities identified in aCGH including the numeral chromosomal abnormalities (again or loss) in the embryo samples and the structural (partial deletion and duplication) in the Coriell cell lines. Both technologies can identify a segmental imbalance as small as 1.8 Mb in size. Among the 41 TE samples with abnormal karyotypes in this study, eight (19.5 %) samples presented as multiple chromosome abnormalities. The abnormalities occurred to almost all chromosomes, except chromosome 6, 7, 17 and Y chromosome. Given its reliability and high level of consistency with an established aCGH methodology, NGS has demonstrated a robust high-throughput methodology ready for extensive clinical application in reproductive medicine, with potential advantages of reduced costs and enhanced precision. Then, a randomized controlled clinical trial confirming its clinical effectiveness is advisable to obtain a larger sequencing dataset

  16. Stress response to cadmium and manganese in Paracentrotus lividus developing embryos is mediated by nitric oxide

    Energy Technology Data Exchange (ETDEWEB)

    Migliaccio, Oriana; Castellano, Immacolata [Laboratory of Cellular and Developmental Biology, Stazione Zoologica Anton Dohrn, Villa Comunale, 80121 Naples (Italy); Romano, Giovanna [Laboratory of Functional and Evolutionary Ecology, Stazione Zoologica Anton Dohrn, Villa Comunale, 80121 Naples (Italy); Palumbo, Anna, E-mail: anna.palumbo@szn.it [Laboratory of Cellular and Developmental Biology, Stazione Zoologica Anton Dohrn, Villa Comunale, 80121 Naples (Italy)

    2014-11-15

    Highlights: • NO is produced in sea urchin embryos in response to cadmium and manganese. • Cadmium and manganese affect the expression of specific genes. • NO levels regulate directly or indirectly the expression of some metal-induced genes. • NO is proposed as a sensor of different stress agents in sea urchin embryos. - Abstract: Increasing concentrations of contaminants, often resulting from anthropogenic activities, have been reported to occur in the marine environment and affect marine organisms. Among these, the metal ions cadmium and manganese have been shown to induce developmental delay and abnormalities, mainly reflecting skeleton elongation perturbation, in the sea urchin Paracentrotus lividus, an established model for toxicological studies. Here, we provide evidence that the physiological messenger nitric oxide (NO), formed by L-arginine oxidation by NO synthase (NOS), mediates the stress response induced by cadmium and manganese in sea urchins. When NO levels were lowered by inhibiting NOS, the proportion of abnormal plutei increased. Quantitative expression of a panel of 19 genes involved in stress response, skeletogenesis, detoxification and multidrug efflux processes was followed at different developmental stages and under different conditions: metals alone, metals in the presence of NOS inhibitor, NO donor and NOS inhibitor alone. These data allowed the identification of different classes of genes whose metal-induced transcriptional expression was directly or indirectly mediated by NO. These results open new perspectives on the role of NO as a sensor of different stress agents in sea urchin developing embryos.

  17. Composition of single-step media used for human embryo culture.

    Science.gov (United States)

    Morbeck, Dean E; Baumann, Nikola A; Oglesbee, Devin

    2017-04-01

    To determine compositions of commercial single-step culture media and test with a murine model whether differences in composition are biologically relevant. Experimental laboratory study. University-based laboratory. Inbred female mice were superovulated and mated with outbred male mice. Amino acid, organic acid, and ions content were determined for single-step culture media: CSC, Global, G-TL, and 1-Step. To determine whether differences in composition of these media are biologically relevant, mouse one-cell embryos were cultured for 96 hours in each culture media at 5% and 20% oxygen in a time-lapse incubator. Compositions of four culture media were analyzed for concentrations of 30 amino acids, organic acids, and ions. Blastocysts at 96 hours of culture and cell cycle timings were calculated, and experiments were repeated in triplicate. Of the more than 30 analytes, concentrations of glucose, lactate, pyruvate, amino acids, phosphate, calcium, and magnesium varied in concentrations. Mouse embryos were differentially affected by oxygen in G-TL and 1-Step. Four single-step culture media have compositions that vary notably in pyruvate, lactate, and amino acids. Blastocyst development was affected by culture media and its interaction with oxygen concentration. Copyright © 2017 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  18. PROTECTING EGG DONORS AND HUMAN EMBRYOS-THE FAILURE OF THE SOUTH KOREAN BIOETHICS AND BIOSAFETY ACT

    National Research Council Canada - National Science Library

    Mukta Jhalani

    2008-01-01

    ... Korea passed the Bioethics and Biosafety Act to regulate biotechnology research. In its current form, the Bioethics and Biosafety Act fails to adequately protect egg donors and human embryos. The Bioethics and Biosafety Act does not have adequate safeguards to protect egg donors, such as a requirement of voluntary consent and a requirement tha...

  19. Cryotop vitrification for in vitro produced bovine and buffalo (Bubalus bubalis) embryos at different stages of development

    OpenAIRE

    B. Gasparrini; G. Campanile; D. Vecchio; L. Boccia; L. Attanasio; De Rosa, A.

    2010-01-01

    The aim of this study was to evaluate the possibility to vitrify in vitro produced (IVP) buffalo and bovine embryos at different stages of development by an advanced version of the “minimal volume approaches”: the Cryotop method. In both experiments, the embryos were vitrified at the tight morula (TM), early blastocyst (eBl), blastocyst (Bl), expanded blastocyst (xBl) and, only for buffalo, at the hatched blastocyst (hBl) stage. After warming, the embryos were cultured in vitro fo...

  20. Biopsy of embryos produced by in vitro fertilization affects development in C57BL/6 mouse strain

    OpenAIRE

    Sugawara, Atsushi; Ward, Monika A.

    2012-01-01

    Preimplantation genetic diagnosis (PGD) is considered highly successful in respect to its accuracy in detecting genetic anomalies but the effects of embryo biopsy on embryonic/fetal growth and development are less known, particularly in conjunction with in vitro fertilization (IVF). Here, we compared biopsied (B) and non-biopsied (NB) mouse embryos for their developmental competence. Embryos C57BL/6 (B6) and B6D2F2 (F2) generated by IVF were subjected to single blastomere biopsy at the 4-cell...

  1. In vitro and in vivo Development of Cloned Ovine Embryos using in vitro and in vivo Matured Oocytes

    DEFF Research Database (Denmark)

    Holm, P; Nagashima, H; Sun, F-J

    1995-01-01

    Cloning of sheep embryos by nucleus transplantation can be achieved by using in vivo matured (oviductal) oocytes and in vivo culture. However, these steps involve cumbersome procedures. Therefore, the effects of in vivo vs. the equivalent in vitro procedures on the pre-implantation development...... matured oocytes were enucleated and fused with inserted blastomeres from donor embryos. In vitro matured oocytes were enucleated and allowed to age prior to blastomere insertion and electrofusion. Fused embryos were cultured for approximately 132 h either in vivo in ligated sheep oviducts or in vitro...

  2. Development of the cardiac conduction system in atrioventricular septal defect in human trisomy 21

    NARCIS (Netherlands)

    Blom, Nico A.; Ottenkamp, Jaap; Deruiter, Marco C.; Wenink, Arnold C. G.; Gittenberger-de Groot, Adriana C.

    2005-01-01

    In patients with atrioventricular septal defect (AVSD), the occurrence of nonsurgical AV block has been reported. We have looked for an explanation in the development of the AV conduction system. Human embryos with AVSD and trisomy 21 and normal embryos were examined (age 5-16 wk gestation).

  3. Developmental anatomy of the liver from computerized three-dimensional reconstructions of four human embryos (from Carnegie stage 14 to 23).

    Science.gov (United States)

    Lhuaire, Martin; Tonnelet, Romain; Renard, Yohann; Piardi, Tullio; Sommacale, Daniele; Duparc, Fabrice; Braun, Marc; Labrousse, Marc

    2015-07-01

    Some aspects of human embryogenesis and organogenesis remain unclear, especially concerning the development of the liver and its vasculature. The purpose of this study was to investigate, from a descriptive standpoint, the evolutionary morphogenesis of the human liver and its vasculature by computerized three-dimensional reconstructions of human embryos. Serial histological sections of four human embryos at successive stages of development belonging to three prestigious French historical collections were digitized and reconstructed in 3D using software commonly used in medical radiology. Manual segmentation of the hepatic anatomical regions of interest was performed section by section. In this study, human liver organogenesis was examined at Carnegie stages 14, 18, 21 and 23. Using a descriptive and an analytical method, we showed that these stages correspond to the implementation of the large hepatic vascular patterns (the portal system, the hepatic artery and the hepatic venous system) and the biliary system. To our knowledge, our work is the first descriptive morphological study using 3D computerized reconstructions from serial histological sections of the embryonic development of the human liver between Carnegie stages 14 and 23. Copyright © 2015 Elsevier GmbH. All rights reserved.

  4. Glutathione redox dynamics and expression of glutathione-related genes in the developing embryo

    Science.gov (United States)

    Timme-Laragy, Alicia R.; Goldstone, Jared V.; Imhoff, Barry R.; Stegeman, John J.; Hahn, Mark E.; Hansen, Jason M.

    2013-01-01

    Embryonic development involves dramatic changes in cell proliferation and differentiation that must be highly coordinated and tightly regulated. Cellular redox balance is critical for cell fate decisions, but it is susceptible to disruption by endogenous and exogenous sources of oxidative stress. The most abundant endogenous non-protein antioxidant defense molecule is the tri-peptide glutathione (γ-glutamyl-cysteinylglycine, GSH), but the ontogeny of GSH concentration and redox state during early life stages is poorly understood. Here, we describe the GSH redox dynamics during embryonic and early larval development (0–5 days post-fertilization) in the zebrafish (Danio rerio), a model vertebrate embryo. We measured reduced and oxidized glutathione (GSH, GSSG) using HPLC, and calculated the whole embryo total glutathione (GSHT) concentrations and redox potentials (Eh) over 0–120 hours of zebrafish development (including mature oocytes, fertilization, mid-blastula transition, gastrulation, somitogenesis, pharyngula, pre-hatch embryos, and hatched eleutheroembryos). GSHT concentration doubled between 12 hours post fertilization (hpf) and hatching. The GSH Eh increased, becoming more oxidizing during the first 12 h, and then oscillated around −190 mV through organogenesis, followed by a rapid change, associated with hatching, to a more negative (more reducing) Eh (−220 mV). After hatching, Eh stabilized and remained steady through 120 hpf. The dynamic changes in GSH redox status and concentration defined discrete windows of development: primary organogenesis, organ differentiation, and larval growth. We identified the set of zebrafish genes involved in the synthesis, utilization, and recycling of GSH, including several novel paralogs, and measured how expression of these genes changes during development. Ontogenic changes in the expression of GSH-related genes support the hypothesis that GSH redox state is tightly regulated early in development. This study

  5. The Periconceptional Environment and Cardiovascular Disease: Does In Vitro Embryo Culture and Transfer Influence Cardiovascular Development and Health?

    Directory of Open Access Journals (Sweden)

    Monalisa Padhee

    2015-02-01

    Full Text Available Assisted Reproductive Technologies (ARTs have revolutionised reproductive medicine; however, reports assessing the effects of ARTs have raised concerns about the immediate and long-term health outcomes of the children conceived through ARTs. ARTs include manipulations during the periconceptional period, which coincides with an environmentally sensitive period of gamete/embryo development and as such may alter cardiovascular development and health of the offspring in postnatal life. In order to identify the association between ARTs and cardiovascular health outcomes, it is important to understand the events that occur during the periconceptional period and how they are affected by procedures involved in ARTs. This review will highlight the emerging evidence implicating adverse cardiovascular outcomes before and after birth in offspring conceived through ARTs in both human and animal studies. In addition, it will identify the potential underlying causes and molecular mechanisms responsible for the congenital and adult cardiovascular dysfunctions in offspring whom were conceived through ARTs.

  6. Transmission of Campylobacter coli in chicken embryos

    Directory of Open Access Journals (Sweden)

    Daise Aparecida Rossi

    2012-06-01

    Full Text Available 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.

  7. Effect of Mobile Phone Radiation on Cardiovascular Development of Chick Embryo.

    Science.gov (United States)

    Ye, W; Wang, F; Zhang, W; Fang, N; Zhao, W; Wang, J

    2016-06-01

    The biological effects on cardiovascular development of chicken embryos were examined after radiation exposure using mobile phone (900 MHz; specific absorption rate˜1.07 W/kg) intermittently 3 h per day during incubation. Samples were selected by morphological and histological methods. The results showed the rate of embryonic mortality and cardiac deformity increased significantly in exposed group (P findings suggest that long-term exposure of MPR may induce myocardium pathological changes, DNA damage and increased mortality; however, there was little effect on vascular development. © 2015 Blackwell Verlag GmbH.

  8. The impact of nutrition of the cumulus oocyte complex and embryo on subsequent development in ruminants.

    Science.gov (United States)

    Thompson, Jeremy G

    2006-02-01

    Cumulus-oocyte complexes (COCs) and early embryos rely on a histotrophic nutrition source for energy production and the synthesis of macromolecules. There is accumulating evidence suggesting that the balance of supply and demand for energy and other anabolic substrates during oocyte maturation and very early stages of development programmes subsequent developmental potential, and this may include subsequent fetal growth trajectory. One example is the role of glucose (Glc) during cumulus-oocyte complex maturation. Glucose is an essential nutrient for maturation, especially its role during cumulus expansion. Our laboratory has shown that during in vitro culture, too little glucose during cumulus-oocyte complex maturation affects meiotic competence. We have focussed on glucose (Glc) metabolism through the hexosamine biosynthesis pathway (HBP) during COC maturation in vitro. The HBP in somatic cells is regarded as a "fuel-sensing" pathway and its interaction with cell signalling systems and transcriptional regulation is increasingly apparent. Up-regulation of the HBP during oocyte maturation in vitro has negative consequences for subsequent development. Another example is the role of hypoxia (low O2) during peri-compaction development. My laboratory believes that ruminant embryos during compaction, blastulation and subsequent development in the uterine cavity lack a key hypoxia responsive element. Because of this, hypoxia is important for normal development in ruminants but perturbs further development in rodents. The implication of these examples to the fundamental concept of peri-conception nutritional programming of development are discussed.

  9. Impact of protamine transcripts and their proteins on the quality and fertilization ability of sperm and the development of preimplantation embryos.

    Science.gov (United States)

    Depa-Martynow, Magdalena; Kempisty, Bartosz; Jagodziński, Paweł Piotr; Pawelczyk, Leszek; Jedrzejczak, Piotr

    2012-03-01

    Protamine 1 (PRM1) and 2 (PRM2) are major nuclear proteins in spermatozoa known to bind to chromatin during early spermatogenesis. The aim of this study was to evaluate the impact of protamine transcripts and proteins on human spermatozoa and their fertilization ability as well as the development of preimplantation embryos. Oocytes selected from 92 patients were fertilized in vitro (IVF) using their partners' sperm after evaluating its concentration, motility and morphology. Reverse transcription and real-time quantitative PCR (RQ-PCR) were applied to determine protamine mRNA level, while sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting were used to quantify the protamine protein concentration. Next, protamine mRNA and protein levels were correlated with sperm concentration, motility and morphology as well as with the fertilization ability of mature spermatozoa and embryos of different quality obtained from the IVF program. A significant correlation has been found between protamine transcripts/proteins and: sperm concentration, progressive sperm motility, sperm fertilization ability as well as embryo quality. Protamine transcripts and proteins contribute to the quality of spermatozoa, successful fertilization and may have a significant influence on the development of preimplantation embryos.

  10. Should intrauterine human chorionic gonadotropin infusions ever be used prior to embryo transfer?

    Science.gov (United States)

    Volovsky, Michelle; Healey, Martin; MacLachlan, Vivien; Vollenhoven, Beverley J

    2017-09-25

    The aim of this study was to explore the factors that influence the outcome of intrauterine human chorionic gonadotropin (hCG) infusion at the time of embryo transfer (ET), in particular, the effect of hCG infusions on fresh and frozen embryo transfers (FETs) and whether prior recurrent implantation failure (RIF) impacts upon outcomes. This was a case-control study based on a standardized database from a multi-site in vitro fertilization clinic. The analysis contains 458 cases and 749 matched controls, with an intervention group of those given intrauterine hCG prior to ET and a control group of patients receiving no hCG infusion. Outcomes were defined as clinical pregnancy and live birth rates. Two analyses were performed. The first separated FETs (cases n = 224, controls n = 325) and fresh ETs (cases n = 234, controls n = 424), with outcomes calculated in each group. The second analysis divided patients into those with RIF (cases n = 149, controls n = 200) and those without (cases n = 309, controls n = 549). Results in fresh ETs demonstrated a 5.8% reduction (adjusted odds ratio (AOR) = 0.60, p = 0.041) in clinical pregnancy rates with the use of intrauterine hCG. In those without defined RIF, clinical pregnancy rates were reduced by 8.1% (AOR = 0.61, p = 0.023) and live birth rates by 7.2% (AOR = 0.56, p = 0.32) with intrauterine hCG use. There were no significant differences in outcomes in FETs and in the RIF cohort. Intrauterine hCG at the time of ET not only seems to have no benefit, but rather a negative effect in fresh ETs and those without RIF.

  11. A predictive model for blastocyst formation based on morphokinetic parameters in time-lapse monitoring of embryo development.

    Science.gov (United States)

    Milewski, Robert; Kuć, Paweł; Kuczyńska, Agnieszka; Stankiewicz, Bożena; Łukaszuk, Krzysztof; Kuczyński, Waldemar

    2015-04-01

    The aim of the study was to create a predictive model of blastocyst development based on morphokinetic parameters of time-lapse embryoscope monitoring. Time-lapse recordings of 432 embryos (obtained from 77 patients), monitored in Embryoscope, were involved in the study. Patients underwent in vitro fertilization according to standard procedure between June 2012 and April 2013. A retrospective analysis of morphokinetic features, focused on duration of time from the Intracytoplasmic Sperm Injection (ICSI) procedure to consecutive embryo division for 2, 3, 4 and 5 blastomeres, as well as time intervals between each division, was conducted. All embryos were observed for 5 days. Based on the distribution of analyzed morphokinetic parameters and number of embryos developed to blastocyst, a range denoting the possibility of an embryo reaching blastocyst stage was determined. According to the obtained results, univariate and multivariate logistic regression analyses were performed. Based on the times of division for two and five blastomeres and intervals between the second and third division, a multivariate predictive model was created. The predictive equation was constructed based on the parameters of logistic regression analysis (odds ratios). Statistically significant differences (p prediction parameter between the group of embryos developed to blastocyst (the median value: Me = 9.95, and quartiles: Q1 = 7.59, Q3 = 12.30) and embryos that did not develop to the blastocyst stage (Me = 4.66, Q1 = 2.33, Q3 = 8.19) were found. A Receiver Operating Characteristic (ROC) curve was created for the constructed predictive model. The Area Under the Curve was AUC = 0.806 with a 95 % confidence interval (0.747, 0.864). The predictive model constructed in this study has been validated using an independent data set, which indicates that the model is reliable and repeatable. Time-lapse imaging presents a new diagnostic tool for parametric evaluation of

  12. Potential hazards to embryo implantation: A human endometrial in vitro model to identify unwanted antigestagenic actions of chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, L.; Deppert, W.R. [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany); Pfeifer, D. [Department of Hematology and Oncology, University Hospital Freiburg (Germany); Stanzel, S.; Weimer, M. [Department of Biostatistics, German Cancer Research Center, Heidelberg (Germany); Hanjalic-Beck, A.; Stein, A.; Straßer, M.; Zahradnik, H.P. [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany); Schaefer, W.R., E-mail: wolfgang.schaefer@uniklinik-freiburg.de [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany)

    2012-05-01

    Embryo implantation is a crucial step in human reproduction and depends on the timely development of a receptive endometrium. The human endometrium is unique among adult tissues due to its dynamic alterations during each menstrual cycle. It hosts the implantation process which is governed by progesterone, whereas 17β-estradiol regulates the preceding proliferation of the endometrium. The receptors for both steroids are targets for drugs and endocrine disrupting chemicals. Chemicals with unwanted antigestagenic actions are potentially hazardous to embryo implantation since many pharmaceutical antiprogestins adversely affect endometrial receptivity. This risk can be addressed by human tissue-specific in vitro assays. As working basis we compiled data on chemicals interacting with the PR. In our experimental work, we developed a flexible in vitro model based on human endometrial Ishikawa cells. Effects of antiprogestin compounds on pre-selected target genes were characterized by sigmoidal concentration–response curves obtained by RT-qPCR. The estrogen sulfotransferase (SULT1E1) was identified as the most responsive target gene by microarray analysis. The agonistic effect of progesterone on SULT1E1 mRNA was concentration-dependently antagonized by RU486 (mifepristone) and ZK137316 and, with lower potency, by 4-nonylphenol, bisphenol A and apigenin. The negative control methyl acetoacetate showed no effect. The effects of progesterone and RU486 were confirmed on the protein level by Western blotting. We demonstrated proof of principle that our Ishikawa model is suitable to study quantitatively effects of antiprogestin-like chemicals on endometrial target genes in comparison to pharmaceutical reference compounds. This test is useful for hazard identification and may contribute to reduce animal studies. -- Highlights: ► We compare progesterone receptor-mediated endometrial effects of chemicals and drugs. ► 4-Nonylphenol, bisphenol A and apigenin exert weak

  13. In vitro maturation and embryo development of bovine oocytes after meiosis blockage with MPF inhibitors

    Directory of Open Access Journals (Sweden)

    Mariana Groke Marques

    2011-12-01

    Full Text Available This study evaluated the bovine oocyte maturation and embryo development after in vitro fertilization. The maturation of the oocytes was blocked using Butyrolactone I and Roscovitine using pre-maturation medium supplemented with fetal calf serum (FCS. The ocytes were divided in four groups: Control 0 hour, Control (24 hours of maturation, Roscovitine (maturation blockage with 50mM Roscovitine during 24 hours followed by 24 hours of maturation, and Butyrolactone I (maturation blockage with 150mM Butyrolactone I during 24 hours followed by 24 hours of maturation. The oocytes were fixed and stained with aceto orcein to evaluate the nuclear maturation. After the maturation period, the remaining oocytes of the Control group, Roscovitine, and Butyrolactone I were fertilized in vitro. Embryo development was assessed by the cleavage rate (D3 and blastocysts formation (D7. The Butyrolactone I group had similar rates of germinal vesical stage oocytes during blockage, and Metaphase 2 after maturation, comparing to Control group at 0 hour and Control group, respectively. On the other hand, the Roscovitine group had lower rates of vesical stage oocytes during blockage, and Metaphase 2 after maturation comparing to Control groups. After in vitro fertilization, higher rates of cleavage were observed in Control and Butyrolactone I groups. For the blastocyst formation rate, the Control group showed better results than Roscovitine group. In summary, Butyrolactone I group had similar results to the Control group, and for this reason, is suitable for pre-maturation of bovine oocytes using FCS. In contrast, Roscovitine group had lower oocyte maturation and embryo development.

  14. Functional divergence of chloroplast Cpn60α subunits during Arabidopsis embryo development.

    Directory of Open Access Journals (Sweden)

    Xiaolong Ke

    2017-09-01

    Full Text Available Chaperonins are a class of molecular chaperones that assist in the folding and assembly of a wide range of substrates. In plants, chloroplast chaperonins are composed of two different types of subunits, Cpn60α and Cpn60β, and duplication of Cpn60α and Cpn60β genes occurs in a high proportion of plants. However, the importance of multiple Cpn60α and Cpn60β genes in plants is poorly understood. In this study, we found that loss-of-function of CPNA2 (AtCpn60α2, a gene encoding the minor Cpn60α subunit in Arabidopsis thaliana, resulted in arrested embryo development at the globular stage, whereas the other AtCpn60α gene encoding the dominant Cpn60α subunit, CPNA1 (AtCpn60α1, mainly affected embryonic cotyledon development at the torpedo stage and thereafter. Further studies demonstrated that CPNA2 can form a functional chaperonin with CPNB2 (AtCpn60β2 and CPNB3 (AtCpn60β3, while the functional partners of CPNA1 are CPNB1 (AtCpn60β1 and CPNB2. We also revealed that the functional chaperonin containing CPNA2 could assist the folding of a specific substrate, KASI (β-ketoacyl-[acyl carrier protein] synthase I, and that the KASI protein level was remarkably reduced due to loss-of-function of CPNA2. Furthermore, the reduction in the KASI protein level was shown to be the possible cause for the arrest of cpna2 embryos. Our findings indicate that the two Cpn60α subunits in Arabidopsis play different roles during embryo development through forming distinct chaperonins with specific AtCpn60β to assist the folding of particular substrates, thus providing novel insights into functional divergence of Cpn60α subunits in plants.

  15. Assessment of the long-term and transgenerational consequences of perturbing preimplantation embryo development in mice.

    Science.gov (United States)

    Mahsoudi, B; Li, A; O'Neill, C

    2007-11-01

    Perturbations of the development of preimplantation embryos may have long-term consequences for the health of progeny. There are no standardized methods for assessing such risks. The OECD/OCDE 416 Guideline for Testing of Chemicals (Two-Generation Reproduction Toxicity Study) is a standardized assay for detecting potential toxic effects of chemicals. The present study assessed the utility of this guideline for identifying long-term consequences of perturbing preimplantation development. Extended culturing of mammalian zygotes commonly results in retarded preimplantation development. Mouse zygotes were cultured in vitro for 96 h until the blastocyst stage (cultured blastocysts) or blastocysts were collected from the Day-3.5 uterus (in vivo blastocysts). The resulting blastocysts were transferred to the uteri of pseudopregnant recipients (P generation). Progeny from both treatments were mated for a further two generations (F1 and F2 generations). There was no effect of treatment group on gross fertility across the generations tested. Progeny of the cultured blastocysts had lower body weights to the time of weaning compared to in vivo blastocysts in the P and F1 generations, but not in the F2 generation. At maturity, there was no effect of treatment group on body weight, although thyroid weight was higher in the in vivo blastocyst group in the P generation, while the brain, pituitary, and kidneys were larger in the progeny of the cultured blastocysts of the F1 generation. The OECD/OCDE 416 assessment may have a role as a standardized test for the assessment of the biological consequences of perturbing the growth environment of the preimplantation embryo. Embryo culture influenced the somatometric parameters of the resulting progeny, some of which were maintained across a generation.

  16. Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos

    OpenAIRE

    Famiglietti, Amber L.; Wei, Zheng; Beres, Thomas M.; Milac, Adina L.; Tran, Duy T.; Patel, Divya; Angerer, Robert C.; Angerer, Lynne M.; Tabak, Lawrence A.

    2017-01-01

    Mucin-type O-glycosylation is a ubiquitous posttranslational modification in which N-Acetylgalactosamine (GalNAc) is added to the hydroxyl group of select serine or threonine residues of a protein by the family of UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferases (GalNAc-Ts; EC 2.4.1.41). Previous studies demonstrate that O-glycosylation plays essential roles in protein function, cell-cell interactions, cell polarity and differentiation in developing mouse and Drosophila embryos. Alth...

  17. Altered cleavage patterns in human tripronuclear embryos and their association to fertilization method

    DEFF Research Database (Denmark)

    Joergensen, Mette Warming; Agerholm, Inge; Hindkjaer, Johnny

    2014-01-01

    PURPOSE: To analyze the cleavage patterns in dipronuclear (2PN) and tripronuclear (3PN) embryos in relation to fertilization method. METHOD: Time-lapse analysis. RESULTS: Compared to 2PN, more 3PN IVF embryos displayed early cleavage into 3 cells (p < 0.001), displayed longer duration of the 3-ce...

  18. Early development of Drosophila embryos requires Smc5/6 function during oogenesis

    Directory of Open Access Journals (Sweden)

    Martin Tran

    2016-07-01

    Full Text Available Mutations in structural maintenance of chromosomes (Smc proteins are frequently associated with chromosomal abnormalities commonly observed in developmental disorders. However, the role of Smc proteins in development still remains elusive. To investigate Smc5/6 function during early embryogenesis we examined smc5 and smc6 mutants of the fruit fly Drosophila melanogaster using a combination of reverse genetics and microscopy approaches. Smc5/6 exhibited a maternally contributed function in maintaining chromosome stability during early embryo development, which manifested as female subfertility in its absence. Loss of Smc5/6 caused an arrest and a considerable delay in embryo development accompanied by fragmented nuclei and increased anaphase-bridge formation, respectively. Surprisingly, early embryonic arrest was attributable to the absence of Smc5/6 during oogenesis, which resulted in insufficient repair of pre-meiotic and meiotic DNA double-strand breaks. Thus, our findings contribute to the understanding of Smc proteins in higher eukaryotic development by highlighting a maternal function in chromosome maintenance and a link between oogenesis and early embryogenesis.

  19. Early development of Drosophila embryos requires Smc5/6 function during oogenesis.

    Science.gov (United States)

    Tran, Martin; Tsarouhas, Vasilios; Kegel, Andreas

    2016-07-15

    Mutations in structural maintenance of chromosomes (Smc) proteins are frequently associated with chromosomal abnormalities commonly observed in developmental disorders. However, the role of Smc proteins in development still remains elusive. To investigate Smc5/6 function during early embryogenesis we examined smc5 and smc6 mutants of the fruit fly Drosophila melanogaster using a combination of reverse genetics and microscopy approaches. Smc5/6 exhibited a maternally contributed function in maintaining chromosome stability during early embryo development, which manifested as female subfertility in its absence. Loss of Smc5/6 caused an arrest and a considerable delay in embryo development accompanied by fragmented nuclei and increased anaphase-bridge formation, respectively. Surprisingly, early embryonic arrest was attributable to the absence of Smc5/6 during oogenesis, which resulted in insufficient repair of pre-meiotic and meiotic DNA double-strand breaks. Thus, our findings contribute to the understanding of Smc proteins in higher eukaryotic development by highlighting a maternal function in chromosome maintenance and a link between oogenesis and early embryogenesis. © 2016. Published by The Company of Biologists Ltd.

  20. Embryo development after mitochondrial supplementation from induced pluripotent stem cells.

    Science.gov (United States)

    Li, Ruiqi; Wen, Bingqiang; Zhao, Haijing; Ouyang, Nengyong; Ou, Songbang; Wang, Wenjun; Han, Jianyong; Yang, Dongzi

    2017-08-01

    The purpose of this study was to evaluate the effects of mitochondrial supplementation (MS) on early embryonic development and to assess the safety of MS treatments using induced pluripotent stem cells (iPSCs) as the mitochondrial donor. In this study, we evaluated the effect of MS on early embryonic development using induced pluripotent stem cells (iPSCs) as the donor. Mouse zygotes were injected with either mitochondria from iPSCs or a vehicle solution. Several parameters were evaluated, including the rates of blastocyst formation and implantation, the weight of E13.5 embryos and placentas, the distribution of the donor mitochondrial DNA (mtDNA), and the pattern of methylation in the differentially methylated regions (DMRs) of the H19 and Snrpn genes. We found that neither the rates of blastocyst formation and implantation nor the weights of E13.5 embryos and placentas were significantly different between the MS and control groups. Additionally, the mtDNA from the iPSC donors could be detected in the muscle tissue of four fetuses and all placentas in the MS group. Finally, the methylation patterns of H19 and Snrpn DMRs remained unchanged by MS. iPSC-derived mtDNA was directly involved in the process of embryonic development after MS. No adverse effects were seen when using iPSCs as a mitochondrial donor, but it remains to be seen whether this method can improve embryonic development, especially in older mice.

  1. Effect of selection for productive traits in broiler maternal lines on embryo development

    Directory of Open Access Journals (Sweden)

    Schmidt GS

    2003-01-01

    Full Text Available This study used 300 females and 30 males with 36 weeks of age from the selected PP and control PPc maternal broiler lines. PP has been selected for heavy body weight (PC and high egg production for eight generations. Fertile eggs were collected and weighed individually for 4 periods of 5 consecutive days at two-week intervals. In each period, a total of 960 eggs/line were identified and separated in groups of 240 eggs, and stored for later incubation. Embryo weight (PE was evaluated at 9 (P9, 11 (P11, 13 (P13, 15 (P15, 17 (P17 and 21 (P21 days of incubation. The objective was to estimate the effect of selection on embryo development. Egg weight (PO was similar between the two lines. The differences in PE were significant from P15 on, resulting in 1.9g of difference in the chick weight, indicating correlated genetic changes in the embryo development, which can be credited to the selection for PC. Changes in PE while PO was kept unaltered modified the correlations between these two traits. Differences were significant from P13 on and estimated correlations for P21 were 0.72 and 0.70 for PP and PPc, respectively. Chick weight corresponded to 70.91% (PP and 68.48% (PPc of egg weight. The estimated increase in P21 that resulted from the increase of 1.0g in PO was 0.71 in PP and 0.68g in PPc.

  2. Post-Transcriptional Control of Gene Expression in Mouse Early Embryo Development: A View from the Tip of the Iceberg

    Directory of Open Access Journals (Sweden)

    Claudio Sette

    2011-04-01

    Full Text Available Fertilization is a very complex biological process that requires the perfect cooperation between two highly specialized cells: the male and female gametes. The oocyte provides the physical space where this process takes place, most of the energetic need, and half of the genetic contribution. The spermatozoon mostly contributes the other half of the chromosomes and it is specialized to reach and to penetrate the oocyte. Notably, the mouse oocyte and early embryo are transcriptionally inactive. Hence, they fully depend on the maternal mRNAs and proteins stored during oocyte maturation to drive the onset of development. The new embryo develops autonomously around the four-cell stage, when maternal supplies are exhausted and the zygotic genome is activated in mice. This oocyte-to-embryo transition needs an efficient and tightly regulated translation of the maternally-inherited mRNAs, which likely contributes to embryonic genome activation. Full understanding of post-transcriptional regulation of gene expression in early embryos is crucial to understand the reprogramming of the embryonic genome, it might help driving reprogramming of stem cells in vitro and will likely improve in vitro culturing of mammalian embryos for assisted reproduction. Nevertheless, the knowledge of the mechanism(s underlying this fundamental step in embryogenesis is still scarce, especially if compared to other model organisms. We will review here the current knowledge on the post-transcriptional control of gene expression in mouse early embryos and discuss some of the unanswered questions concerning this fascinating field of biology.

  3. Perkembangan Praimplantasi Embrio Mencit dengan Materi Genetik yang Berasal dari Parental, Maternal, dan Inti Sel Somatik (PRE-IMPLANTATION DEVELOPMENT OF MOUSE EMBRYO WITH GENETIC MATERIAL DERIVED FROM PARENTAL, MATERNAL AND SOMATIC CELL NUCLEUS

    Directory of Open Access Journals (Sweden)

    Harry Murti

    2014-05-01

    Full Text Available Cloned embryo and parthenogenetic embryo are a potential source of stem cells for regenerativemedicine. Stem cells derived from those embryos are expected to overcome the ethical issues to the use offertilization embryos for therapeutic purposes. The pre-implantation development is a critical step fordeveloping embryos reach the blastocyst stage. The objectives in vivo of this research are to produce mousecloned embryo, parthenogenetic embryo, and fertilized embryo and to study stages of  in vitro pre-implantation development culture. In vivo fertilized embryos, mouse oocytes, and cumulus cells were usedin this study. Treatment was performed on female mice superovulated with PMSG and hCG injections.Two-cell stage of in vivo fertilized embryos were collected on the second day post hCG injection. Clonedembryos were produced through Somatic Cell Nuclear Transfer (SCNT, which included enucleation, nucleartransfer and artificial activation. Parthenogenetic embryos were produced with artificial activationtechnique. The result of the research indicated that SCNT application was able to produce cloned embryos which could develop to blastocyst stage (3,2%. In addition, artificial activation of oocytes could produceparthenogenetic embryos which were able to develop up to the blastocyst stage (8,6%. In conclusion,efficiency level of parthenogenetic embryos that is able to reach the blastocyst stage was higher than in thecloned embryos. Fertilized embryos shows a better development and more efficient compared to in vitrocloned embryos and parthenogenetic embryos cultures.

  4. Pias1 is essential for erythroid and vascular development in the mouse embryo.

    Science.gov (United States)

    Constanzo, Jerfiz D; Deng, Mi; Rindhe, Smita; Tang, Ke-Jing; Zhang, Cheng-Cheng; Scaglioni, Pier Paolo

    2016-07-01

    The protein inhibitor of activated STAT-1 (PIAS1) is one of the few known SUMO E3 ligases. PIAS1 has been implicated in several biological processes including repression of innate immunity and DNA repair. However, PIAS1 function during development and tissue differentiation has not been studied. Here, we report that Pias1 is required for proper embryonic development. Approximately 90% of Pias1 null embryos die in utero between E10.5 and E12.5. We found significant apoptosis within the yolk sac (YS) blood vessels and concomitant loss of red blood cells (RBCs) resulting in profound anemia. In addition, Pias1 loss impairs YS angiogenesis and results in defective capillary plexus formation and blood vessel occlusions. Moreover, heart development is impaired as a result of loss of myocardium muscle mass. Accordingly, we found that Pias1 expression in primary myoblasts enhances the induction of cardiac muscle genes MyoD, Myogenin and Myomaker. PIAS1 protein regulation of cardiac gene transcription is dependent on transcription factors Myocardin and Gata-4. Finally, endothelial cell specific inactivation of Pias1 in vivo impairs YS erythrogenesis, angiogenesis and recapitulates loss of myocardium muscle mass. However, these defects are not sufficient to recapitulate the lethal phenotype of Pias1 null embryos. These findings highlight Pias1 as an essential gene for YS erythropoiesis and vasculogenesis in vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Functional studies of signaling pathways in peri-implantation development of the mouse embryo by RNAi

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    Bell Graham

    2005-12-01

    Full Text Available Abstract Background Studies of gene function in the mouse have relied mainly on gene targeting via homologous recombination. However, this approach is difficult to apply in specific windows of time, and to simultaneously knock-down multiple genes. Here we report an efficient method for dsRNA-mediated gene silencing in late cleavage-stage mouse embryos that permits examination of phenotypes at post-implantation stages. Results We show that introduction of Bmp4 dsRNA into intact blastocysts by electroporation recapitulates the genetic Bmp4 null phenotype at gastrulation. It also reveals a novel role for Bmp4 in the regulation the anterior visceral endoderm specific gene expression and its positioning. We also show that RNAi can be used to simultaneously target several genes. When applied to the three murine isoforms of Dishevelled, it leads to earlier defects than previously observed in double knock-outs. These include severe delays in post-implantation development and defects in the anterior midline and neural folds at headfold stages. Conclusion Our results indicate that the BMP4 signalling pathway contributes to the development of the anterior visceral endoderm, and reveal an early functional redundancy between the products of the murine Dishevelled genes. The proposed approach constitutes a powerful tool to screen the functions of genes that govern the development of the mouse embryo.

  6. Effect of cysteamine supplementation of in vitro matured bovine oocytes on chilling sensitivity and development of embryos.

    Science.gov (United States)

    Balasubramanian, S; Rho, Gyu-Jin

    2007-04-01

    In vitro techniques for production of bovine embryos including in vitro oocyte maturation (IVM), fertilization (IVF) and culture (IVC) are becoming increasingly employed for a variety of research purposes. However, decreased viability following cryopreservation by conventional methods has limited commercial applications of these technologies. A practical alternative to facilitate transport would be to arrest development by chilling without freezing. The present research was undertaken to evaluate chilling sensitivity of IVM-IVF embryos at different stages of development, and to determine possible beneficial effects of cysteamine treatment during IVM, previously shown to enhance embryo development in culture, on survival following chilling at different stages. Embryos produced by standard IVM-IVF-IVC methods were chilled to 0 degrees C for 30 min at 2-cell (30-34 h post-insemination, hpi), 8-cell (48-52 hpi) or blastocyst (166-170 hpi) stages. Viability after chilling was assessed by IVC with development to expanded blastocyst stage determined on days 7 and 8 post-insemination (pi) and hatching blastocyst stage determined on days 9 and 10 pi. Control embryos at the same stages were handled similarly, but without chilling, and development during culture similarly assessed. The effect of cysteamine supplementation (100 microM) of the IVM medium was determined for both chilled and non-chilled (control) embryos. Cysteamine supplementation during IVM had no significant effect on oocyte maturation or fertilization, but increased the proportions of oocytes developing to blastocyst stage by day 7 (13.7+/-0.9% versus 7.2+/-0.9%; Pchilling of blastocysts produced by IVM-IVF of oocytes matured in media supplemented with cysteamine offers promise for applications requiring short-term storage to facilitate transport of in vitro produced bovine embryos.

  7. Effect of sperm DNA fragmentation on embryo development: clinical and biological aspects.

    Science.gov (United States)

    Alvarez Sedó, Cristian; Bilinski, Melina; Lorenzi, Daniela; Uriondo, Heydy; Noblía, Felicitas; Longobucco, Valeria; Lagar, Estefanía Ventimiglia; Nodar, Florencia

    2017-12-01

    The aim of this study was to investigate the effect of sperm DNA fragmentation on fertilization rate, embryo development (blastulation rate), and pregnancy outcomes for ICSI cycles performed in a cohort of couples using donor eggs and to assess the remaining embryos that were not transferred or frozen for apoptotic markers. Eighty-two women (egg recipients) were included in the study (2016) were included in the study. The recipients' mean age was 41.8±5.1 y/o (36-49), while the egg donors' mean age was 30.8±2.1 y/o (27-33). Even though donor egg cycles with frozen sperm samples are performed regularly in our center, 35 cycles were done using fresh sperm samples. The mean age of the males involved in the procedure was 40.1±5.2 y/o. Fertilization, blastulation, and pregnancy rates were assessed. The patients were divided into two groups, TUNEL fragmentation and blastulation rate. High levels of DNA fragmentation were associated with low blastulation and pregnancy rates (per transfer); however, fertilization rate was not affected. Samples with higher levels of DNA fragmentation were associated with higher levels of DNA fragmentation in blastomeres without activating the apoptotic pathway (9.1% vs. 15.9%) (pfragmentation activated the apoptotic pathway in higher levels than samples with TUNEL fragmentation was negatively correlated with blastulation and pregnancy rates even in good quality oocytes. High levels of DNA damage promote embryo arrest and induce the activation of the apoptotic pathway.

  8. Effect of estrous cow serum during bovine embryo culture on blastocyst development and cryotolerance after slow freezing or vitrification.

    Science.gov (United States)

    Mucci, N; Aller, J; Kaiser, G G; Hozbor, F; Cabodevila, J; Alberio, R H

    2006-05-01

    The present study investigated the effect of estrous cow serum (ECS) during culture of bovine embryos on blastocyst development and survival after cryopreservation by slow freezing or vitrification. Embryos were derived from in vitro maturation (IVM) and in vitro fertilization (IVF) of abbatoir-derived oocytes. At Day 3, embryos were cultured in three different media: Charles Ronsenkrans medium + amino acids (CR1aa; without bovine serum albumin (BSA)) + 5% estrous cow serum (CR1-ECS), CR1aa + 3 mg/mL BSA (CR1-BSA) or CR1aa + 5% ECS + 3 mg/mL BSA (CR1-ECS-BSA). At 7.5 d post-insemination (PI), blastocyst yield and quality were evaluated; blastocysts and expanded blastocysts from each media were cryopreserved by Open Pulled Straw (OPS) vitrification method or slow freezing (1.5 M ethylene glycol, EM). Total blastocyst yield did not differ among CR1-ECS, CR1-BSA and CR1-ECS-BSA (30.9, 33.1 and 32.9%, respectively, P Embryo survival (hatching rate) was higher in vitrified versus slow-frozen embryos (43% versus 12%, respectively, P embryos cultured in CR1-BSA (40.3%) compared with those cultured in serum-containing media (CR1-ECS, 21.5% and CR1-ECS-BSA, 19.8%; P produce in vitro bovine embryos in serum-free culture medium without affecting blastocyst yield and quality; (b) serum-free medium produced the best quality embryos (in terms of post-cryopreservation survival); and (c) vitrification yielded the highest post-cryopreservation survival rates, regardless of the presence of serum in the culture medium.

  9. Cell wall invertase as a regulator in determining sequential development of endosperm and embryo through glucose signaling early in seed development.

    Science.gov (United States)

    Wang, Lu; Liao, Shengjin; Ruan, Yong-Ling

    2013-01-01

    Seed development depends on coordination among embryo, endosperm and seed coat. Endosperm undergoes nuclear division soon after fertilization, whereas embryo remains quiescent for a while. Such a developmental sequence is of great importance for proper seed development. However, the underlying mechanism remains unclear. Recent results on the cellular domain- and stage-specific expression of invertase genes in cotton and Arabidopsis revealed that cell wall invertase may positively and specifically regulate nuclear division of endosperm after fertilization, thereby playing a role in determining the sequential development of endosperm and embryo, probably through glucose signaling.

  10. Comparison of Starch and ADP-Glucose Pyrophosphorylase Levels in Nonembryogenic Cells and Developing Embryos from Induced Carrot Cultures.

    Science.gov (United States)

    Keller, G L; Nikolau, B J; Ulrich, T H; Wurtele, E S

    1988-02-01

    Cultures of carrot (Daucus carota L.) in a medium without added 2,4-dichlorophenoxyacetic acid were separated into fractions of embryos at different stages of development (large globular and heart, torpedo, and germinating) and nonembryogenic cells. The average starch content per cell in these fractions was similar. However, due to the smaller sizes of the cells of the embryos relative to the nonembryogenic cells, starch content per weight of tissue was higher in the embryos. The ADP-glucose pyrophosphorylase activity per cell in the nonembryogenic cells was double that of the embryo cells. Furthermore, the ratio of ADP-glucose pyrophosphorylase to starch was over 2-fold higher in the nonembryogenic cells, indicating that starch content is not simply determined by ADP-glucose pyrophosphorylase levels. ADP-glucose pyrophosphorylase activity of all culture fractions was directly proportional to the level of a single 50 kilodalton polypeptide detected by immunoblot analysis, using antiserum raised to the purified spinach leaf enzyme. In the same immunoblot analysis, novel polypeptides of 63 and 100 kilodalton were detected in embryos but were absent from nonembryogenic cells. This is one of the few reported examples of specific proteins which differentially accumulate in embryos and nonembryogenic cells.

  11. Influences of somatic donor cell sex on and embryo development following somatic cell nuclear transfer in pigs

    Directory of Open Access Journals (Sweden)

    Jae-Gyu Yoo

    2017-04-01

    Full Text Available Objective The present study investigates pre- and post-implantation developmental competence of nuclear-transferred porcine embryos derived from male and female fetal fibroblasts. Methods Male and female fetal fibroblasts were transferred to in vitro-matured enucleated oocytes and in vitro and in vivo developmental competence of reconstructed embryos was investigated. And, a total of 6,789 female fibroblast nuclear-transferred embryos were surgically transferred into 41 surrogate gilts and 4,746 male fibroblast nuclear-transferred embryos were surgically transferred into 25 surrogate gilts. Results The competence to develop into blastocysts was not significantly different between the sexes. The mean cell number of female and male cloned blastocysts obtained by in vivo culture (143.8±10.5 to 159.2±14.8 was higher than that of in vitro culture of somatic cell nuclear transfer (SCNT groups (31.4±8.3 to 33.4±11.1. After embryo transfer, 5 pregnant gilts from each treatment delivered 15 female and 22 male piglets. The average birth weight of the cloned piglets, gestation length, and the postnatal survival rates were not significantly different (p<0.05 between sexes. Conclusion The present study found that the sex difference of the nuclear donor does not affect the developmental rate of porcine SCNT embryos. Furthermore, postnatal survivability of the cloned piglets was not affected by the sex of the donor cell.

  12. Cell and tissue-autonomous development of the circadian clock in mouse embryos.

    Science.gov (United States)

    Inada, Yutaka; Uchida, Hitoshi; Umemura, Yasuhiro; Nakamura, Wataru; Sakai, Takayoshi; Koike, Nobuya; Yagita, Kazuhiro

    2014-01-31

    The emergence of the circadian rhythm is a dramatic and physiologically essential event for mammals to adapt to daily environmental cycles. It has been demonstrated that circadian rhythms develop during the embryonic stage even when the maternal central pacemaker suprachiasmatic nucleus has been disrupted. However, the mechanisms controlling development of the circadian clock are not yet fully understood. Here, we show that the circadian molecular oscillation in primary dispersed embryonic cells and explanted salivary glands obtained from mPER2(Luc) mice embryos developed cell- or tissue-autonomously even in tissue culture conditions. Moreover, the circadian clock in the primary mPER2(Lu)(c) fibroblasts could be reprogrammed by the expression of the reprogramming factors. These findings suggest that mammalian circadian clock development may interact with cellular differentiation mechanisms. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  13. Effect of titanium dioxide nanoparticles on zebrafish embryos and developing retina

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    Ya-Jie Wang

    2014-12-01

    Full Text Available AIM:To investigate the impact of titanium dioxide nanoparticles (TiO2 NPs on embryonic development and retinal neurogenesis. METHODS:The agglomeration and sedimentation of TiO2 NPs solutions at different dilutions were observed, and the ultraviolet-visible spectra of their supernatants were measured. Zebrafish embryos were experimentally exposed to TiO2 NPs until 72h postfertilization (hpf. The retinal neurogenesis and distribution of the microglia were analyzed by immunohistochemistry and whole mount in situ hybridization. RESULTS: The1 mg/L was determined to be an appropriate exposure dose. Embryos exposed to TiO2 NPs had a normal phenotype. The neurogenesis was initiated on time, and ganglion cells, cones and rods were well differentiated at 72 hpf. The expression of fms mRNA and the 4C4 antibody, which were specific to microglia in the central nervous system (CNS, closely resembled their endogenous profile. CONCLUSION:These data demonstrate that short-term exposure to TiO2 NPs at a low dose does not lead to delayed embryonic development or retinal neurotoxicity.

  14. Absorption of the Wolffian duct and duplicated ureter by the urogenital sinus: morphological study using human fetuses and embryos.

    Science.gov (United States)

    Naito, Michiko; Hinata, Nobuyuki; Rodriguez-Vazquez, Jose Francisco; Murakami, Gen; Aizawa, Shin; Fujisawa, Masato

    2015-07-01

    To describe the embryological origin of the duplicated ureter and to investigate whether the urogenital sinus absorbs not only the Wolffian duct (WD) but also the ureter. During studies using sections of human fetuses (45 specimens), we incidentally found a specific type of ureteric duplication (at ~7 weeks) in which two unilateral ureters joined at the vesico-ureteric junction, apparently representing a morphology arising at an intermediate stage between complete and partial ureteric duplication. The existing literature lacks any photographic representation of early development of the vesico-ureteric junction, and we therefore studied horizontal sections of 10 human embryos (at ~5-6 weeks' gestation) in which the ureter did not join the urogenital sinus (future bladder) but instead joined the WD (future vas deferens). The sinus consistently showed a reversed Y-shape, the arms of which extended posteriorly to receive the WD. When absorption of the duct into the sinus wall reached the distal end of the ureter, the arm-like parts appeared to enlarge posteriorly for further involvement of the duct, with little or no incorporation of the ureter; therefore, the future trigone of the bladder might develop from these arm-like parts of the sinus posterior wall. Consequently, in the case of ureteric duplication included in the present study, it is considered that the ureters would probably have merged with the WD at closely adjacent sites. The present study represents the first photographic illustration of the early development of the human vesico-ureteric junction. © 2014 The Authors BJU International © 2014 BJU International Published by John Wiley & Sons Ltd.

  15. Characterization of microRNA in bovine in vitro culture media associated with embryo quality and development.

    Science.gov (United States)

    Kropp, Jenna; Khatib, Hasan

    2015-09-01

    Dairy cattle fertility has declined over time due to factors including reduced fertilization and early embryonic loss. To counter fertility problems and better study preimplantation embryonic development, in vitro production systems have been developed. These systems largely assess embryos based on their morphology, which is not a strong indicator of developmental potential. Currently, no biomarkers can be used to noninvasively survey an embryo's potential in terms of its development and ability to establish a pregnancy. Thus, the objective of this study was to characterize and identify microRNA (miRNA) in culture media of embryos of differing developmental competence for future development as noninvasive biomarkers of embryo quality. The MiRNA sequencing of media conditioned by blastocyst and degenerate (those that failed to develop from the morula to blastocyst stage) embryos, revealed 11 differentially expressed miRNA; all were higher in concentration in degenerate conditioned media. Differential expression of mature microRNA (miR)-24, miR-191, and miR-148a was further validated using quantitative real-time PCR. Functional analysis of miR-24 revealed that addition of a mimic miRNA to culture media of morulae embryos resulted in a 27.3% decrease in development to the blastocyst stage. Furthermore, expression of miR-24 was 44.29-fold higher in blastocysts cultured with a miR-24 mimic compared with control blastocysts. Interestingly, the expression of CDKN1b, a target gene of miR-24 was repressed in embryos grown in the presence of the miRNA mimic. Mimic supplementation experiments suggest that miRNA are taken up by the embryo and that extracellular miRNA affect embryonic development. Overall, identification of a rich extracellular milieu in conditioned media sets the framework for future studies to determine the long-term predictive ability of embryo-based miRNA biomarkers on pregnancy outcome. Copyright © 2015 American Dairy Science Association. Published by

  16. Efficacy of mechanical micro-vibration in the development of bovine embryos during in vitro maturation and culture.

    Science.gov (United States)

    Takahashi, Masahiro; Honda, Tatsutoshi; Hatoya, Shingo; Inaba, Toshio; Kawate, Noritoshi; Tamada, Hiromichi

    2018-02-06

    It is currently unclear how mechanical micro-vibration affects the in vitro culture of embryos in Japanese Black cow. In the experimental groups, immature oocytes and fertilized embryos were cultured using the micro-vibration culture system with the vibration set for 5 sec at intervals of 60 min and frequency of 20, 40, or 80 Hz, respectively, during in vitro maturation and in vitro development. Compared with the control group, the rate of blastocyst development significantly increased in the 40 Hz group. In addition, the number of blastocyst cells reduced significantly in the 80 Hz group. In conclusion, the development of blastocysts in cows is facilitated by providing moderate mechanical micro-vibration to immature oocytes and embryos during the in vitro maturation and in vitro development.

  17. Embryo development and corresponding factors affecting in vitro germination of Cymbidium faberi × C. sinense hybrid seeds

    Directory of Open Access Journals (Sweden)

    Li Fengtong

    2016-01-01

    Full Text Available A better understanding of embryo development would provide insights into seed quality and subsequent germination events in the interspecific hybridization of Cymbidium faberi ‘Jiepeimei’ × C. sinense ‘Qijianheimo’. At the mature stage, 26.1% of the ovules were abnormal. Most of the hybrid embryos could develop normally. Abortions mainly occurred at the zygote (9.5% and 2-4-celled embryo (15.1% stages. No germination was observed at 90 and 105 days after pollination (DAP, when the embryo was at the early globular stage, with abundant organelles but no storage materials. During 110-130 DAP, the globular embryo was formed and the starch grains began to accumulate in plastids. The hybrid seeds collected at 120 DAP showed initiation of germination. Germination significantly increased at 135 DAP and was maximal at 150 DAP, during which period the hybrid embryos developed into the late globular stage. The storage materials, i.e. lipid and protein bodies, began to accumulate and the filamentary structures derived from suspensor cells still persisted. After the seeds matured (160 DAP, the germination percentage declined sharply. Safranin staining revealed that the outer seed coat was totally cuticularized and the inner seed coat appeared as a cuticle layer enclosing the embryo proper tightly, which may be the main factor inhibiting the subsequent germination of hybrid seeds. In conclusion, 150 DAP should be the opportune time for the in vitro germination of C. faberi ‘Jiepeimei’ × C. sinense ‘Qijianheimo’ hybrid seeds.

  18. In Vitro Development of Ovine Embryos Following Maturation Under Limited CO2

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    ENDANG TRI MARGAWATI

    2005-09-01

    Full Text Available An experiment was conducted to examine the influence of CO2 during in vitro oocyte maturation on the in vitro ovine embryo development. Three treatments of CO2 were subjected to the oocyte development. Those were 2h gasses prior to maturation in incubator (T1; without CO2 either prior to or over maturation (T2 and CO2 exposure both prior to and over 22h maturation (T3. A total of 324 oocytes were used. Putative zygotes were cultured for seven days and evaluated for their developmental stage. Presence of CO2 (T3 increased the proportion of oocytes reaching Metaphase II ( 66.50 + 3.5%; p0.05. This study suggests that it is possible to mature ovine oocytes in the absence of CO2 without loss its potensial development. It may therefore be an effective method of maturing ovine oocytes during transportation to IVP (in vitro production laboratory.

  19. In Vitro Development of Ovine Embryos Following Maturation Under Limited CO2

    Directory of Open Access Journals (Sweden)

    ENDANG TRI MARGAWATI

    2005-09-01

    Full Text Available An experiment was conducted to examine the influence of CO2 during in vitro oocyte maturation on the in vitro ovine embryo development. Three treatments of CO2 were subjected to the oocyte development. Those were 2h gasses prior to maturation in incubator (T1; without CO2 either prior to or over maturation (T2 and CO2 exposure both prior to and over 22h maturation (T3. A total of 324 oocytes were used. Putative zygotes were cultured for seven days and evaluated for their developmental stage. Presence of CO2 (T3 increased the proportion of oocytes reaching Metaphase II (66.50 ± 3.5%; p0.05. This study suggests that it is possible to mature ovine oocytes in the absence of CO2 without loss its potensial development. It may therefore be an effective method of maturing ovine oocytes during transportation to IVP (in vitro production laboratory.

  20. Use of novel silver nanoparticles with Hyaluronan as potential biological labels for determining the quality of embryos development

    Science.gov (United States)

    Syrvatka, Vasyl J.; Slyvchuk, Yurij I.; Rozgoni, Ivan I.; Hevkan, Ivan I.; Bilyi, Oleksandr I.; Osypchuk, Oleksandr S.; Zyuzyun, Aza B.

    2013-09-01

    In reproductive medicine it is important to determine the quality of embryo development without interference in their function and viability. The surface plasmon resonance of silver nanoparticles makes them promising candidates for optical sensing, molecular labeling and imaging applications. Furthermore unique optical properties of silver nanoparticles provide an opportunity to use them as real time analytic tools in living state especially for observation of dynamic processes in gametes and embryos. The main aim of the study was to investigate the physicochemical properties and biological activities of novel silver nanoparticles with prospect of their use for the determining the quality of embryo development. For this purpose, we investigated the optical properties of new silver nanoparticles in biological mediums during cultivation and their influence on rabbit's embryos development in vitro. The physicochemical and biological properties of novel silver nanoparticles were compared with silver nanoparticles identical in size and shapes but with different chemical surfaces modifications by polyvinylpyrrolidone and bovine serum albumin. The results suggest that silver nanoparticles with hyaluronic acid were disintegrated with the formation of new complexes with proteins in biological mediums. This property with strong optical surface plasmon resonance of novel silver nanoparticles with hyaluronan makes them promising candidates in diagnostic area and gives reasons to explore them as biomarkers of target molecules. Nevertheless novel silver nanoparticles with hyaluronan at the concentrations of 0.1-1 μg/ml have no toxic effect on rabbit's embryos development and can be successfully applied in reproductive medicine.

  1. No cytotoxic effects from application of pentoxifylline to spermatozoa on subsequent pre-implantation embryo development in mice

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    Mohammad Ali Khalili

    2017-06-01

    Full Text Available The aim was to assess the effect of spermatozoa exposed to PTX on the rates of fertilization and embryo development and apoptotic cells within blastocysts in an animal model. Mice Oocytes were inseminated with spermatozoa exposed to 3.6 mmol PTX for 30 min, or with neat spermatozoa. Then fertilization and embryo development rate, blastocyst formation and quality, as well as total cell number of blastocyst, and DNA fragmentation index (DFI in blastocysts were surveyed in both groups. Fertilization and embryo development rate were similar between the groups. The rates of blastocyst formation did not differ significantly between control and PTX groups (52.4% vs. 51.8%. The average of total cell count in blastocysts and DFI in control and PTX groups were also insignificant (31.08 ± 1.5 vs. 34.14 ± 1.5 and 9.76 ± 5.0 vs. 11.77 ± 5.4. Application of PTX for enhancing sperm motility does not cause a cytotoxic effect on subsequent embryo development and embryo genome integrity.

  2. Development of half-embryo test and germination test for detection of irradiated fruits and grains

    Energy Technology Data Exchange (ETDEWEB)

    Kawamura, Y.; Murayama, M.; Uchiyamam, S.; Saito, Y. [National Inst. of Health Sciences, Tokyo (Japan)

    1996-12-31

    Irradiation treatment of fruit and grains is allowed in many countries for purposes such as insect disinfestation and the extension of shelf-life; the irradiation doses used being mainly below 1 kGy. The detection of such low doses is difficult for most techniques. On the other hand, biological systems are sensitive to low doses of {gamma}-irradiation. Therefore, a detection method based on biological changes would be applicable. It is well known that germination, or sprouting, is inhibited by {gamma}-irradiation and it has been observed that the seed germination of grapefruit is affected by irradiation and shoot growth is totally inhibited by a 0.3 kGy dose. However, the immediate use of this inhibition for the detection of irradiated grapefruit was prevented by variability caused by such factors as variety, harvest date and storage conditions. The time required to conduct the detection was also too lengthy. Accordingly, the germination test was improved and the half-embryo test was established for the detection of irradiated grapefruit and was applied to other citrus fruits, e.g. apples and cherries. Moreover, the test was also applied as a screening method for irradiated rice and wheat. In this report, the development of a standardized half-embryo test and germination test for the detection of irradiated fruit and grains is described. (author).

  3. Finding biomarkers in non-model species: literature mining of transcription factors involved in bovine embryo development.

    Science.gov (United States)

    Turenne, Nicolas; Tiys, Evgeniy; Ivanisenko, Vladimir; Yudin, Nikolay; Ignatieva, Elena; Valour, Damien; Degrelle, Séverine A; Hue, Isabelle

    2012-08-29

    arrangement of data from model organisms. This approach was applied to identify novel transcription factors during bovine blastocyst elongation, a process that is not observed in rodents and primates. As a result, searching through human and mouse corpuses, we identified numerous bovine homologs, among which 11 to 14% of transcription factors including the gold standard TF as well as novel TF potentially important to gene regulation in ruminant embryo development. The scripts of the workflow are written in Perl and available on demand. They require data input coming from all various databases for any kind of biological issue once the data has been prepared according to keywords for the studied topic and species; we can provide data sample to illustrate the use and functionality of the workflow. To do so, we created a workflow that allowed the pipeline processing of literature data and biological data, extracted from Web of Science (WoS) or PubMed but also from Gene Expression Omnibus (GEO), Gene Ontology (GO), Uniprot, HomoloGene, TcoF-DB and TFe (TF encyclopedia). First, the human and mouse homologs of the bovine proteins were selected, filtered by text corpora and arranged by score functions. The score functions were based on the gene name frequencies in corpora. Then, transcription factors were identified using TcoF-DB and double-checked using TFe to characterise TF groups and families. Thus, among a search space of 18,670 bovine homologs, 489 were identified as transcription factors. Among them, 243 were absent from the high-throughput data available at the time of the study. They thus stand so far for putative TF acting during bovine embryo elongation, but might be retrieved from a recent RNA sequencing dataset (Mamo et al. , 2012). Beyond the 246 TF that appeared expressed in bovine elongating tissues, we restricted our interpretation to those occurring within a list of 50 top-ranked genes. Among the transcription factors identified therein, half belonged to the gold

  4. Finding biomarkers in non-model species: literature mining of transcription factors involved in bovine embryo development

    Directory of Open Access Journals (Sweden)

    Turenne Nicolas

    2012-08-01

    approach based on literature-mining and score arrangement of data from model organisms. This approach was applied to identify novel transcription factors during bovine blastocyst elongation, a process that is not observed in rodents and primates. As a result, searching through human and mouse corpuses, we identified numerous bovine homologs, among which 11 to 14% of transcription factors including the gold standard TF as well as novel TF potentially important to gene regulation in ruminant embryo development. The scripts of the workflow are written in Perl and available on demand. They require data input coming from all various databases for any kind of biological issue once the data has been prepared according to keywords for the studied topic and species; we can provide data sample to illustrate the use and functionality of the workflow. Results To do so, we created a workflow that allowed the pipeline processing of literature data and biological data, extracted from Web of Science (WoS or PubMed but also from Gene Expression Omnibus (GEO, Gene Ontology (GO, Uniprot, HomoloGene, TcoF-DB and TFe (TF encyclopedia. First, the human and mouse homologs of the bovine proteins were selected, filtered by text corpora and arranged by score functions. The score functions were based on the gene name frequencies in corpora. Then, transcription factors were identified using TcoF-DB and double-checked using TFe to characterise TF groups and families. Thus, among a search space of 18,670 bovine homologs, 489 were identified as transcription factors. Among them, 243 were absent from the high-throughput data available at the time of the study. They thus stand so far for putative TF acting during bovine embryo elongation, but might be retrieved from a recent RNA sequencing dataset (Mamo et al. , 2012. Beyond the 246 TF that appeared expressed in bovine elongating tissues, we restricted our interpretation to those occurring within a list of 50 top-ranked genes. Among the transcription

  5. Interactions of acid-base balance and hematocrit regulation during environmental respiratory gas challenges in developing chicken embryos (Gallus gallus).

    Science.gov (United States)

    Burggren, Warren W; Andrewartha, Sarah J; Tazawa, Hiroshi

    2012-08-15

    How the determinants of hematocrit (Hct) - alterations in mean corpuscular volume (MCV) and/or red blood cell concentration ([RBC]) - are influenced by acid-base balance adjustments across development in the chicken embryo is poorly understood. We hypothesized, based on oxygen transport needs of the embryos, that Hct will increase during 1 day of hypercapnic hypoxia (5%CO(2), 15%O(2)) or hypoxia alone (0%CO(2), 15%O(2)), but decrease in response to hyperoxia (0%CO(2), 40%O(2)). Further, age-related differences in acid-base disturbances and Hct regulation may arise, because the O(2) transport and hematological regulatory systems are still developing in embryonic chickens. Our studies showed that during 1 day of hypoxia (with or without hypercapnia) Hct increased through both increased MCV and [RBC] in day 15 (d15) embryo, but only through increased MCV in d17 embryo and therefore enhancement of O(2) transport was age-dependent. Hypercapnia alone caused a ≈ 14% decrease in Hct through decreased [RBC] and therefore did not compensate for decreased blood oxygen affinity resulting from the Bohr shift. The 11% (d15) and 14% (d17) decrease in Hct during hyperoxia in advanced embryos was because of an 8% and 9% decrease, respectively, in [RBC], coupled with an associated 3% and 5% decrease in MCV. Younger, d13 embryos were able to metabolically compensate for respiratory acidosis induced by hypercapnic hypoxia, and so were more tolerant of disturbances in acid-base status induced via alterations in environmental respiratory gas composition than their more advanced counterparts. This counter-intuitive increased tolerance likely results from the relatively low [Formula: see text] and immature physiological functions of younger embryos. Copyright © 2012 Elsevier B.V. All rights reserved.

  6. Ultrastructural study of polyspermy during early embryo development in pigs, observed by scanning electron microscope and transmission electron microscope.

    Science.gov (United States)

    Xia, P; Wang, Z; Yang, Z; Tan, J; Qin, P

    2001-02-01

    Polyspermy is generally considered a pathological phenomenon in mammals. Incidence of polyspermy in porcine eggs in vivo is extremely high (30-40%) compared with other species, and polyspermy rate in the in vitro fertilized eggs in pigs can reach 65%. It is still unknown whether polyspermy to a certain degree is a physiological condition in pigs, and whether porcine eggs have any capability with which to remove the accessory sperm in the cytoplasm. The objectives in the present study are to observe the ultrastructural changes of accessory sperm during early embryonic development in pigs. A total of 58 normal, early embryos at one-, two, three-, and four-cell and morular stages were collected from gilts and were studied by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The surface ultrastructure showed that sperm fusion with the zona pellucida was a continuous process during one-, two-, three-, and four-cell and morular stages, as observed by the SEM. Accessory sperm were present in the cytoplasm of cleaved embryos. The sperm heads in the cytoplasm of cleaved embryos did not decondense. TEM revealed the presence of a condensed sperm head within a lysosome (or phagolysosome) in a three-cell embryo. These observations suggest that polyspermy may be a physiological condition in pigs and that early embryos may develop to term if accessory sperm do not interrupt the embryo genome. Furthermore, lysosome activity could be another physiological mechanism for removing accessory sperm in the cytoplasm of fertilized eggs and cleaved embryos after fertilization in pigs.

  7. Transcription in pronuclei and one- to four-cell embryos drives early development in a nematode

    Science.gov (United States)

    Wang, Jianbin; Garrey, Julianne; Davis, Richard E.

    2014-01-01

    Summary Background A long-standing view of development is that transcription is silenced in the oocyte until early divisions in the embryo. The point at which major transcription is reactivated varies between organisms, but is usually after the 2-cell stage. However, this model may not be universal. Results We used RNA-seq and exploited the protracted development of the parasitic nematode Ascaris suum, to provide a comprehensive time course of mRNA expression, degradation, and translation during early development. Surprisingly, we find that ~4,000 genes are transcribed prior to pronuclear fusion and in the 1–4 cell embryos. Intriguingly, we do not detect maternal contribution of many orthologs of maternal C. elegans mRNAs, but instead find these are newly transcribed in the A. suum zygote prior to pronuclear fusion. Ribosome profiling demonstrates that, in general, early embryonic mRNAs are not stored for subsequent translation, but are directly translated following their synthesis. The role of maternally contributed and zygotically transcribed genes differs between the nematodes A. suum and C. elegans despite the fact that the two nematodes appear to exhibit highly similar morphological patterns during early development. Conclusions Our study indicates that major transcription can occur immediately after fertilization and prior to pronuclear fusion in metazoa, suggesting that newly transcribed genes appear to drive A. suum early development. Furthermore, the mechanisms used for controlling the timing of the expression of key conserved genes has been altered between the two nematodes, illustrating significant plasticity in the regulatory networks that play important roles in developmental outcomes in nematodes. PMID:24374308

  8. Expression of intracellular interferon-alpha confers antiviral properties in transfected bovine fetal fibroblasts and does not affect the full development of SCNT embryos.

    Directory of Open Access Journals (Sweden)

    Dawei Yu

    Full Text Available Foot-and-mouth disease, one of the most significant diseases of dairy herds, has substantial effects on farm economics, and currently, disease control measures are limited. In this study, we constructed a vector with a human interferon-α (hIFN-α (without secretory signal sequence gene cassette containing the immediate early promoter of human cytomegalovirus. Stably transfected bovine fetal fibroblasts were obtained by G418 selection, and hIFN-α transgenic embryos were produced by somatic cell nuclear transfer (SCNT. Forty-six transgenic embryos were transplanted into surrogate cows, and five cows (10.9% became pregnant. Two male cloned calves were born. Expression of hIFN-α was detected in transfected bovine fetal fibroblasts, transgenic SCNT embryos, and different tissues from a transgenic SCNT calf at two days old. In transfected bovine fetal fibroblasts, expression of intracellular IFN-α induced resistance to vesicular stomatitis virus infection, increased apoptosis, and induced the expression of double-stranded RNA-activated protein kinase gene (PKR and the 2'-5'-oligoadenylate synthetase gene (2'-5' OAS, which are IFN-inducible genes with antiviral activity. Analysis by qRT-PCR showed that the mRNA expression levels of PKR, 2'-5' OAS, and P53 were significantly increased in wild-type bovine fetal fibroblasts stimulated with extracellular recombinant human IFN-α-2b, showing that intracellular IFN-α induces biological functions similar to extracellular IFN-α. In conclusion, expression of intracellular hIFN-α conferred antiviral properties in transfected bovine fetal fibroblasts and did not significantly affect the full development of SCNT embryos. Thus, IFN-α transgenic technology may provide a revolutionary way to achieve elite breeding of livestock.

  9. Effect of copper nanoparticles on metabolic rate and development of chicken embryos

    DEFF Research Database (Denmark)

    Pineda, Lane Manalili; Sawosz, E.; Vadalasetty, K. P.

    2013-01-01

    The objective of the study was to investigate the effects of an in ovo injection of CuNano and the timing of injection on metabolic rate (O consumption and heat production, HP) and development of layer hatchlings. On day 1 of incubation, 192 fertile eggs from 29-week-old Lohmann breeder strain...... chickens were distributed into four groups that were administered colloidal CuNano on: day 1 and/or 10. Gaseous exchange was measured in an open-air-circuit respiration unit, and HP was calculated for 16- and 19-day-old embryos. Yolk free body weight (YFBW) at 24h after hatching and the relative organ...... weights were used as a measure of hatchling development. In ovo injection of CuNano on different days during incubation significantly decreased O consumption and HP compared with the control group. The residual yolk sac weight in the treated groups was significantly higher than in the control group (P0...

  10. A potential determinant role of adiponectin and receptors for the early embryo development in PCOS patients with obesity hinted by quantitative profiling.

    Science.gov (United States)

    Zhang, Ning; Hao, Cuifang; Liu, Xiaoyan; Zhang, Shouxin; Zhang, Fengrong; Zhuang, Lili; Zhao, Dongmei

    2017-02-01

    To identify the quantitative profiling of adiponectin and its receptors (AdipoR1, AdipoR2, and T-cadherin) in cumulus cells (CCs) and to evaluate their roles in the early embryo development of polycystic ovary syndrome (PCOS) patients, in part, with obesity. Fifty-five subjects were divided into two groups according to the body mass index. Oocytes were further inseminated and only mature and normal fertilized oocytes (2PN) were included in this research. Real-time PCR and western blot were performed to identify adiponectin and its receptors in CCs. Adiponectin and receptors were ubiquitously expressed in CCs of PCOS and non-PCOS patients. The level of AdipoR2 in CCs from the oocytes yielding blastocyst after 5/6 days in vitro culture was markedly higher than in those from oocytes could not develop to blastocyst stage after Day 6, for non-obese or obese PCOS patients (0.1647 ± 0.0161 versus 0.0783 ± 0.0385, 0.1948 ± 0.0307 versus 0.1057 ± 0.0236, respectively, p < 0.05). In addition, only in patients with PCOS and concurrent obesity the AdipoR1 in CCs was considerably increased in CC-B+ compared with CC-B- subgroup (0.5162 ± 0.0371 versus 0.2448 ± 0.0333, p < 0.01). The development of early embryo was associated with the up-regulation of AdipoR1 and AdipoR2 in PCOS patients. Our results suggested that adiponectin could positively modulate embryo development in humans. Further investigations should be carried out to unlock the crucial role that adiponectin plays in embryo development.

  11. Effect of CO2 on somatic embryos development Coffea arabica L. cv. ‘Caturra rojo’ and Clematis tangutica K.

    Directory of Open Access Journals (Sweden)

    Raúl Barbon

    2016-07-01

    Full Text Available Studies to optimize somatic embryogenesis have traditionally focused on the components of the culture medium but little other in vitro environment factors have been analyzed such as the composition of the gaseous atmosphere. The objective of this work was to determine the influence of CO2 on the development of the somatic embryo during the transition from the globular to the torpedo stage. The research was carried out on two model species for somatic embryogenesis that they are developed in different climatic zones: Coffea arabica L. cv. ‘Caturra rojo’ and Clematis tangutica K. Three CO2 concentrations (2.5, 5.0 and 10.0% combined with 21% O2 and two controls (passive exchange and forced ventilation were used. The effect of CO2 on the differentiation of somatic embryos from globular to torpedo stage in coffee and clematis was demonstrated, since in the treatments with passive exchange, where there was accumulation of CO2, the differentiation of somatic embryos was superior to treatments with forced ventilation. With 5.0% CO2 the process of differentiation of the embryos in the globular stage was stimulated, because in the treatment with this concentration of CO2 for coffee and clematis the highest proportion of embryos in torpedo stages and low levels of malformation were obtained.   Keywords: carbon dioxide, differentiation, in vitro environment, somatic embryogenesis

  12. Second heart field and the development of the outflow tract in human embryonic heart.

    Science.gov (United States)

    Yang, Yan-Ping; Li, Hai-Rong; Cao, Xi-Mei; Wang, Qin-Xue; Qiao, Cong-Jin; Ya, Jing

    2013-04-01

    The second heart field (SHF) is indicated to contribute to the embryonic heart development. However, less knowledge is available about SHF development of human embryo due to the difficulty of collecting embryos. In this study, serial sections of human embryos from Carnegie stage 10 (CS10) to CS16 were stained with antibodies against Islet-1 (Isl-1), Nkx2.5, GATA4, myosin heavy chain (MHC) and α-smooth muscle actin (α-SMA) to observe spatiotemporal distribution of SHF and its contribution to the development of the arterial pole of cardiac tube. Our findings suggest that during CS10 to CS12, SHF of the human embryo is composed of the bilateral pharyngeal mesenchyme, the central mesenchyme of the branchial arch and splanchnic mesoderm of the pericardial cavity dorsal wall. With development, SHF translocates and consists of ventral pharyngeal mesenchyme and dorsal wall of the pericardial cavity. Hence, the SHF of human embryo shows a dynamic spatiotemporal distribution pattern. The formation of the Isl-1 positive condense cell prongs provides an explanation for the saddle structure formation at the distal pole of the outflow tract. In human embryo, the Isl-1 positive cells of SHF may contribute to the formation of myocardial outflow tract (OFT) and the septum during different development stages. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

  13. Embryos of the viviparous dermapteran, Arixenia esau develop sequentially in two compartments: terminal ovarian follicles and the uterus.

    Science.gov (United States)

    Tworzydlo, Waclaw; Kisiel, Elzbieta; Bilinski, Szczepan M

    2013-01-01

    Three main reproductive strategies have been described among insects: most common oviparity, ovoviviparity and viviparity. In the latter strategy, the embryonic development takes place within the body of the mother which provides gas exchange and nutrients for embryos. Here we present the results of histological and EM analyses of the female reproductive system of the viviparous earwig, Arixenia esau, focusing on all the modifications related to the viviparity. We show that in the studied species the embryonic development consists of two "physiological phases" that take place in two clearly disparate compartments, i.e. the terminal ovarian follicle and the uterus. In both compartments the embryos are associated with synthetically active epithelial cells. We suggest that these cells are involved in the nourishment of the embryo. Our results indicate that viviparity in arixeniids is more complex than previously considered. We propose the new term "pseudoplacento-uterotrophic viviparity" for this unique two-phase reproductive strategy.

  14. Embryos of the viviparous dermapteran, Arixenia esau develop sequentially in two compartments: terminal ovarian follicles and the uterus.

    Directory of Open Access Journals (Sweden)

    Waclaw Tworzydlo

    Full Text Available Three main reproductive strategies have been described among insects: most common oviparity, ovoviviparity and viviparity. In the latter strategy, the embryonic development takes place within the body of the mother which provides gas exchange and nutrients for embryos. Here we present the results of histological and EM analyses of the female reproductive system of the viviparous earwig, Arixenia esau, focusing on all the modifications related to the viviparity. We show that in the studied species the embryonic development consists of two "physiological phases" that take place in two clearly disparate compartments, i.e. the terminal ovarian follicle and the uterus. In both compartments the embryos are associated with synthetically active epithelial cells. We suggest that these cells are involved in the nourishment of the embryo. Our results indicate that viviparity in arixeniids is more complex than previously considered. We propose the new term "pseudoplacento-uterotrophic viviparity" for this unique two-phase reproductive strategy.

  15. Effects of copper oxide nanoparticles on developing zebrafish embryos and larvae.

    Science.gov (United States)

    Sun, Yan; Zhang, Gong; He, Zizi; Wang, Yajie; Cui, Jianlin; Li, Yuhao

    2016-01-01

    Copper oxide nanoparticles (CuO NPs) are used for a variety of purposes in a wide range of commercially available products. Some CuO NPs probably end up in the aquatic systems, thus raising concerns about aqueous exposure toxicity, and the impact of CuO NPs on liver development and neuronal differentiation remains unclear. In this study, particles were characterized using Fourier transform infrared spectra, scanning electron microscopy, and transmission electron microscopy. Zebrafish embryos were continuously exposed to CuO NPs from 4 hours postfertilization at concentrations of 50, 25, 12.5, 6.25, or 1 mg/L. The expression of gstp1 and cyp1a was examined by quantitative reverse transcription polymerase chain reaction. The expression of tumor necrosis factor alpha and superoxide dismutase 1 was examined by quantitative reverse transcription polymerase chain reaction and Western blotting. Liver development and retinal neurodifferentiation were analyzed by whole-mount in situ hybridization, hematoxylin-eosin staining, and immunohistochemistry, and a behavioral test was performed to track the movement of larvae. We show that exposure of CuO NPs at low doses has little effect on embryonic development. However, exposure to CuO NPs at concentrations of 12.5 mg/L or higher leads to abnormal phenotypes and induces an inflammatory response in a dose-dependent pattern. Moreover, exposure to CuO NPs at high doses results in an underdeveloped liver and a delay in retinal neurodifferentiation accompanied by reduced locomotor ability. Our data demonstrate that short-term exposure to CuO NPs at high doses shows hepatotoxicity and neurotoxicity in zebrafish embryos and larvae.

  16. Embryo sac development in yellow passion fruit Passiflora edulis f. flavicarpa (Passifloraceae

    Directory of Open Access Journals (Sweden)

    Margarete Magalhães de Souza

    2002-01-01

    Full Text Available The yellow passion fruit, Passiflora edulis f. flavicarpa, is one of the most important Brazilian fruit crops. It is an allogamous, diploid, and self-incompatible species. It has hermaphrodite, solitary flowers, located in the leaf axils and protected by leaf bracts. The flower has an androgynophore, which is a straight stalk supporting its reproductive parts. There are usually five anthers, located at the tip of each of the five filaments. The ovary is borne just above the filaments, at the top of the androgynophore; there are three styles that are united at their base, and at the top there are three stigmas. The objective of this research was to observe embryo sac development in yellow passion flowers. Ovaries at different stages of development were fixed in FAA (formalin, acetic acid and alcohol solution, hydrated, stained with Mayer’s hemalum, and dehydrated. Ovules were cleared by using methyl salicylate, mounted on slides, and observed through a confocal scanning laser microscope. The yellow passion fruit ovule is bitegmic, crassinucellate, and anatropous, and its gametophyte development is of the Polygonum type. After meiosis, functional megaspores under go three successive mitotic divisions, resulting in an eight-nucleate megagametophyte: the egg apparatus at the micropylar end, two polar nuclei at the cell center, and three antipodals at the chalazal end. The egg apparatus is formed by an egg cell and two synergids, each with a filiform apparatus. The mature embryo sac has an egg cell, two synergids, two polar nuclei, and three antipodes, as has been described for most angiosperms.

  17. High environmental temperature increases glucose requirement in the developing chicken embryo.

    Directory of Open Access Journals (Sweden)

    Roos Molenaar

    Full Text Available Environmental conditions during the perinatal period influence metabolic and developmental processes in mammals and avian species, which could impact pre- and postnatal survival and development. The current study investigated the effect of eggshell temperature (EST on glucose metabolism in broiler chicken embryos. Broiler eggs were incubated at a high (38.9°C or normal (37.8°C EST from day 10.5 of incubation onward and were injected with a bolus of [U-(13C]glucose in the chorio-allantoic fluid at day 17.5 of incubation. After [U-(13C]glucose administration, (13C enrichment was determined in intermediate pools and end-products of glucose metabolism. Oxidation of labeled glucose occurred for approximately 3 days after injection. Glucose oxidation was higher in the high than in the normal EST treatment from day 17.6 until 17.8 of incubation. The overall recovery of (13CO2 tended to be 4.7% higher in the high than in the normal EST treatment. An increase in EST (38.9°C vs 37.8°C increased (13C enrichment in plasma lactate at day 17.8 of incubation and (13C in hepatic glycogen at day 18.8 of incubation. Furthermore, high compared to normal EST resulted in a lower yolk-free body mass at day 20.9 (-2.74 g and 21.7 (-3.81 g of incubation, a lower hepatic glycogen concentration at day 18.2 (-4.37 mg/g and 18.8 (-4.59 mg/g of incubation, and a higher plasma uric acid concentration (+2.8 mg/mL/+43% at day 21.6 of incubation. These results indicate that the glucose oxidation pattern is relatively slow, but the intensity increased consistently with an increase in developmental stage of the embryo. High environmental temperatures in the perinatal period of chicken embryos increased glucose oxidation and decreased hepatic glycogen prior to the hatching process. This may limit glucose availability for successful hatching and could impact body development, probably by increased gluconeogenesis from glucogenic amino acids to allow anaerobic glycolysis.

  18. Single nucleotide polymorphism microarray-based concurrent screening of 24-chromosome aneuploidy and unbalanced translocations in preimplantation human embryos.

    Science.gov (United States)

    Treff, Nathan R; Northrop, Lesley E; Kasabwala, Khushabu; Su, Jing; Levy, Brynn; Scott, Richard T

    2011-04-01

    To develop, validate, and apply a single nucleotide polymorphism (SNP) microarray-based method for simultaneous preimplantation genetic diagnosis (PGD) of unbalanced inheritance of rearranged chromosomes and 24-chromosome aneuploidy screening. Prospective clinical research study. Academic reproductive medicine center. Eighteen couples carrying a balanced reciprocal or Robertsonian chromosomal rearrangement. PGD on blastocyst trophectoderm biopsy specimens. Aneuploidy, implantation, pregnancy, and delivery rates after SNP microarray-based aneuploidy and translocation screening. Single nucleotide polymorphism microarray was capable of detecting translocation-associated imbalances as small as 9.0 megabases. In the 12 transfers performed, sustained implantation occurred for 9 (45%) of 20 balanced-normal and euploid embryos replaced. The clinical pregnancy rate in patients receiving a transfer was 75% with six singleton deliveries and three ongoing singleton pregnancies thus far. Significantly fewer embryos were eligible for transfer with the incorporation of simultaneous 24-chromosome aneuploidy screening. Arrested embryos were also significantly more likely to possess unbalanced chromosomes when compared with developmentally competent blastocysts. This SNP microarray-based method provides the first opportunity to improve outcomes through comprehensive identification of euploid embryos from translocation carrier couples. Copyright © 2011 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  19. Human pancreas development.

    Science.gov (United States)

    Jennings, Rachel E; Berry, Andrew A; Strutt, James P; Gerrard, David T; Hanley, Neil A

    2015-09-15

    A wealth of data and comprehensive reviews exist on pancreas development in mammals, primarily mice, and other vertebrates. By contrast, human pancreatic development has been less comprehensively reviewed. Here, we draw together those studies conducted directly in human embryonic and fetal tissue to provide an overview of what is known about human pancreatic development. We discuss the relevance of this work to manufacturing insulin-secreting β-cells from pluripotent stem cells and to different aspects of diabetes, especially permanent neonatal diabetes, and its underlying causes. © 2015. Published by The Company of Biologists Ltd.

  20. Supplementation of fetal bovine serum alters histone modification H3R26me2 during preimplantation development of in vitro produced bovine embryos

    Directory of Open Access Journals (Sweden)

    Daniel R. Arnold

    2015-07-01

    Full Text Available Abstract In vitro production (IVP of bovine embryos is not only of great economic importance to the cattle industry, but is also an important model for studying embryo development. The aim of this study was to evaluate the histone modification, H3R26me2 during pre-implantation development of IVP bovine embryos cultured with or without serum supplementation and how these in vitro treatments compared to in vivo embryos at the morula stage. After in vitro maturation and fertilization, bovine embryos were cultured with either 0 or 2.5% fetal bovine serum (FBS. Development was evaluated and embryos were collected and fixed at different stages during development (2-, 4-, 8-, 16-cell, morula and blastocyst. Fixed embryos were then used for immunofluorescence utilizing an antibody for H3R26me2. Images of stained embryos were analyzed as a percentage of total DNA. Embryos cultured with 2.5% FBS developed to blastocysts at a greater rate than 0%FBS groups (34.85±5.43% vs. 23.38±2.93%; P<0.05. Levels of H3R26me2 changed for both groups over development. In the 0%FBS group, the greatest amount of H3R26me2 staining was at the 4-cell (P<0.05, 16-cell (P<0.05 and morula (P<0.05 stages. In the 2.5%FBS group, only 4-cell stage embryos were significantly higher than all other stages (P<0.01. Morula stage in vivo embryos had similar levels as the 0%FBS group, and both were significantly higher than the 2.5%FBS group. These results suggest that the histone modification H3R26me2 is regulated during development of pre-implantation bovine embryos, and that culture conditions greatly alter this regulation.

  1. Early embryo development in a sequential versus single medium: a randomized study

    Directory of Open Access Journals (Sweden)

    D'Hooghe Thomas M

    2010-07-01

    Full Text Available Abstract Background The success of in vitro fertilization techniques is defined by multiple factors including embryo culture conditions, related to the composition of the culture medium. In view of the lack of solid scientific data and in view of the current general belief that sequential media are superior to single media, the aim of this randomized study was to compare the embryo quality in two types of culture media. Methods In this study, the embryo quality on day 3 was measured as primary outcome. In total, 147 patients younger than 36 years treated with IVF/ICSI during the first or second cycle were included in this study. Embryos were randomly cultured in a sequential (group A or a single medium (group B to compare the embryo quality on day 1, day 2 and day 3. The embryo quality was compared in both groups using a Chi-square test with a significance level of 0.05. Results At day 1, the percentage of embryos with a cytoplasmic halo was higher in group B (46% than in group A (32%. At day 2, number of blastomeres, degree of fragmentation and the percentage of unequally sized blastomeres were higher in group B than in group A. At day 3, a higher percentage of embryos had a higher number of blastomeres and unequally sized blastomeres in group B. The number of good quality embryos (GQE was comparable in both groups. The embryo utilization rate was higher in group B (56% compared to group A (49%. Conclusions Although, no significant difference in the number of GQE was found in both media, the utilization rate was significantly higher when the embryos were cultured in the single medium compared to the sequential medium. The results of this study have a possible positive effect on the cumulative cryo-augmented pregnancy rate. Trial registration number NCT01094314

  2. Beyond the 'embryo question': human embryonic stem cell ethics in the context of biomaterial donation in the UK.

    Science.gov (United States)

    Bahadur, G; Morrison, M; Machin, L

    2010-12-01

    Discussion about the ethics of human embryonic stem cell (ESC) research in the UK tends to be dominated by the divisive and potentially intractable issue of the moral status of the embryo. This can have the effect of silencing or marginalizing other concerns, especially in the context of public engagement with science in this field. One such area of potential public concern is the donation of oocytes and embryos to stem cell research. Contemporary research on the views of donors and potential donors about a wide range of biomaterials, from solid organs to gametes and bone marrow, is reviewed and used to illustrate the range and types of ethical concerns articulated by this important group of stakeholders. Attitudes to donation are found to vary according to the type of tissue being donated or collected, the purpose for which donation is being sought and the nature of the recipient of the donation. Pertinently, attitudes towards donating oocytes are found to differ in some respects from donation of embryos or fetal tissue. The implications of these findings for ensuring ethically robust informed consent and publicly acceptable sourcing of human biomaterials for stem cell research are then considered. Copyright © 2010 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  3. Inhibition of fumonisin B1 cytotoxicity by nanosilicate platelets during mouse embryo development.

    Directory of Open Access Journals (Sweden)

    Yu-Jing Liao

    Full Text Available Nanosilicate platelets (NSP, the form of natural silicate clay that was exfoliated from montmorillonite (MMT, is widely used as a feed additive for its high non-specific binding capacity with mycotoxins such as fumonisin B1 (FB1, and has been evaluated its safety for biomedical use including cytotoxicity, genotoxicity, and lethal dosage (LD. In the study, we further examined its toxicity on the development of CD1 mouse embryos and its capacity to prevent teratogenesis-induced by FB1. In vitro cultures, NSP did not disturb the development and the quality of intact pre-implantation mouse embryos. Further, newborn mice from females consumed with NSP showed no abnormalities. NSP had an unexpected high adsorption capacity in vitro. In contrast to female mice consumed with FB1 only, a very low residual level of FB1 in the circulation, reduced incidence of neutral tube defects and significantly increased fetal weight were observed in the females consumed with FB1 and NSP, suggesting a high alleviation effect of NSP on FB1 in vivo. Furthermore, FB1 treatment disturbed the gene expression of sphingolipid metabolism enzymes (longevity assurance homolog 5, LASS 5; sphingosine kinase 1, Sphk1; sphingosine kinase 2, Sphk2; sphingosine 1- phosphate lyase, Sgpl1; sphingosine 1-phosphate phosphatase, Sgpp1 in the maternal liver, uterus, fetus, and placenta, but NSP administration reversed the perturbations. Based on these findings, we conclude that NSP is a feasible and effective agent for supplementary use in reducing the toxicity of FB1 to animals.

  4. Marketing Human Resource Development.

    Science.gov (United States)

    Frank, Eric, Ed.

    1994-01-01

    Describes three human resource development activities: training, education, and development. Explains marketing from the practitioners's viewpoint in terms of customer orientation; external and internal marketing; and market analysis, research, strategy, and mix. Shows how to design, develop, and implement strategic marketing plans and identify…

  5. Embryo rescue as a method to develop and multiply a backcross ...

    African Journals Online (AJOL)

    Jane

    2010-10-18

    Oct 18, 2010 ... growing as a food security crop in sub Saharan Africa, where malnutrition is a menace. ... seeds in embryo culture was high (66%). ... Embryo culture provides a simple technique for breaking seed dormancy and ensuring a fairly uniform germination rate (Biggs et al., 1986). Raising cassava plantlets in a ...

  6. "Thin" property and controversial subject matter: Yanner v. Eaton and property rights in human tissue and embryos.

    Science.gov (United States)

    Moses, Lyria Bennett; Gollan, Nicola

    2013-12-01

    This article examines the definitions of "property" offered by the majority of the High Court of Australia in the case of Yanner v Eaton (1999) 201 CLR 351, which involved a statute giving the Crown "property" in fauna. It argues that the majority judges in that case endorsed a flexible or "thin" conception of property that is consistent with recognition of property in "things" such as excised human tissue and in vitro human embryos, despite the many differences between such "things" and ordinary chattels. A similar flexible conception of property was also an important factor in the United Kingdom case of Yearworth v North Bristol NHS Trust[2010] QB 1.

  7. Phase variance optical coherence microscopy for label-free imaging of the developing vasculature in zebrafish embryos

    Science.gov (United States)

    Chen, Yu; Trinh, Le A.; Fingler, Jeff; Fraser, Scott E.

    2016-12-01

    A phase variance optical coherence microscope (pvOCM) has been created to image blood flow in the microvasculature of zebrafish embryos, without the use of exogenous labels. The pvOCM imaging system has axial and lateral resolutions of 2.8 μm in tissue and imaging depth of more than 100 μm. Images of 2 to 5 days postfertilization zebrafish embryos identified the detailed anatomical structure based on OCM intensity contrast. Phase variance contrast offered visualization of blood flow in the arteries, veins, and capillaries. The pvOCM images of the vasculature were confirmed by direct comparisons with fluorescence microscopy images of transgenic embryos in which the vascular endothelium is labeled with green fluorescent protein. The ability of pvOCM to capture activities of regional blood flow permits it to reveal functional information that is of great utility for the study of vascular development.

  8. The role of RNA polymerase I transcription and embryonic genome activation in nucleolar development in bovine preimplantation embryos

    DEFF Research Database (Denmark)

    Østrup, Olga; Strejcek, F.; Petrovicova, I.

    2008-01-01

    was lacking and clustering of nucleolar proteins was hampered. In conclusion, rDNA transcription is not required for targeting of rRNA processing proteins, rRNA is maternally inherited and target to rDNA independent of transcription, and de novo transcription is required for proper nucleologenesis in cattle.......The aim of the present study was to investigate the role of RNA polymerase I (RPI) transcription in nucleolar development during major transcriptional activation (MTA) in cattle. Late eight-cell embryos were cultured in the absence (control group) or presence of actinomycin D (AD) (RPI inhibition......, Ad 0.2 µg/ml; total transcriptional inhibition, AD 2.0 µg/ml). Late four-cell embryos were cultured to late eight-cell stage in 0.2 µg/ml AD (MTA prevention, ADLT (long-term total transcriptional inhibition group). Embryos were processed for autoradiography, transmission electron microscopy...

  9. Human Rights, Human Needs, Human Development, Human Security

    OpenAIRE

    Gasper, D.R.

    2007-01-01

    Human rights, human development and human security form increasingly important, partly interconnected, partly competitive and misunderstood ethical and policy discourses. Each tries to humanize a pre-existing and unavoidable major discourse of everyday life, policy and politics; each has emerged within the United Nations world; each relies implicitly on a conceptualisation of human need; each has specific strengths. Yet mutual communication, understanding and co-operation are deficient, espec...

  10. Glycerol-3-phosphate acyltransferase 4 is essential for the normal development of reproductive organs and the embryo in Brassica napus.

    Science.gov (United States)

    Chen, Xue; Chen, Guanqun; Truksa, Martin; Snyder, Crystal L; Shah, Saleh; Weselake, Randall J

    2014-08-01

    The enzyme sn-glycerol-3-phosphate acyltransferase 4 (GPAT4) is involved in the biosynthesis of plant lipid poly-esters. The present study further characterizes the enzymatic activities of three endoplasmic reticulum-bound GPAT4 isoforms of Brassica napus and examines their roles in the development of reproductive organs and the embryo. All three BnGPAT4 isoforms exhibited sn-2 acyltransferase and phosphatase activities with dicarboxylic acid-CoA as acyl donor. When non-substituted acyl-CoA was used as acyl donor, the rate of acylation was considerably lower and phosphatase activity was not manifested. RNA interference (RNAi)-mediated down-regulation of all GPAT4 homologues in B. napus under the control of the napin promoter caused abnormal development of several reproductive organs and reduced seed set. Microscopic examination and reciprocal crosses revealed that both pollen grains and developing embryo sacs of the B. napus gpat4 lines were affected. The gpat4 mature embryos showed decreased cutin content and altered monomer composition. The defective embryo development further affected the oil body morphology, oil content, and fatty acid composition in gpat4 seeds. These results suggest that GPAT4 has a critical role in the development of reproductive organs and the seed of B. napus. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  11. Spatio-temporal micromeasurements of the oxygen uptake in the developing chick embryo.

    Science.gov (United States)

    Kucera, P; Raddatz, E

    1980-02-01

    A method has been developed for the determination of the oxygen uptake of small areas (0.01 mm2) in an entire chick embryo cultured in vitro under defined metabolic conditions. It is based on the recordings of the spectral changes of the hemoglobin used as oxygen source for the respiring tissue (Barzu and Borza, 1967). Rapid scanning of the hemoglobin absorbance over the preparation allows a comparison of the O2 uptake of various regions. Values of the order of 10(-2) 1 O2 . min-2 are measured in less than 10 sec with a spatial resolution of 100 micron. The differentiation of embryonic tissue is not disturbed by the measurements. The O2 diffusion in the media and in the tissue has been analyzed by digital simulation. The O2 uptake of the Hensen's node was measured from embryos starting at the stage of definitive primitive streak (stage 4) up to the stage of 10 somites. It increases from 0.6 to 1.1 nl . h-1 with a marked acceleration between stages 4 and 5. The values corrected for the protein content of the Hensen's node at stage 4, 5, 6 and 8 are 32, 30 and 28 microliter . mg-1 . h-1 respectively. The first scanning results show different patterns of the O2 utake at the level of the Hensen's node and of the neural plate. At stage 6-7, the corrected O2 uptake is 30 microliter . mg-1 . h-1 for . the former and 43 microliter . mg-1 . h-1 for the latter.

  12. Effect of antithyroid drug on chick embryos during the last week of development: delayed hatching and decreased cerebellar acetylcholinesterase activity.

    Science.gov (United States)

    Haba, Gen; Nishigori, Hidekazu; Tezuka, Yu; Kagami, Keisuke; Sugiyama, Toru; Nishigori, Hideo

    2011-11-01

    Hypothyroid state during embryogenesis disturbs normal growth and brain development, influencing later life. To evaluate the harmful consequences of the state during embryogenesis using an animal model, we inhibited thyroid hormone biosynthesis in chick embryos by using methimazole (MMI). Typically, embryos were treated with MMI (20 µmol/egg) on day 14, and examined on specific days.  Of the control embryos, 94% hatched on day 21, whereas 0% and 60% of MMI-treated embryos hatched on days 21 and 24, respectively. MMI retarded the rates of bodyweight gain as well as liver and heart development, and delayed hatching. However, the external differences in appearance and differences in the weights of the newly hatched control chicks on day 21 and the MMI-treated chicks on day 24 were less obvious. Embryos treated with MMI exhibited increased mass in their brain parts on day 24. Most notably, the treatment resulted in a 1.35-fold increase in cerebellum weight compared to that of the untreated animals. Acetylcholinesterase activity in the cerebellum on the day of hatching decreased significantly to 0.85-fold that of the untreated controls. Thyroid hormone receptor β mRNA was detected from day 12 and dramatically expressed from day 19 to the day of hatching. The 'fertilized hen's egg-chick embryo-chick system' is an appropriate animal model for investigating the hypothyroid state during embryogenesis. Decreased cerebellar acetylcholinesterase activity after MMI treatment was assumed to relate to a mechanism of motor and cognitive deficits in congenital hypothyroidism. © 2011 The Authors. Journal of Obstetrics and Gynaecology Research © 2011 Japan Society of Obstetrics and Gynecology.

  13. Peroxidized mineral oil increases the oxidant status of culture media and inhibits in vitro porcine embryo development.

    Science.gov (United States)

    Martinez, C A; Nohalez, A; Ceron, J J; Rubio, C P; Roca, J; Cuello, C; Rodriguez-Martinez, H; Martinez, E A; Gil, M A

    2017-11-01

    The use of oils with undetected alterations is a long-recognized problem for in vitro embryo production systems. Since peroxides in oils have been associated with reduced embryo production outcomes, our goals were (1) to evaluate the effects of a batch of mineral oil (MO) that was suspected to be altered on the in vitro production of pig embryos and (2) to determine oil peroxide values throughout culture and the transfer of oxidant agents from oil to culture media. Sunflower oil, which has a completely different chemical composition than MO but a higher oxidative status, and unaltered MO were used as controls. Oocyte maturation, fertilization and embryo development were affected differently depending on the oil overlay used. While the suspected MO was not able to sustain in vitro maturation and fertilization, the oocytes incubated in the presence of sunflower oil were matured and fertilized similarly to those of the unaltered MO group. Moreover, the cleavage rate of presumed zygotes cultured under the suspected MO was severely reduced compared with those cultured under the other oils, and none of the cleaved embryos developed to the blastocyst stage. Although the cleavage rates in the sunflower oil and unaltered MO groups were similar, embryos cultured under sunflower oil also failed to develop to the blastocyst stage. Our results revealed that the suspected MO and sunflower oil had similar levels of peroxides and that these levels were much higher than those of the unaltered MO. The total oxidant status was higher in media incubated under peroxidized oils than in fresh media or media incubated without an oil overlay or under unaltered MO, indicating that oxidant agents were transferred to the incubation media. However, unlike the sunflower oil group, the culture media incubated under the suspected MO had high levels of total oxidant status and low levels of hydrogen peroxide and reactive oxygen species, suggesting the presence of other unknown oxidant agents in

  14. Evaluation of the safety and pharmacokinetics of the multi-targeted receptor tyrosine kinase inhibitor sunitinib during embryo-fetal development in rats and rabbits.

    Science.gov (United States)

    Patyna, S; Haznedar, J; Morris, D; Freshwater, K; Peng, G; Sukbuntherng, J; Chmielewski, G; Matsumoto, D

    2009-06-01

    Angiogenesis plays a key role in embryo-fetal development and, based on nonclinical safety data, the majority of vascular endothelial growth factor (VEGF)-targeted antiangiogenic agents used in cancer therapy are not recommended during pregnancy. We investigated the effects of sunitinib (an oral inhibitor of multiple receptor tyrosine kinases [RTKs] including VEGF-receptors) on embryo-fetal development. Presumed-pregnant Sprague-Dawley rats and New Zealand White rabbits received repeated daily oral doses of sunitinib (0-30 mg/kg/day), during the major period of organogenesis. Clinical/physical examinations were performed throughout the gestation phase, and blood samples were collected to determine systemic exposure. Necropsy (including uterine examination) was performed on all animals and fetal morphology was examined. The no-observed-adverse-effect level was 1-5 mg/kg/day for maternal toxicity and 3 mg/kg/day for developmental toxicity in rats; 1 and 0.5 mg/kg/day, respectively, in rabbits. Embryo-fetal toxicity included decreases in the number of live fetuses and increases in the numbers of resorptions and post-implantation/complete litter losses; these were observed at doses of > or =5 mg/kg/day in rats and 5 mg/kg/day in rabbits. Malformations included fetal skeletal malformations (generally thoracic/lumbar vertebral alterations) in rats and cleft lip/palate in rabbits. These developmental effects were observed at approximately 5.5- (rats) and approximately 0.3-times (rabbits) the human systemic exposure at the approved sunitinib dose (50 mg/day). Similar effects have been reported with the prototype monoclonal antibody bevacizumab. As is typically observed for potent inhibitors of RTKs involved in angiogenesis, sunitinib was associated with embryo-fetal developmental toxicity in rats and rabbits at clinically relevant dose levels. 2009 Wiley-Liss, Inc.

  15. The Effect of Oral Morphine Consumption on Ependymal Duct and Spinal Cord Development in Wistar Rats Embryos

    Directory of Open Access Journals (Sweden)

    Masoomeh Kazemi

    2011-04-01

    Full Text Available Background: Previous studies have shown that morphine consumption during pregnancy may delay embryo development or cause abnormal nervous system function. The present study focused on the effects of maternal morphine consumption on ependyma duct and spinal cord development in Wistar rats. Methods: Wistar rats (170-200g were used throughout. The experimental groups after fertility received 0.05 mg/ml of morphine by tap water while, the control group received water. On 17th day of pregnancy, the pregnant animals were anesthetized by chloroform and the embryos were removed surgically. The embryos were fixed in formalin 10% for 4 weeks. Then, tissue processing, sectioning and staining hematoxylin and eosin (H&E, were applied for the embryos. The sections were examined for ependyma duct and spinal cord development by light microscope and MOTIC ,SPSS software. Results: Severe reduction of the area ependyma duct and an increase in the marginal layer of spinal cord area were observed in the experimental group. In addition, an increase in the mantle layer area and number cells of spinal cord in the experimental group regarding to controls was identified. Conclusion: The study showed that oral morphine consumption has caused to a decrease ependyma duct and spinal cord .This defect may cause postponed on function and development central neuron system. such as, changes observed in the fetus born by opioid addicted women.

  16. Effective Silencing of Sry Gene with RNA Interference in Developing Mouse Embryos Resulted in Feminization of XY Gonad

    Directory of Open Access Journals (Sweden)

    Ning Wu

    2012-01-01

    Full Text Available Delivering siRNA or shRNA into the developing embryos is still a main challenge to use of RNAi in mammalian systems. Here we analyze several factors influencing RNAi-mediated silencing of Sry gene, which is a tightly controlled spatiotemporal expressed gene and only shortly expressed in developing mouse embryo gonad. A Sry gene-specific shRNAs expression vector (pSilencer4.1/Sry565 was constructed. The shRNA constructs were mixed with polyethylenimines (PEIs to form a complex and then injected into pregnant mice though tail vein. Our results showed that Sry gene was downregulated significantly in developing embryos. Further study revealed that knocking-down of Sry expression resulted in feminization of gonad development in mouse embryos and the expression level of Sox9 and Wt1 gene was also significantly changed by downregulation of Sry. The transfection efficiency is associated with the amount of plasmid DNA injection, injection time, injection speed, and volume. Our studies suggest that transplacental RNAi could be implemented by tail vein injection of plasmid vector into pregnant mice.

  17. Vertical transmission and successive location of symbiotic bacteria during embryo development and larva formation in Corticium candelabrum (Porifera: Demospongiae)

    OpenAIRE

    Caralt Bosch, de, S.; Uriz, M. J.; Wijffels, R.H.

    2007-01-01

    This study reports on the transfer of heterotrophic bacteria from parental tissue to oocytes in the Mediterranean bacteriosponge Corticium candelabrum (Homosclerophorida) and the description of the successive locations of the microsymbionts during embryo development through transmission and scanning electron microscopy. Eight different types of symbiotic bacteria are described morphologically. These eight bacteria morphotypes are found in both adult individuals and larvae. Sym...

  18. Development of the pancreas in medaka, Oryzias latipes, from embryo to adult.

    Science.gov (United States)

    Otsuka, Takayoshi; Tsukahara, Tatsuya; Takeda, Hiroyuki

    2015-10-01

    To address conserved and unique features of fish pancreas development, we performed extensive analyses of pancreatic development in medaka embryos and adults using pdx1- and ptf1a-transgenic medaka, in situ hybridization and immunohistochemistry. The markers used in these analyses included pdx1, nkx6.1, nkx6.2, nkx2.2, Islet1, insulin, Somatostatin, glucagon, ptf1a, ela3l, trypsin, and amylase. The double transgenic (Tg) fish produced in the present study visualizes the development of endocrine (pdx1+) and exocrine (ptf1a+) parts simultaneously in living fishes. Like other vertebrates, the medaka pancreas develops as two (dorsal and ventral) buds in the anterior gut tube, which soon fuse into a single anlagen. The double Tg fish demonstrates that the differential property between the two buds is already established at the initial phase of bud development as indicated by strong pdx1 expression in the dorsal one. This Tg fish also allowed us to examine the gross morphology and the structure of adult pancreas and revealed unique characters of medaka pancreas such as broad and multiple connections with the gut tube along the anterior-posterior axis. © 2015 Japanese Society of Developmental Biologists.

  19. Comparative expression pattern analysis of WUSCHEL-related homeobox 2 (WOX2) and WOX8/9 in developing seeds and somatic embryos of the gymnosperm Picea abies.

    Science.gov (United States)

    Palovaara, Joakim; Hallberg, Henrik; Stasolla, Claudio; Hakman, Inger

    2010-10-01

    • In seed plants, current knowledge concerning embryonic pattern formation by polar auxin transport (PAT) and WUSCHEL-related homeobox (WOX) gene activity is primarily derived from studies on angiosperms, while less is known about these processes in gymnosperms. In view of the differences in their embryogeny, and the fact that somatic embryogenesis is used for mass propagation of conifers, a better understanding of embryo development is vital. • The expression patterns of PaWOX2 and PaWOX8/9 were followed with quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH) during seed and somatic embryo development in Norway spruce (Picea abies), and in somatic embryos treated with the PAT inhibitor N-1-naphthylphthalamic acid (NPA). • Both PaWOX2 and PaWOX8/9 were highly expressed at the early growth stages of zygotic and somatic embryos, and shared a similar expression pattern over the entire embryo. At later embryo stages, high expression of PaWOX8/9 became restricted to cotyledon primordia, epidermis, procambium and root apical meristem (RAM), which became most evident in NPA-treated somatic embryos, while expression of PaWOX2 was much lower. • Our results suggest an ancestral role of WOX in seed plant embryo development, and strengthen the proposed connection between PAT, PIN-FORMED (PIN) and WOX in the regulation of embryo patterning in seed plants.

  20. Assessment of imaging parameters correlated with the effects of cryopreservation on embryo development

    Science.gov (United States)

    Zarnescu, Livia; Abeyta, Mike; Baer, Thomas M.; Behr, Barry; Ellerbee, Audrey K.

    2014-03-01

    Embryo cryopreservation is an increasingly common technique that allows patients to undergo multiple cycles of in vitro fertilization (IVF) without being subjected to repeated ovarian stimulation and oocyte retrieval. There are two types of cryopreservation commonly used in IVF clinics today: slow freezing and vitrification. Because vitrification has been shown to result in higher rates of embryo survival post-thaw compared to slow freezing, it is rapidly gaining popularity in clinics worldwide. However, several studies have shown that vitrification can still cause damage to embryos in the form of DNA fragmentation, altered mitochondrial distribution and changes in transcriptional activity, all of which are impossible to assess noninvasively. In this paper we demonstrate a new method of quantitatively and noninvasively assessing changes in embryo appearance due to vitrification. Using full-field optical coherence tomography (FF-OCT), we show that vitrification causes striking changes in the appearance of the cytoplasm that are not visible under conventional brightfield microscopy. Using an automated algorithm that extracts parameters to describe these changes, we show that these parameters can also predict viability in embryos that have undergone vitrification. An automated, noninvasive assessment of embryo viability after vitrification and thawing could have significant clinical impact: allowing clinicians to more accurately choose the most viable embryos to transfer back to patients could reduce the average number of IVF cycles that patients must undergo to achieve pregnancy.

  1. Supplementation with small-extracellular vesicles from ovarian follicular fluid during in vitro production modulates bovine embryo development

    Science.gov (United States)

    Andrade, Gabriella M.; del Collado, Maite; Sampaio, Rafael V.; Sangalli, Juliano R.; Silva, Luciano A.; Pinaffi, Fábio V. L.; Jardim, Izabelle B.; Cesar, Marcelo C.; Nogueira, Marcelo F. G.; Cesar, Aline S. M.; Coutinho, Luiz L.; Pereira, Rinaldo W.; Perecin, Felipe; Meirelles, Flávio V.

    2017-01-01

    Pregnancy success results from the interaction of multiple factors, among them are folliculogenesis and early embryonic development. Failure during these different processes can lead to difficulties in conception. Alternatives to overcome these problems are based on assisted reproductive techniques. Extracellular vesicles are cell-secreted vesicles present in different body fluids and contain bioactive materials, such as messenger RNA, microRNAs (miRNAs), and proteins. Thus, our hypothesis is that extracellular vesicles from follicular fluid from 3–6 mm ovarian follicles can modulate bovine embryo development in vitro. To test our hypothesis follicular fluid from bovine ovaries was aspirated and small-extracellular vesicles (extracellular vesicles (EVs) were utilized for functional experiments investigating their role in modulating messenger RNA, microRNA as well as global DNA methylation and hydroxymethylation levels of bovine blastocysts. EVs from 3–6 mm follicles were used for RNA-seq and miRNA analysis. Functional annotation analysis of the EVs transcripts revealed messages related to chromatin remodeling and transcriptional regulation. EVs treatment during oocyte maturation and embryo development causes changes in blastocyst rates, as well as changes in the transcription levels of genes related to embryonic metabolism and development. Supplementation with EVs from 3–6 mm follicles during oocyte maturation and early embryo development (until the 4-cell stage) increased the levels of bta-miR-631 (enriched in EVs from 3–6 mm follicles) in embryos. Interestingly, the addition of EVs from 3–6 mm follicles induced changes in global DNA methylation and hydroxymethylation levels compared to embryos produced by the standard in vitro production system. Our results indicate that the supplementation of culture media with EVs isolated from the follicular fluid of 3–6 mm follicles during oocyte maturation and early embryo development can partially modify

  2. Differences in the timing of cardio-respiratory development determine whether marine gastropod embryos survive or die in hypoxia.

    Science.gov (United States)

    Rudin-Bitterli, Tabitha S; Spicer, John I; Rundle, Simon D

    2016-04-01

    Physiological plasticity of early developmental stages is a key way by which organisms can survive and adapt to environmental change. We investigated developmental plasticity of aspects of the cardio-respiratory physiology of encapsulated embryos of a marine gastropod, Littorina obtusata, surviving exposure to moderate hypoxia (PO2 =8 kPa) and compared the development of these survivors with that of individuals that died before hatching. Individuals surviving hypoxia exhibited a slower rate of development and altered ontogeny of cardio-respiratory structure and function compared with normoxic controls (PO2 >20 kPa). The onset and development of the larval and adult hearts were delayed in chronological time in hypoxia, but both organs appeared earlier in developmental time and cardiac activity rates were greater. The velum, a transient, 'larval' organ thought to play a role in gas exchange, was larger in hypoxia but developed more slowly (in chronological time), and velar cilia-driven, rotational activity was lower. Despite these effects of hypoxia, 38% of individuals survived to hatching. Compared with those embryos that died during development, these surviving embryos had advanced expression of adult structures, i.e. a significantly earlier occurrence and greater activity of their adult heart and larger shells. In contrast, embryos that died retained larval cardio-respiratory features (the velum and larval heart) for longer in chronological time. Surviving embryos came from eggs with significantly higher albumen provisioning than those that died, suggesting an energetic component for advanced development of adult traits. © 2016. Published by The Company of Biologists Ltd.

  3. Release of sICAM-1 in oocytes and in vitro fertilized human embryos.

    Directory of Open Access Journals (Sweden)

    Monica Borgatti

    Full Text Available During the last years, several studies have reported the significant relationship between the production of soluble HLA-G molecules (sHLA-G by 48-72 hours early embryos and an increased implantation rate in IVF protocols. As consequence, the detection of HLA-G modulation was suggested as a marker to identify the best embryos to be transferred. On the opposite, no suitable markers are available for the oocyte selection.The major finding of the present paper is that the release of ICAM-1 might be predictive of oocyte maturation. The results obtained are confirmed using three independent methodologies, such as ELISA, Bio-Plex assay and Western blotting. The sICAM-1 release is very high in immature oocytes, decrease in mature oocytes and become even lower in in vitro fertilized embryos. No significant differences were observed in the levels of sICAM-1 release between immature oocytes with different morphological characteristics. On the contrary, when the mature oocytes were subdivided accordingly to morphological criteria, the mean sICAM-I levels in grade 1 oocytes were significantly decreased when compared to grade 2 and 3 oocytes.The reduction of the number of fertilized oocytes and transferred embryos represents the main target of assisted reproductive medicine. We propose sICAM-1 as a biochemical marker for oocyte maturation and grading, with a possible interesting rebound in assisted reproduction techniques.

  4. Human Capital and Development

    OpenAIRE

    Rodolfo E. Manuelli

    2015-01-01

    Perhaps no question has attracted as much attention in the economics literature as “Why are some countries richer than others?” In this article, the author revisits the “development problem” and provides some estimates of the importance of human capital in accounting for cross-country differences in output per worker. His results suggest that human capital has a central role in determining the wealth of nations and that the quality of human capital varies systematically with the level of deve...

  5. Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos.

    Directory of Open Access Journals (Sweden)

    Amber L Famiglietti

    Full Text Available Mucin-type O-glycosylation is a ubiquitous posttranslational modification in which N-Acetylgalactosamine (GalNAc is added to the hydroxyl group of select serine or threonine residues of a protein by the family of UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferases (GalNAc-Ts; EC 2.4.1.41. Previous studies demonstrate that O-glycosylation plays essential roles in protein function, cell-cell interactions, cell polarity and differentiation in developing mouse and Drosophila embryos. Although this type of protein modification is highly conserved among higher eukaryotes, little is known about this family of enzymes in echinoderms, basal deuterostome relatives of the chordates. To investigate the potential role of GalNAc-Ts in echinoderms, we have begun the characterization of this enzyme family in the purple sea urchin, S. purpuratus. We have fully or partially cloned a total of 13 genes (SpGalnts encoding putative sea urchin SpGalNAc-Ts, and have confirmed enzymatic activity of five recombinant proteins. Amino acid alignments revealed high sequence similarity among sea urchin and mammalian glycosyltransferases, suggesting the presence of putative orthologues. Structural models underscored these similarities and helped reconcile some of the substrate preferences observed. Temporal and spatial expression of SpGalnt transcripts, was studied by whole-mount in situ hybridization. We found that many of these genes are transcribed early in developing embryos, often with restricted expression to the endomesodermal region. Multicolor fluorescent in situ hybridization (FISH demonstrated that transcripts encoding SpGalnt7-2 co-localized with both Endo16 (a gene expressed in the endoderm, and Gcm (a gene expressed in secondary mesenchyme cells at the early blastula stage, 20 hours post fertilization (hpf. At late blastula stage (28 hpf, SpGalnt7-2 message co-expresses with Gcm, suggesting that it may play a role in secondary mesenchyme development. We

  6. [Effects of cytosine-arabinofuranoside on the development of reptilian embryos (Lacerta viridis, Laur. and Anguis fragilis, L.)].

    Science.gov (United States)

    Raynaud, A

    1982-01-01

    Administered into the yolk sac of eggs of Lacerta viridis as a single dose of 17 to 40 micrograms, cytosine-arabinoside (Ara-C) was compatible with survival of the embryo, from the sixth day of incubation, for at least 20 to 25 days. The LD50 was 40 to 50 micrograms per egg. Doses of 20 to 40 micrograms of Ara-C introduced in the yolk sac of eggs of the slow-worm (Anguis fragilis) cultured in vitro, at stages of the allantoid bud of 0,5 mm to 2,5 mm long, killed the embryo in 4 to 8 days (possibly due to alterations of capillary blood vessels of allantois and area vasculosa). In the two species, these doses caused cytotoxic effects on embryonic proliferating tissues, growth inhibition and a variety of developmental defects. In young embryos of Anguis fragilis, similar doses of 20 to 40 micrograms of Ara-C caused, in 2 to 4 days, death of many cells in the anlagen of growing organs: neural tube, sensory organs, bronchi, mesoderm of the limb bud, subcutaneous mesenchyme, anlage of dorsal skeletal structures, etc.; followed by growth inhibition and malformations. On the other hand, in the limb bud, the apical ridge was less retrogressed than in control embryos; the limb buds showed slightly better development in treated embryos than in controls, but, Ara-C induced severe damage in their mesoderm. In all embryos of Lacerta viridis, treated at the stage of 6 days or of 10 days of incubation by doses of 20 to 40 micrograms of Ara-C and killed 15 to 35 days later, there was a general reduction of size and of weight and external and internal malformations, more or less severe, were present: modifications of the form of the head, shortening of the lower jaw, labial clefts, microphthalmia, micromelia and other limbs defects, developmental defects of the tail. In some embryos, the only external defects observed were missing fingers and toes; in three of these embryos, the same digits were missing in the four limbs. Modifications of limb morphogenesis induced by Ara-C are

  7. Sublethal effects of atrazine on embryo-larval development of Rhinella arenarum (Anura: Bufonidae).

    Science.gov (United States)

    Svartz, Gabriela V; Herkovits, Jorge; Pérez-Coll, Cristina S

    2012-05-01

    Atrazine (ATR), one of the most widely used herbicides in the world, affects not only target organisms but also the biota in general. Here, the teratogenic and neurotoxic effects of ATR on Rhinella arenarum (South American toad) embryos, and larvae were evaluated by means of standardized bioassays during acute and chronic exposures. The herbicide had a significant incidence of malformations, with a Teratogenic Index (TI) of 3.28. The main effects were delayed development, reduced body size, microcephaly, axial flexures, wavy tail and edema. In addition, delayed development, reduced development of forelimbs, and edema were recorded at metamorphosis stages. Scanning electron microscopy allowed observing different degrees of cellular dissociation and persistent cilliar cells in specific regions like the adhesive structure and tail fin. Results obtained by ATR 24 h pulse exposures at six developmental stages pointed out blastula as the most susceptible developmental stage both for immediate and delayed adverse effects. A noteworthy recovery capacity from acute toxic effects was recorded from the neural plate stage onwards. Regarding neurotoxic effects, abnormal, and erratic swimming and spasmodic contractions were recorded. Both the teratogenic and neurotoxic effects reported in this study demonstrate the importance of evaluating sublethal effects in non-target organisms as they could imply reduced fitness of individuals and eventually a population decline. The Hazard Quotients (HQ) for ATR ranged from 0.14 to 10.80, and the fact that some of these values are above USEPA's level of concern indicate that ATR is likely a risk to R. arenarum.

  8. The Bsister MADS gene FST determines ovule patterning and development of the zygotic embryo and endosperm.

    Directory of Open Access Journals (Sweden)

    Dong Sun Lee

    Full Text Available Many homeotic MADS-box genes have been identified as controllers of the floral transition and floral development. However, information regarding Bsister (Bs-function genes in monocots is still limited. Here, we describe the functional characterization of a Bs-group MADS-box gene FEMALE-STERILE (FST, whose frame-shift mutation (fst results in abnormal ovules and the complete abortion of zygotic embryos and endosperms in rice. Anatomical analysis showed that the defective development in the fst mutant exclusively occurred in sporophytic tissues including integuments, fertilized proembryos and endosperms. Analyses of the spatio-temporal expression pattern revealed that the prominent FST gene products accumulated in the inner integument, nucellar cell of the micropylar side, apical and base of the proembryos and free endosperm nuclei. Microarray and gene ontology analysis unraveled substantial changes in the expression level of many genes in the fst mutant ovules and seeds, with a subset of genes involved in several developmental and hormonal pathways appearing to be down-regulated. Using both forward and reverse genetics approaches, we demonstrated that rice FST plays indispensable roles and multiple functions during ovule and early seed development. These findings support a novel function for the Bs-group MADS-box genes in plants.

  9. Potential toxicity of engineered nanoparticles in mammalian germ cells and developing embryos: treatment strategies and anticipated applications of nanoparticles in gene delivery.

    Science.gov (United States)

    Das, Joydeep; Choi, Yun-Jung; Song, Hyuk; Kim, Jin-Hoi

    2016-09-01

    Engineered nanoparticles (ENPs) offer technological advantages for a variety of industrial and consumer products as well as show promise for biomedical applications. Recent progress in the field of nanotechnology has led to increased exposure to nanoparticles by humans. To date, little is known about the adverse effects of these ENPs on reproductive health, although interest in nanotechnology area is growing. A few biocompatible ENPs have a high loading capacity for exogenous substances, including drugs, DNA or proteins, and can selectively deliver molecular cargo into cells; however, they represent a potential tool for gene delivery into gametes and embryos. Understanding the reprotoxicological aspects of these ENPs is of the utmost importance to reliably estimate its potential impact on human health. In addition, a search for protective agents to combat ENP-mediated reproductive toxicity is warranted. Therefore, in this review we summarize the toxic effects of a few ENPs (metal and metal oxides, carbon-based nanoparticles, quantum dots and chitosan) in mammalian germ cells and developing embryos, and propose some treatment strategies that could mitigate nanoparticle-mediated toxicity. In addition, we outline the anticipated applications of ENPs in transgenic animal production in order to generate models for investigations into the mechanisms for human disease. A literature search was performed using the National Center for Biotechnology Information PubMed database up until March 2016 and relevant keywords were used to obtain information regarding mammalian germ cell-specific toxicity and embryotoxicity of ENPs, possible treatment strategies, as well as the anticipated applications of nanoparticles in gene delivery in germ cells and embryos. Only English language publications were included. Here, we demonstrate the toxicological effects of ENPs in mammalian germ cells and developing embryos by considering both in vitro and in vivo experimental models based on the

  10. Late embryos and bony skull development in Bothropoides jararaca (Serpentes, Viperidae).

    Science.gov (United States)

    Polachowski, Katja M; Werneburg, Ingmar

    2013-02-01

    In recent years, developmental anatomy received increasing interest as a potential new source for phylogenetic research. For skeletal development, studies mainly rely on the first appearance of ossification centers. However, informative events occur during the whole course of skeletogenesis; interactions between external and internal development occur and morphometric changes take place - all of which present potential sources for phylogenetic analyses. Therefore, the Standard Event System (SES) was used to traceably describe the external development of the snake species Bothropoides jararaca and external measurements were analyzed. We then applied micro-computed tomography (μCT), clearing and double-staining, and 2D and 3D morphometric methods to describe, illustrate, and analyze the development of the head in great detail. We found a 3D flattening of the skull during ontogeny, a pattern that is not reflected in external development. This may be explained by a different relationship of skeletogenesis and external characters to the developing jaw musculature or simply by the different type of data. Clearing and double-staining and μCT-scanning revealed a broadly similar sequence in the onset of ossification. Minute differences may be due to the treatment of embryos. Bones of the dermatocranium are among the first to ossify and the development of the calcified endolymph may reflect its function as a calcium source during development. The value of phylogenetic observations using the sequence of first ossifications is critically discussed. The related heterochronic changes are interpreted to contribute at least to the very first phase of divagating skull formation among taxa. Copyright © 2012 Elsevier GmbH. All rights reserved.

  11. ARID1A, a component of SWI/SNF chromatin remodeling complexes, is required for porcine embryo development.

    Science.gov (United States)

    Tseng, Yu-Chun; Cabot, Birgit; Cabot, Ryan A

    2017-12-01

    Mammalian embryos undergo dramatic epigenetic remodeling that can have a profound impact on both gene transcription and overall embryo developmental competence. Members of the SWI/SNF (Switch/Sucrose non-fermentable) family of chromatin-remodeling complexes reposition nucleosomes and alter transcription factor accessibility. These large, multi-protein complexes possess an SNF2-type ATPase (either SMARCA4 or SMARCA2) as their core catalytic subunit, and are directed to specific loci by associated subunits. Little is known about the identity of specific SWI/SNF complexes that serve regulatory roles during cleavage development. ARID1A, one of the SWI/SNF complex subunits, can affect histone methylation in somatic cells; here, we determined the developmental requirements of ARID1A in porcine oocytes and embryos. We found ARID1A transcript levels were significantly reduced in 4-cell porcine embryos as compared to germinal vesicle-stage oocytes, suggesting that ARID1A would be required for porcine cleavage-stage development. Indeed, injecting in vitro-matured and fertilized porcine oocytes with double-stranded interfering RNAs that target ARID1A, and evaluating their phenotype after seven days, revealed that the depletion of ARID1A results in significantly fewer cells than their respective control groups (p < 0.001). © 2017 Wiley Periodicals, Inc.

  12. Effects of ulipristal acetate on human embryo attachment and endometrial cell gene expression in an in vitro co-culture system.

    Science.gov (United States)

    Berger, C; Boggavarapu, N R; Menezes, J; Lalitkumar, P G L; Gemzell-Danielsson, K

    2015-04-01

    Does ulipristal acetate (UPA) used for emergency contraception (EC) interfere with the human embryo implantation process? UPA, at the dosage used for EC, does not affect human embryo implantation process, in vitro. A single pre-ovulatory dose of UPA (30 mg) acts by delaying or inhibiting ovulation and is recommended as first choice among emergency contraceptive pills due to its efficacy. The compound has also been demonstrated to have a dose-dependent effect on the endometrium, which theoretically could impair endometrial receptivity but its direct action on human embryo implantation has not yet been studied. Effect of UPA on embryo implantation process was studied in an in vitro endometrial construct. Human embryos were randomly added to the cultures and cultured for 5 more days with UPA (n = 10) or with vehicle alone (n = 10) to record the attachment of embryos. Endometrial biopsies were obtained from healthy, fertile women on cycle day LH+4 and stromal and epithelial cells were isolated. A three-dimensional in vitro endometrial co-culture system was constructed by mixing stromal cells with collagen covered with a layer of epithelial cells and cultured in progesterone containing medium until confluence. The treatment group received 200 ng/ml of UPA. Healthy, viable human embryos were placed on both control and treatment cultures. Five days later the cultures were tested for the attachment of embryos and the 3D endometrial constructs were analysed for endometrial receptivity markers by real-time PCR. There was no significant difference in the embryo attachment rate between the UPA treated group and the control group as 5 out of 10 human embryos exposed to UPA and 7 out of 10 embryos in the control group attached to the endometrial cell surface (P = 0.650). Out of 17 known receptivity genes studied here, only 2 genes, HBEGF (P = 0.009) and IL6 (P = 0.025) had a significant up-regulation and 4 genes, namely HAND2 (P = 0.003), OPN (P = 0.003), CALCR (P = 0.016) and

  13. Embryo culture in teratological surveillance and serum proteins in development. Progress report, 1979-1980

    Energy Technology Data Exchange (ETDEWEB)

    Klein, N.W.

    1980-07-01

    Research progress for the period 1979-1980 is reported. The feasibility of using rat embryo cultures to test the teratogenic activity of serum was studied. The mechanisms regulating the synthesis of serum proteins were investigated. (ACR)

  14. The Relationship between Cell Number, Division Behavior and Developmental Potential of Cleavage Stage Human Embryos: A Time-Lapse Study.

    Science.gov (United States)

    Kong, Xiangyi; Yang, Shuting; Gong, Fei; Lu, Changfu; Zhang, Shuoping; Lu, Guangxiu; Lin, Ge

    2016-01-01

    Day 3 cleavage embryo transfer is routine in many assisted reproductive technology centers today. Embryos are usually selected according to cell number, cell symmetry and fragmentation for transfer. Many studies have showed the relationship between cell number and embryo developmental potential. However, there is limited understanding of embryo division behavior and their association with embryo cell number and developmental potential. A retrospective and observational study was conducted to investigate how different division behaviors affect cell number and developmental potential of day 3 embryos by time-lapse imaging. Based on cell number at day 3, the embryos (from 104 IVF/intracytoplasmic sperm injection (ICSI) treatment cycles, n = 799) were classified as follows: less than 5 cells (10C; n = 42). Division behavior, morphokinetic parameters and blastocyst formation rate were analyzed in 5 groups of day 3 embryos with different cell numbers. In 10C embryos increased compared to 7-8C embryos (45.8%, 33.3% vs. 11.1%, respectively). In ≥5C embryos, FR and DC significantly reduced developmental potential, whereas potential irrespective of division behaviors. In NB embryos, the blastocyst formation rate increased with cell number from 7.4% (10C). In NB embryos, the cell cycle elongation or shortening was the main cause for abnormally low or high cell number, respectively. After excluding embryos with abnormal division behaviors, the developmental potential, implantation rate and live birth rate of day 3 embryos increased with cell number.

  15. Post-hatching development of the porcine and bovine embryo-defining criteria for expected development in vivo and in vitro

    DEFF Research Database (Denmark)

    Vejlsted, Morten; Du, Yutao; Vajta, Gábor

    2006-01-01

    Particular attention has been paid to the pre-hatching period of embryonic development although blastocyst development is a poor indicator of embryo viability. Post-hatching embryonic dev elopment in vitro would allow for establishment of more accurate tools for evaluating developmental potential...... without the need for transfer to recipient animals. Such a system would require (1) definition of milestones of expected post-hatching embryonic development in vivo; and (2) development of adequate culture systems. We propose a stereomicroscopical staging system for post-hatching embryos defining...

  16. Reflectin genes and development of iridophore patterns in Sepia officinalis embryos (Mollusca, Cephalopoda).

    Science.gov (United States)

    Andouche, Aude; Bassaglia, Yann; Baratte, Sébastien; Bonnaud, Laure

    2013-05-01

    In the cuttlefish Sepia officinalis, iridescence is known to play a role in patterning and communication. In iridophores, iridosomes are composed of reflectins, a protein family, which show great diversity in all cephalopod species. Iridosomes are established before hatching, but very little is known about how these cells are established, their distribution in embryos, or the contribution of each reflectin gene to iridosome structures. Six reflectin genes are expressed during the development of iridosomes in Sepia officinalis. We show that they are expressed in numerous parts of the body before hatching. Evidence of the colocalization of two different genes of reflectin was found. Curiously, reflectin mRNA expression was no longer detectable at the time of hatchling, while reflectin proteins were present and gave rise to visible iridescence. These data suggest that several different forms of reflectins are simultaneously used to produce iridescence in S. officinalis and that mRNA production and translation are decoupled in time during iridosome development. Copyright © 2013 Wiley Periodicals, Inc.

  17. Effect of copper nanoparticles and copper sulphate on metabolic rate and development of broiler embryos

    DEFF Research Database (Denmark)

    Scott, Abdullah Talal Abudllah; Vadalasetty, Krishna Prasad; Sawosz, E.

    2016-01-01

    Copper (Cu) is regularly used as a growth promoter in poultry production. However, it has been demonstrated that the content of Cu inside eggs might not be sufficient to support the embryonic development. It is possible to supply the embryo with extra nutrients by in-ovo administration. Recently...... consumption — O2 and energy expenditure — EE) and development during embryogenesis. Fertilised broiler eggs were divided into six groups: a non-injected control, a placebo injected with demineralised water, two groups injected, at day one of incubation, with CuSO4 (50 and 100 mg/kg) and two groups injected....../kg Cu-NP and CuSO4 significantly increased O2 consumption and EE on the 16th and 19th day of incubation compared with the control group; Cu-NP had the largest effect on the metabolic rate. However, organ weights (intestine, heart, liver, and breast) relative to the yolk-free body weight were...

  18. Immunocytochemical localization and subcellular site of lectin synthesis in developing wheat embryos

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, N.V.; Peumans, W.J.

    1986-04-01

    Appearance of wheat germ agglutinin (WGA) during wheat germ embryogenesis was studied using indirect immunocytochemical methods at the light and electron microscope levels. Developing embryos were labelled with (/sup 35/S) cysteine in pulse and pulse-chase experiments to study the synthesis and transport of the lectin to protein bodies (PB). The radical, and coleorhiza first accumulated WGA around 10 days post anthesis (DPA), while WGA was found in the epiblast and coleoptile 15 and 20 DPA respectively. At the subcellular level WGA can be seen first in a small developing PB which later fused with larger ones. WGA was distributed evenly throughout the PB. When tissue was pulse-labelled, fractionated on an isopycnic sucrose gradient and exposed to detergent, the incorporated radioactivity of released lectin coincided with the position of the endoplasmic reticulum (ER) marker enzyme NADH-cytochrome c reductase. Both radioactivity and enzyme activity shifted to a higher density in the presence of 2 mM Mg acetate, indicating that radioactive lectin was associated with the rough ER.

  19. Human Endometrial Exosomes Contain Hormone-Specific Cargo Modulating Trophoblast Adhesive Capacity: Insights into Endometrial-Embryo Interactions.

    Science.gov (United States)

    Greening, David W; Nguyen, Hong P T; Elgass, Kirstin; Simpson, Richard J; Salamonsen, Lois A

    2016-02-01

    Embryo implantation into receptive endometrium requires synergistic endometrial-blastocyst interactions within the uterine cavity and is essential for establishing pregnancy. We demonstrate that exosomes (40-150 nm nanovesicles) released from endometrial epithelial cells are an important component of these interactions. We defined the proteome of purified endometrial epithelial-derived exosomes (Exos) influenced by menstrual cycle hormones estrogen (E; proliferative phase) and estrogen plus progesterone (EP; receptive phase) and examined their potential to modify trophoblast function. E-/EP-Exos were uniquely enriched with 254 and 126 proteins, respectively, with 35% newly identified proteins not previously reported in exosome databases. Importantly, EP-Exos protein cargo was related to fundamental changes in implantation: adhesion, migration, invasion, and extracellular matrix remodeling. These findings from hormonally treated ECC1 endometrial cancer cells were validated in human primary uterine epithelial cell-derived exosomes. Functionally, exosomes were internalized by human trophoblast cells and enhanced their adhesive capacity, a response mediated partially through active focal adhesion kinase (FAK) signaling. Thus, exosomes contribute to the endometrial-embryo interactions within the human uterine microenvironment essential for successful implantation. © 2016 by the Society for the Study of Reproduction, Inc.

  20. The Necessity of OCT-4 and CDX2 for Early Development and Gene Expression Involved in Differentiation of Inner Cell Mass and Trophectoderm Lineages in Bovine Embryos.

    Science.gov (United States)

    Sakurai, Nobuyuki; Takahashi, Kazuki; Emura, Natsuko; Fujii, Takashi; Hirayama, Hiroki; Kageyama, Soichi; Hashizume, Tsutomu; Sawai, Ken

    2016-10-01

    The functions of POU class 5 transcription factor 1 (Oct-4) and caudal-type homeobox 2 (Cdx2) in the differentiation of the murine inner cell mass (ICM) and trophectoderm (TE) have been described in detail. However, little is known about the roles of OCT-4 and CDX2 in preimplantation bovine embryos. To elucidate their functions during early development in bovine embryos, we performed OCT-4 and CDX2 downregulation using RNA interference. We injected OCT-4- or CDX2-specific short interfering RNAs (siRNAs) into bovine zygotes. The rate of blastocyst development of OCT-4-downregulated embryos was lower compared with uninjected or control siRNA-injected embryos. Gene expression analysis revealed decreased CDX2 and fibroblast growth factor 4 expression in OCT-4-downregulated embryos. CDX2-downregulated embryos developed to the blastocyst stage; however, in most cases, blastocoel formation was delayed. Gene expression analysis revealed decreased GATA3 expression and elevated NANOG expression in CDX2-downregulated embryos. In conclusion, OCT-4 and CDX2 are essential for early development and gene expression involved in differentiation of ICM and TE lineages in bovine embryos.

  1. "Healthy" Human Development Indices

    Science.gov (United States)

    Engineer, Merwan; Roy, Nilanjana; Fink, Sari

    2010-01-01

    In the Human Development Index (HDI), life expectancy is the only indicator used in modeling the dimension "a long and healthy life". Whereas life expectancy is a direct measure of quantity of life, it is only an indirect measure of healthy years lived. In this paper we attempt to remedy this omission by introducing into the HDI the morbidity…

  2. Developing Human Performance Measures

    Energy Technology Data Exchange (ETDEWEB)

    Jeffrey Joe; Bruce Hallbert; Larry Blackwood; Donald Dudehoeffer; Kent Hansen

    2006-05-01

    Through the reactor oversight process (ROP), the U.S. Nuclear Regulatory Commission (NRC) monitors the performance of utilities licensed to operate nuclear power plants. The process is designed to assure public health and safety by providing reasonable assurance that licensees are meeting the cornerstones of safety and designated crosscutting elements. The reactor inspection program, together with performance indicators (PIs), and enforcement activities form the basis for the NRC’s risk-informed, performance based regulatory framework. While human performance is a key component in the safe operation of nuclear power plants and is a designated cross-cutting element of the ROP, there is currently no direct inspection or performance indicator for assessing human performance. Rather, when human performance is identified as a substantive cross cutting element in any 1 of 3 categories (resources, organizational or personnel), it is then evaluated for common themes to determine if follow-up actions are warranted. However, variability in human performance occurs from day to day, across activities that vary in complexity, and workgroups, contributing to the uncertainty in the outcomes of performance. While some variability in human performance may be random, much of the variability may be attributed to factors that are not currently assessed. There is a need to identify and assess aspects of human performance that relate to plant safety and to develop measures that can be used to successfully assure licensee performance and indicate when additional investigation may be required. This paper presents research that establishes a technical basis for developing human performance measures. In particular, we discuss: 1) how historical data already gives some indication of connection between human performance and overall plant performance, 2) how industry led efforts to measure and model human performance and organizational factors could serve as a data source and basis for a

  3. The coincident time-space patterns of septate junction development in normal and exogastrulated sea urchin embryos.

    Science.gov (United States)

    Spiegel, E; Howard, L

    1985-11-01

    Earlier studies have shown that two types of septate junction are formed during early sea urchin morphogenesis. One type is the straight, unbranched, double septum septate (SUDS) which is found in the ectodermal layer throughout early development. The second type is formed only in cells which invaginate to become endoderm and to form the digestive tract. This junction is characterized by pleated, anastomosing, single septum septates (PASS). In order to ascertain in which parts of the digestive tract these junctions are formed, we studied exogastrulae because the endoderm is everted and forms constricted areas of the gut which are easily recognizable. Our results show that, in control embryos, SUDS septates are found in the mouth, esophagus and coelom and that PASS septates are found in the stomach, intestine and anus. These junctional types are also found in the same areas in exogastrulae; SUDS septates are found in the stomadeum, esophagus and coelom, and PASS septates are found in the stomach and intestine. The transition from SUDS to PASS junctions takes place within the same time period in exogastrulae as in normal embryos, i.e., from the time of mid-gastrulation through the pluteus stage. These results indicate that septate junction formation in the sea urchin embryo digestive tract may be genetically programmed in terms of both time and spatial location. This program is not altered either by the major dislocation of cells from their normal position within the embryo or from normal contacts with neighboring cells.

  4. The Role of Peroxisome Proliferator-Activated Receptors in the Development and Physiology of Gametes and Preimplantation Embryos

    Directory of Open Access Journals (Sweden)

    Jaou-Chen Huang

    2008-01-01

    Full Text Available In several species, a family of nuclear receptors, the peroxisome proliferator-activated receptors (PPARs composed of three isotypes, is expressed in somatic cells and germ cells of the ovary as well as the testis. Invalidation of these receptors in mice or stimulation of these receptors in vivo or in vitro showed that each receptor has physiological roles in the gamete maturation or the embryo development. In addition, synthetic PPARγ ligands are recently used to induce ovulation in women with polycystic ovary disease. These results reveal the positive actions of PPAR in reproduction. On the other hand, xenobiotics molecules (in herbicides, plasticizers, or components of personal care products, capable of activating PPAR, may disrupt normal PPAR functions in the ovary or the testis and have consequences on the quality of the gametes and the embryos. Despite the recent data obtained on the biological actions of PPARs in reproduction, relatively little is known about PPARs in gametes and embryos. This review summarizes the current knowledge on the expression and the function of PPARs as well as their partners, retinoid X receptors (RXRs, in germ cells and preimplantation embryos. The effects of natural and synthetic PPAR ligands will also be discussed from the perspectives of reproductive toxicology and assisted reproductive technology.

  5. Intrinsic vs. extrinsic influences on life history expression: metabolism and parentally induced temperature influences on embryo development rate

    Science.gov (United States)

    Martin, Thomas E.; Ton, Riccardo; Nikilson, Alina

    2013-01-01

    Intrinsic processes are assumed to underlie life history expression and trade-offs, but extrinsic inputs are theorised to shift trait expression and mask trade-offs within species. Here, we explore application of this theory across species. We do this based on parentally induced embryo temperature as an extrinsic input, and mass-specific embryo metabolism as an intrinsic process, underlying embryonic development rate. We found that embryonic metabolism followed intrinsic allometry rules among 49 songbird species from temperate and tropical sites. Extrinsic inputs via parentally induced temperatures explained the majority of variation in development rates and masked a relationship with metabolism; metabolism explained a minor proportion of the variation in development rates among species, and only after accounting for temperature effects. We discuss evidence that temperature further obscures the expected interspecific trade-off between development rate and offspring quality. These results demonstrate the importance of considering extrinsic inputs to trait expression and trade-offs across species.

  6. Contrasting Storage Protein Synthesis and Messenger RNA Accumulation during Development of Zygotic and Somatic Embryos of Alfalfa (Medicago sativa L.) 1

    Science.gov (United States)

    Krochko, Joan E.; Pramanik, Saroj K.; Bewley, J. Derek

    1992-01-01

    During development on hormone-free media, somatic embryos pass through distinct morphological stages that superficially resemble those of zygotic embryo development (globular, heart, torpedo, cotyledonary stages). Despite these similarities, they differ from zygotic embryos in the extent of cotyledonary development and the patterns of synthesis and quantitative expression of seed-specific storage proteins (7S, 11S, and 2S proteins). Alfin (7S) is the first storage protein synthesized in developing zygotic embryos (stage IV). The 11S (medicagin) and 2S (Low Molecular Weight, LMW) storage proteins are not detectable until the following stage of development (stage V), although all three are present before the completion of embryo enlargement. Likewise, the 7S storage protein is the first to be synthesized in developing somatic embryos (day 5). Medicagin is evident by day 7 and the LMW protein by day 10. In contrast to zygotic embryos, alfin remains the predominant storage protein in somatic embryos throughout development. Not only are the relative amounts of medicagin and the LMW protein reduced in somatic embryos but the LMW protein is accumulated much later than the other proteins. Quantification of the storage protein mRNAs (7S, 11S, and 2S) by northern blot analysis confirms that there are substantial differences in the patterns of message accumulation in zygotic and somatic embryos of alfalfa (Medicago sativa). In zygotic embryos, the 7S, 11S, and 2S storage protein mRNAs are abundant during maturation and, in particular, during the stages of maximum protein synthesis (alfin, stages VI and VII; medicagin, stage VII; LMW, stage VII). In somatic embryos, the predominance of the 7S storage protein is correlated with increased accumulation of its mRNA, whereas the limited synthesis of the 11S storage protein is associated with much lower steady-state levels of its message. The mRNA for the LMW protein is present already by 3 days after transfer to hormone-free media

  7. The Relationship between Cell Number, Division Behavior and Developmental Potential of Cleavage Stage Human Embryos: A Time-Lapse Study.

    Directory of Open Access Journals (Sweden)

    Xiangyi Kong

    Full Text Available Day 3 cleavage embryo transfer is routine in many assisted reproductive technology centers today. Embryos are usually selected according to cell number, cell symmetry and fragmentation for transfer. Many studies have showed the relationship between cell number and embryo developmental potential. However, there is limited understanding of embryo division behavior and their association with embryo cell number and developmental potential. A retrospective and observational study was conducted to investigate how different division behaviors affect cell number and developmental potential of day 3 embryos by time-lapse imaging. Based on cell number at day 3, the embryos (from 104 IVF/intracytoplasmic sperm injection (ICSI treatment cycles, n = 799 were classified as follows: less than 5 cells (10C; n = 42. Division behavior, morphokinetic parameters and blastocyst formation rate were analyzed in 5 groups of day 3 embryos with different cell numbers. In 10C embryos increased compared to 7-8C embryos (45.8%, 33.3% vs. 11.1%, respectively. In ≥5C embryos, FR and DC significantly reduced developmental potential, whereas 10C. In NB embryos, the cell cycle elongation or shortening was the main cause for abnormally low or high cell number, respectively. After excluding embryos with abnormal division behaviors, the developmental potential, implantation rate and live birth rate of day 3 embryos increased with cell number.

  8. Hematocrit and blood osmolality in developing chicken embryos (Gallus gallus): in vivo and in vitro regulation.

    Science.gov (United States)

    Andrewartha, Sarah J; Tazawa, Hiroshi; Burggren, Warren W

    2011-12-15

    Hematocrit (Hct) regulation is a complex process involving potentially many factors. How such regulation develops in vertebrate embryos is still poorly understood. Thus, we investigated the role of blood pH in the regulation of Hct across developmental time in chicken embryos. We hypothesized that blood pH alterations in vitro (i.e., in a test tube) would affect Hct far more than in vivo because of in vivo compensatory regulatory processes for Hct. Large changes in Hct (through mean corpuscular volume (MCV)) and blood osmolality (Osm) occur when the blood was exposed to varying ambient temperatures (T(a)'s) and P(CO2) in vitro alongside an experimentally induced blood pH change from ~7.3 to 8.2. However, homeostatic regulatory mechanisms apparently limited these alterations in vivo. Changes in blood pH in vitro were accompanied by hydration or dehydration of red blood cells depending on embryonic age, resulting in changes in Hct that also were specific to developmental stage, due likely to initial blood gas and [HCO(3)(-)](v) values. Significant linear relationships between Hct and pH (Hct/ΔpH=-21.4%/(pH unit)), Hct and [HCO(3)(-)] (ΔHct/Δ[HCO(3)(-)]=1.6%/(mEq L(-1))) and the mean buffer value (Δ[HCO(3)(-)]/ΔpH=-13.4 (mEq L(-1))/(pH unit)) demonstrate that both pH and [HCO(3)(-)] likely play a role in the regulation of Hct through MCV at least in vitro. Low T(a) (24°C) resulted in relatively large changes in pH with small changes in Hct and Osm in vitro with increased T(a) (42°C) conversely resulting in larger changes in both Hct and Osm. In vivo exposure to altered T(a) caused age-dependent changes in Hct, demonstrating a trend towards increased Hct at higher T(a). Further, exposing embryos to a gas mixture where P(CO2) = 5.1 kPa for >4 h period at T(a) of 37 or 42°C also did not elicit a change in Hct or Osm. Presumably, homeostatic mechanisms ensured that in vivo Hct was stable during a 4-6 h temperature and/or hypercapnic stress. Thus, although blood p

  9. The potential role of As-sumo-1 in the embryonic diapause process and early embryo development of Artemia sinica.

    Science.gov (United States)

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

    2014-01-01

    During embryonic development of Artemia sinica, environmental stresses induce the embryo diapause phenomenon, required to resist apoptosis and regulate cell cycle activity. The small ubiquitin-related modifier-1 (SUMO), a reversible post-translational protein modifier, plays an important role in embryo development