26 results on '"animal embryo"'
Search Results
2. Whole Mount In Situ Hybridization (WM-ISH) for miRNA Expression Profiling During Vertebrate Development
- Author
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Wang, Zhiguo, Yang, Baofeng, Wang, Zhiguo, and Yang, Baofeng
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- 2010
- Full Text
- View/download PDF
3. From the beginning: the basal transcription machinery and onset of transcription in the early animal embryo.
- Author
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Zurita, M., Reynaud, E., and Aguilar-Fuentes, J.
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- *
EMBRYOS , *EMBRYOLOGY , *ZYGOTES , *GENETIC transcription , *DEVELOPMENTAL biology - Abstract
Transcription onset in the early animal embryo is a fundamental process required for proper embryonic development. Depending on the species, transcription onset occurs at what specifically appears to be different developmental stages. However, studies in early embryos from different animal models have shown that components of the basal transcription machinery play fundamental and highly regulated roles at the onset of transcription. The state of the basal transcription machinery in the embryo seems to be equivalent in different organisms at transcription onset. The dynamic balance between putative activators and repressors as well as the chromatin/cytoplasmic ratio seem to be coordinated with basal transcription factors in order to activate zygotic transcription. Here we discuss and compare the regulation of the basal transcription machineries and their activation in early embryos of different model organisms. [ABSTRACT FROM AUTHOR]
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- 2008
- Full Text
- View/download PDF
4. The Freezing of Early Human Embryos and Blastocysts
- Author
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Ashwood-Smith, M. J., Simons, R., Feichtinger, Wilfried, editor, and Kemeter, Peter, editor
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- 1987
- Full Text
- View/download PDF
5. Comparing Intracellular Stability and Targeting of Sulfobetaine Quantum Dots with Other Surface Chemistries in Live Cells
- Author
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Muro, E., Fragola, A., Pons, T., Lequeux, N., Ioannou, Androulla, Skourides, Paris A., Dubertret, B., and Skourides, Paris A. [0000-0003-3502-5729]
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Cytoplasm ,Embryo, Nonmammalian ,Quantum Dot ,Nanoparticle ,Ligands ,Xenopus laevis ,chemistry.chemical_compound ,thioctic acid ,microinjection ,Semiconductor quantum dots ,animal ,betaine ,General Materials Science ,dihydrolipoate ,Thioctic Acid ,Chemistry ,specific staining ,Electroporation ,Pinocytosis ,article ,Living cell ,intracellular stability ,Biotinylation ,cytoplasm ,Stability ,HeLa cell ,living cells ,Intracellular ,Biotechnology ,electroporation ,Streptavidin ,Microinjections ,Cells ,quantum dots ,Nanotechnology ,Conjugated system ,chemistry ,animal embryo ,Biomaterials ,sulfobetaine ,Quantum Dots ,Animals ,Humans ,human ,technology, industry, and agriculture ,Proteins ,General Chemistry ,equipment and supplies ,Surface chemistry ,Betaine ,Quantum dot ,drug derivative ,Cytology ,metabolism ,HeLa Cells - Abstract
The in vivo labeling of intracellular components with quantum dots (QDs) is very limited because of QD aggregation in the cell cytoplasm and/or QD confinement into lysosomal compartments. In order to improve intracellular targeting with QDs, various surface chemistries and delivery methods have been explored, but they have not yet been compared systematically with respect to the QD intracellular stability. In this work, the intracellular aggregation kinetics of QDs for three different surface chemistries based on ligand exchange or encapsulation with amphiphilic polymers are compared. For each surface chemistry, three delivery methods for bringing the nanoparticles into the cells are compared: electroporation, microinjection, and pinocytosis. It is concluded that the QD intracellular aggregation behavior is strongly dependent on the surface chemistry. QDs coated with dihydrolipoic acid-sulfobetaine (DHLA-SB) ligands diffuse freely in cells for longer periods of time than for QDs in the other chemistries tested, and they can access all cytoplasmic compartments. Even when conjugated to streptavidin, these DHLA-SB QDs remain freely diffusing inside the cytoplasm and unaggregated, and they are able to reach a biotinylated target inside HeLa cells. Such labeling was more efficient when compared to commercial streptavidin-conjugated QDs, which may be due to the smaller size of DHLA-SB QDs and/or to their superior intracellular stability. Quantum dots (QDs) solubilized with the zwitterionic ligand dihydrolipoic acid-sulfobetaine present excellent in vivo stability when introduced into cultured cells and embryos. This intracellular stability is much higher than with other QD surface chemistries. When conjugated to streptavidin, these zwitterionic QDs can label an intracellular target with great specificity, higher than commercial streptavidin QDs. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. 8 1029 1037 Cited By :29
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- 2012
6. Down-Regulation of Circadian Clock GenePeriod 2in Uterine Endometrial Stromal Cells of Pregnant Rats During Decidualization
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Nobuhiko Yamauchi, Miho Uchikawa, Madoka Kawamura, and Masa-aki Hattori
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Physiology ,Circadian clock ,Uterus ,circadian rhythm signaling protein ,genetics ,rat ,endometrium ,transcription factor ,comparative study ,Cells, Cultured ,Decidua ,Decidualization ,article ,Per2 ,transgenic rat ,Cell Differentiation ,Period Circadian Proteins ,Implantation ,Circadian Rhythm ,PER2 ,female ,medicine.anatomical_structure ,stroma cell ,pregnancy ,Rats, Transgenic ,endocrine system ,medicine.medical_specialty ,Stromal cell ,nidation ,Down-Regulation ,Per2 protein, rat ,Rodentia ,Biology ,Vegf ,animal embryo ,Circadian Clocks ,Physiology (medical) ,Internal medicine ,medicine ,Animals ,Embryo Implantation ,Circadian rhythm ,Cell Proliferation ,cell culture ,uterus ,Rattus ,Embryo, Mammalian ,Rats ,Endocrinology ,breeding ,Pregnancy, Animal ,Stromal Cells ,metabolism ,Transcription Factors - Abstract
Circadian rhythms are modulated in a variety of peripheral tissues, including in the uterus where endometrial stromal cells (UESCs) undergo proliferation and differentiation (decidualization) during gestation. Here the authors focused on circadian rhythms in UESCs during implantation and decidualization in rodents. As revealed by analyses of cultured UESCs from pregnant Per2 promoter-dLuc transgenic rats, Per2 oscillation of ∼24 h was observed in response to dexamethasone. Per2 oscillation was enhanced in UESCs during implantation, whereas they were attenuated during decidualization. In vivo studies showed that PER2 protein in the uteri displayed a peak at zeitberger time 4 (ZT 4) (day 4.50 of gestation) and a trough at ZT 12 (day 4.83), indicating its circadian rhythmicity. Conversely, no significant circadian rhythm of the PER2 protein was observed during decidualization. Fluorescent immunohistochemical studies also supported circadian rhythmicity of the PER2 protein in its intracellular distribution. In accordance with Per2 mRNA expression, a circadian rhythm of vascular endothelial growth factor (Vegf) gene expression, having several E-box or E-box-like sites at the upstream of the transcription start site, was observed during implantation, showing a peak at ZT 0 and a trough at ZT 12. In contrast, Vegf mRNA expression displayed no circadian rhythm during decidualization. Collectively, the present results prove that Per2 oscillation is down-regulated in UESCs during decidualization. It is strongly suggested that cellular differentiation in UESCs interferes with circadian clockwork.
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- 2010
7. A serum-free and defined medium for the culture of mammalian postimplantation embryos
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Drakou, Katerina, Georgiades, Pantelis, and Georgiades, Pantelis [0000-0002-5538-3163]
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Developmental toxicity ,Biochemistry ,Culture Media, Serum-Free ,Embryo Culture Techniques ,Mice ,animal ,Defined medium ,Mice, Inbred ICR ,Primitive streak ,catalase ,Embryo ,N2B27 ,superoxide dismutase ,Cell biology ,embryo culture ,priority journal ,histochemistry ,mammalian embryo ,Institute for Cancer Research mouse ,serum-free medium ,insulin ,culture medium ,Biophysics ,embryo ,Embryonic Development ,Biology ,chemistry ,Article ,animal embryo ,bovine serum albumin ,Animals ,primitive streak ,transferrin ,procedures ,Molecular Biology ,mouse ,nonhuman ,Mouse whole embryo culture ,Embryogenesis ,Gastrulation ,Embryo culture ,embryo development ,Cell Biology ,Embryo, Mammalian ,Embryonic stem cell ,Chemically defined medium ,Immunology ,physiology ,cytology ,lectin ,in situ hybridization ,metabolism ,Serum-free - Abstract
Whole embryo culture (WEC) of postimplantation rodent embryos is widely used for the study of mammalian embryogenesis and developmental toxicity testing. Its major advantage is that it allows direct access to embryos for experimental manipulations and the monitoring of their consequences that would otherwise not be possible or technically difficult to perform in utero. However, a major drawback of mammalian WEC is that the culture media currently in use display batch variations and are undefined, as they contain serum or serum replacements of unknown composition. Moreover, these media possess cell-signalling activities important for embryogenesis. Therefore, reproducibility of mammalian postimplantation WEC results may be affected by batch variation and their interpretation is complicated because the experimenter is unsure whether the embryo response to experimental perturbations is solely due to their action, or modified as a result of influences from undefined substances/signaling activities present in culture media. To alleviate these problems we investigated whether N2B27, a serum-free and defined medium, can support the in vitro development of postimplantation mammalian embryos. We show that N2B27 allows pre-gastrulation mouse embryos isolated at embryonic day 5.5 to develop to advanced gastrulation, reaching the mid- and late primitive streak stages. This is the first demonstration that postimplantation mammalian embryos can develop in vitro in a defined medium in the absence of serum and provides a novel WEC system for studying developmental mechanisms and testing for developmental toxicity during the early postimplantation period. © 2015 Elsevier Inc. All rights reserved. 468 813 819
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- 2015
8. Detecting brain injury in neonatal hypoxic ischemic encephalopathy: closing the gap between experimental and clinical research.
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Aridas J.D., Ditchfield M., Wong F.Y., Miller S.L., Sutherland A.E., Nitsos I., Yawno T., Fahey M.C., Malhotra A., Wallace E.M., Jenkin G., Aridas J.D., Ditchfield M., Wong F.Y., Miller S.L., Sutherland A.E., Nitsos I., Yawno T., Fahey M.C., Malhotra A., Wallace E.M., and Jenkin G.
- Abstract
Moderate to severe neonatal hypoxic ischemic encephalopathy remains an important cause of infant death and childhood disability. Early and accurate diagnosis of encephalopathy is difficult but critical for timely intervention. Thus, we have utilized a clinically relevant large animal model of asphyxia in-utero, followed by immediate lamb delivery, resuscitation and clinical care over the next 72h for assessment of potential biomarkers of brain injury. In-utero asphyxia was induced in twelve near-term lambs and outcomes compared with seven controls. Asphyxia resulted in bradycardia (97+/-12beats/min), hypotension (12.1+/-1mm Hg) and metabolic acidosis (pH6.9+/-0.02; base-excess -13.8+/-0.8mmol/l). 72h following asphyxia, cerebrospinal concentrations of malondialdehyde and S100B were elevated 2-fold and 5-fold, respectively, in asphyxic lambs compared to control lambs. Magnetic resonance spectroscopy (MRS) at 72h showed a significant decrease in n-acetyl aspartate: choline ratio in asphyxia lambs compared to that observed at 12h (0.56+/-0.23 vs. 0.82+/-0.15, respectively); lactate:choline ratio was not changed over this time. Marked neuropathology was observed in asphyxia lambs with neuronal degeneration in the hippocampus, thalamus, striatum and cortex. Astrogliosis was observed in the hippocampus and thalamus. Early blood markers of metabolic state showed limited predictive value of histological damage at 72h. MRS outcomes at 72h showed a modest but significant correlation with histological evidence of neuronal brain injury (lactate:N-acetyl aspartate ratio in the thalamus r(2)=0.2, p<0.01). MRS at 72h was best able to detect established brain injury, but a combination of biomarkers over multiple phases of injury may be able to assess the evolution of neonatal brain injury. Copyright © 2014 Elsevier Inc. All rights reserved.
- Published
- 2015
9. A model to predict the thermal reaction norm for the embryo growth rate from field data
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Yakup Kaska and Marc Girondot
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temperature sensitivity ,Embryo, Nonmammalian ,incubation temperature ,sex determination ,Growth ,adaptation ,Biochemistry ,incubation time ,Turkey (republic) ,Body Temperature ,heat sensitivity ,Nest ,hatching ,animal ,Growth rate ,Incubation ,Caretta caretta ,Ecology ,embryo growth ,Norm of reaction ,Temperature ,Adaptation, Physiological ,Turtles ,female ,growth rate ,egg laying ,General Agricultural and Biological Sciences ,nesting ,phenotype ,Zoology ,embryo ,Reptile ,Biology ,Models, Biological ,Article ,animal embryo ,Incubation period ,process model ,embryology ,Animals ,controlled study ,sex determination process ,Hatchling ,Egg incubation ,nonhuman ,Hatching ,embryo development ,prediction ,turtle ,biological model ,Reaction norm ,validation process ,time series analysis ,physiology ,Developmental Biology - Abstract
The incubation of eggs is strongly influenced by temperature as observed in all species studied to date. For example, incubation duration, sexual phenotype, growth, and performances in many vertebrate hatchlings are affected by incubation temperature. Yet it is very difficult to predict temperature effect based on the temperature within a field nest, as temperature varies throughout incubation. Previous works used egg incubation at constant temperatures in the laboratory to evaluate the dependency of growtProd. Type: FTPh rate on temperature. However, generating such data is time consuming and not always feasible due to logistical and legislative constraints. This paper therefore presents a methodology to extract the thermal reaction norm for the embryo growth rate directly from a time series of incubation temperatures recorded within natural nests. This methodology was successfully applied to the nests of the marine turtle Caretta caretta incubated on Dalyan Beach in Turkey, although it can also be used for any egg-laying species, with some of its limitations being discussed in the paper. Knowledge about embryo growth patterns is also important when determining the thermosensitive period for species with temperature-dependent sex determination. Indeed, in this case, sexual phenotype is sensitive to temperature only during this window of embryonic development. © 2014 Elsevier Ltd.
- Published
- 2014
10. Adult human stem cells incorporated into pig embryos grew into precursors of heart, neural, and other types of specific tissue, according to researchers at the Salk Institute
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Animal embryo ,Bioethics ,Human embryo ,Stem cell research -- Ethical aspects ,Political science - Abstract
Adult human stem cells incorporated into pig embryos grew into precursors of heart, neural, and other types of specific tissue, according to researchers at the Salk Institute. Headlines about pig-human [...]
- Published
- 2017
11. Canonical WNT signaling regulates development of bovine embryos to the blastocyst stage
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Anna C. Denicol, Silvia F. Carambula, Peter J. Hansen, Kanyon M. McLean, Kyle B. Dobbs, B. Loureiro, University of Florida, and Universidade Estadual Paulista (Unesp)
- Subjects
2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine ,pyrimidine derivative ,Inner cell mass ,animal ,genetics ,Wnt Signaling Pathway ,Regulation of gene expression ,Multidisciplinary ,drug effect ,artificial insemination ,Wnt signaling pathway ,Gene Expression Regulation, Developmental ,1,3 benzodioxole derivative ,LRP5 ,Embryo ,gene expression regulation ,Cell biology ,female ,medicine.anatomical_structure ,embryonic structures ,Intercellular Signaling Peptides and Proteins ,Female ,2 amino 4 (3,4 (methylenedioxy)benzylamino) 6 (3 methoxyphenyl)pyrimidine ,Embryonic Development ,Biology ,chemistry ,Article ,animal embryo ,Insemination ,medicine ,Animals ,signal peptide ,Benzodioxoles ,Blastocyst ,drug potentiation ,blastocyst ,Embryogenesis ,embryo development ,Embryo, Mammalian ,Wnt protein ,Molecular biology ,Wnt Proteins ,Pyrimidines ,DKK1 ,cattle ,cytology ,Cattle ,metabolism - Abstract
Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:28:35Z No. of bitstreams: 0 Made available in DSpace on 2014-05-27T11:28:35Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-03-01 Objectives were to evaluate the role of canonical WNT signaling in development of the preimplantation embryo. Signaling was activated with 2-Amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (AMBMP) and inhibited with Dickkopf-related protein 1 (DKK1). Treatment of bovine embryos with AMBMP at day 5 after insemination decreased development to the blastocyst stage at day 7 and reduced numbers of trophectoderm and inner cell mass cells. At high concentrations, AMBMP caused disorganization of the inner cell mass. DKK1 blocked actions of AMBMP but did not affect development in the absence of AMBMP. Examination of gene expression in day 6 morulae by microarray revealed expression of 16 WNT genes and other genes involved in WNT signaling; differences in relative expression were confirmed by PCR for 7 genes. In conclusion, the preimplantation embryo possesses a functional WNT signaling system and activation of the canonical pathway can inhibit embryonic development. Department of Animal Sciences Genetics Institute University of Florida, Gainesville, FL 32611-0910 Departamento de Farmacologia Instituto de Biociências Universidade Estadual Paulista (UNESP), Botucatu - SP Departamento de Farmacologia Instituto de Biociências Universidade Estadual Paulista (UNESP), Botucatu - SP
- Published
- 2013
- Full Text
- View/download PDF
12. Dimerized glycosaminoglycan chains increase FGF signaling during zebrafish development
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Duraikkannu Loganathan, Lena Kjellén, Mamoru Koketsu, Akinori Kojima, Thao Kim Nu Nguyen, Chi Bin Chien, Venkataswamy Sorna, Richard I. Dorsky, Inger Eriksson, Vy M. Tran, and Balagurunathan Kuberan
- Subjects
Embryo, Nonmammalian ,Morpholino ,xylosides ,Fibroblast growth factor ,Biochemistry ,3 [4 methyl 2 (2 oxo 3 indolinylmethylidenyl) 3 pyrrolyl]propionic acid ,Animals, Genetically Modified ,zebra fish ,animal ,genetics ,Glycosides ,In Situ Hybridization ,Zebrafish ,Glycosaminoglycans ,Regulation of gene expression ,dimerization ,biology ,protein kinase inhibitor ,Gene Expression Regulation, Developmental ,General Medicine ,gene expression regulation ,priority journal ,Fibroblast growth factor receptor ,embryonic structures ,fibroblast growth factor receptor ,Molecular Medicine ,Signal transduction ,signal transduction ,glycoside ,Cell signaling ,animal structures ,Molecular Sequence Data ,embryo ,Tyrosine Kinase Inhibitor SU5402 ,chemistry ,animal embryo ,Wnt signaling pathway ,fibroblast growth factor ,glycosaminoglycan ,embryology ,Animals ,controlled study ,Pyrroles ,development ,Protein Kinase Inhibitors ,antagonists and inhibitors ,nonhuman ,Base Sequence ,nucleotide sequence ,Fgf8 protein, zebrafish ,zebrafish protein ,Zebrafish Proteins ,Molecular biology ,Receptors, Fibroblast Growth Factor ,transgenic animal ,Fibroblast Growth Factors ,Proteoglycan ,scaffold protein ,drug effects ,molecular genetics ,biology.protein ,Syndecan-1 ,pyrrole derivative ,syndecan 1 ,metabolism - Abstract
Proteoglycans (PGs) modulate numerous signaling pathways during development through binding of their glycosaminoglycan (GAG) side chains to various signaling molecules, including fibroblast growth factors (FGFs). A majority of PGs possess two or more GAG side chains, suggesting that GAG multivalency is imperative for biological functions in vivo. However, only a few studies have examined the biological significance of GAG multivalency. In this report, we utilized a library of bis- and tris-xylosides that produce two and three GAG chains on the same scaffold, respectively, thus mimicking PGs, to examine the importance of GAG valency and chain type in regulating FGF/FGFR interactions in vivo in zebrafish. A number of bis- and tris-xylosides, but not mono-xylosides, caused an elongation phenotype upon their injection into embryos. In situ hybridization showed that elongated embryos have elevated expression of the FGF target gene mkp3 but unchanged expression of reporters for other pathways, indicating that FGF/FGFR signaling was specifically hyperactivated. In support of this observation, elongation can be reversed by the tyrosine kinase inhibitor SU5402, mRNA for the FGFR antagonist sprouty4, or FGF8 morpholino. Endogenous GAGs seem to be unaffected after xyloside treatment, suggesting that this is a gain-of-function phenotype. Furthermore, expression of a multivalent but not a monovalent GAG containing syndecan-1 proteoglycan recapitulates the elongation phenotype observed with the bivalent xylosides. On the basis of these in vivo findings, we propose a new model for GAG/FGF/FGFR interactions in which dimerized GAG chains can activate FGF-mediated signal transduction pathways. � 2013 American Chemical Society.
- Published
- 2013
13. Activation of endogenous FAK via expression of its amino terminal domain in Xenopus embryos
- Author
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Petridou, Nicoletta I., Stylianou, Panayiota, Christodoulou, Neophytos, Rhoads, Daniel S., Guan, Junlin, Skourides, Paris A., and Skourides, Paris A. [0000-0003-3502-5729]
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Integrins ,Embryo, Nonmammalian ,Xenopus ,Gene Expression ,lcsh:Medicine ,animal cell ,Mesoderm ,Mice ,Xenopus laevis ,Molecular Cell Biology ,Morphogenesis ,Signaling in Cellular Processes ,enzyme phosphorylation ,Phosphorylation ,Tyrosine ,lcsh:Science ,Genes, Dominant ,Multidisciplinary ,FERM domain ,biology ,Mus ,article ,protein domain ,Animal Models ,Protein-Tyrosine Kinases ,Cadherins ,blastula ,Extracellular Matrix ,Cell biology ,Protein Transport ,src-Family Kinases ,Cell Movement Signaling ,Tyrosine kinase ,signal transduction ,Research Article ,Signal Transduction ,enzyme localization ,integrin ,PTK2 ,embryo ,morphogenesis ,Cell Migration ,Extracellular Matrix Signaling ,animal embryo ,Cell Line ,in vivo study ,Focal adhesion ,Structure-Activity Relationship ,Model Organisms ,Cell Adhesion ,Animals ,Cell adhesion ,protein expression ,Biology ,carboxy terminal sequence ,nonhuman ,focal adhesion kinase ,Cell Membrane ,lcsh:R ,enzyme activation ,biology.organism_classification ,Protein Structure, Tertiary ,Enzyme Activation ,developmental stage ,Focal Adhesion Protein-Tyrosine Kinases ,amino terminal sequence ,lcsh:Q ,tyrosine ,Developmental Biology - Abstract
Background: The Focal Adhesion Kinase is a well studied tyrosine kinase involved in a wide number of cellular processes including cell adhesion and migration. It has also been shown to play important roles during embryonic development and targeted disruption of the FAK gene in mice results in embryonic lethality by day 8.5. Principal Findings: Here we examined the pattern of phosphorylation of FAK during Xenopus development and found that FAK is phosphorylated on all major tyrosine residues examined from early blastula stages well before any morphogenetic movements take place. We go on to show that FRNK fails to act as a dominant negative in the context of the early embryo and that the FERM domain has a major role in determining FAK's localization at the plasma membrane. Finally, we show that autonomous expression of the FERM domain leads to the activation of endogenous FAK in a tyrosine 397 dependent fashion. Conclusions: Overall, our data suggest an important role for the FERM domain in the activation of FAK and indicate that integrin signalling plays a limited role in the in vivo activation of FAK at least during the early stages of development. © 2012 Petridou et al. 7 Cited By :6
- Published
- 2012
14. Effects of heat stress on development, quality and survival of Bos indicus and Bos taurus embryos produced in vitro
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Anthony C. S. Castilho, Raquel Zaneti Puelker, C. F. Silva, J. S. Ticianelli, Eduardo M. Razza, H. P. Eduardo, Ciro Moraes Barros, R. A. Satrapa, E.S. Sartorelli, B. Loureiro, Universidade Estadual Paulista (Unesp), and Progest Ltda.
- Subjects
Hot Temperature ,Gene Expression ,Apoptosis ,Heat stress ,Human fertilization ,Food Animals ,animal ,Small Animals ,comparative study ,Genetics ,biology ,Embryo ,Bovine ,Breed ,Embryo transfer ,medicine.anatomical_structure ,female ,embryonic structures ,Female ,animal structures ,Embryonic Development ,Bos indicus ,prenatal development ,Fertilization in Vitro ,animal embryo ,Bovinae ,Andrology ,Species Specificity ,medicine ,Animals ,Blastocyst ,oocyte ,embryo transfer ,Equine ,blastocyst ,Embryogenesis ,Embryos ,species difference ,embryo development ,Oocyte ,biology.organism_classification ,Embryo Transfer ,Embryo, Mammalian ,Bos taurus ,cattle ,physiology ,Oocytes ,Animal Science and Zoology ,Cattle ,heat ,animal disease - Abstract
Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:28:10Z No. of bitstreams: 0 Made available in DSpace on 2014-05-27T11:28:10Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-01-15 Heat stress is an important cause of poor development and low survival rates in bovine embryos. Experiments were conducted to test the hypothesis that Bos indicus embryos are more resistant to heat stress than are Bos taurus embryos. In experiment 1, Nelore and Jersey embryos from oocyte pick-up-derived oocytes were submitted to heat stress (96 hours post-insemination, 41 °C, 6 hours), developmental ratios were assessed at Day 7 (Day 0 = day of fertilization), and blastocysts were frozen for RNA extraction. Experiment 2 evaluated expression of COX2, CDX2, HSF1, and PLAC8 in previously frozen blastocysts. In experiment 3, Nellore and Angus embryos from oocyte pick-up-derived oocytes were submitted to heat stress (96 hours post-insemination, 41 °C, 12 hours) and transferred to recipients on Day 7. In experiment 4, embryos developed as in experiment 3 were fixed for Terminal deoxynucleotidyl transferase dUTP nick end labeling labeling and total cell counting. In experiment 1, heat stress decreased the percentage of Jersey oocytes that became blastocysts, but had no effect on Nellore embryos (34.6%, 25.0%, 39.5%, and 33.0% for Jersey control, Jersey heat-stressed, Nellore control, and Nellore heat-stressed oocytes, respectively; P < 0.05). In experiment 2, heat stress decreased (P < 0.05) expression of CDX2 and PLAC8, with higher expression of these genes in Nellore embryos than in Jersey embryos. Heat stress also decreased (P < 0.05) expression of COX2 in Jersey embryos, but had no effect on Nellore embryos. Expression of HSF1 was decreased (P < 0.05) by heat stress in both breeds, with a greater effect in Nellore embryos. In experiment 3, heat stress tended (P = 0.1) to decrease the percentage of pregnancies among cows (Day 30 to 35) that received Angus embryos. In experiment 4, heat stress increased (P < 0.05) the percentage of apoptotic blastomeres, but had no breed-specific effects. In addition, Nellore embryos had fewer (P < 0.05) Terminal deoxynucleotidyl transferase dUTP nick end labeling- positive blastomeres than did Angus embryos. We concluded that the detrimental effects of heat stress were dependent upon embryo breed and were more evident in Bos taurus embryos than in Bos indicus embryos. © 2013 Elsevier Inc. Department of Pharmacology Institute of Bioscience University of São Paulo State (UNESP), Botucatu, São Paulo Progest Ltda., Botucatu, São Paulo Department of Pharmacology Institute of Bioscience University of São Paulo State (UNESP), Botucatu, São Paulo
- Published
- 2011
15. Prohormone Processing in Zebrafish
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Michael G. Morash, Kelly Soanes, and Younes Anini
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Proteases ,Embryo, Nonmammalian ,animal structures ,enzymology ,Proteolysis ,tissue distribution ,Prohormone ,DNA sequence ,protein precursor ,morphogenesis ,Biology ,chemistry ,animal embryo ,Animals, Genetically Modified ,gene silencing ,serine proteinase ,Alzheimer Disease ,Neoplasms ,zebra fish ,medicine ,genetics ,RNA, Messenger ,Protein Precursors ,Receptor ,Zebrafish ,Gene knockdown ,Danio rerio ,medicine.diagnostic_test ,messenger RNA ,Proteolytic enzymes ,methodology ,zebrafish protein ,Sequence Analysis, DNA ,Zebrafish Proteins ,biology.organism_classification ,infection ,humanities ,transgenic animal ,Cell biology ,Gene Knockdown Techniques ,Proprotein Convertases ,metabolism ,neoplasm ,medicine.drug - Abstract
Proprotein convertases (PCs) are secretory proteolytic enzymes that activate precursor proteins into biologically active forms by limited proteolysis at one or multiple internal sites. PCs are implicated in the processing of multiple protein precursors, including hormones, proteases, growth factors, angiogenic factors, and receptors. PCs have been linked recently to various pathologies such as Alzheimer's disease, tumorigenesis, and infections. The zebrafish has emerged as an attractive model for studying the role of PCs not only in substrate production but also in development. Herein we describe methods that are used to characterize DNA sequences of PCs in zebrafish, as well as to evaluate the ontogeny and tissue distribution of their transcripts. We also provide information on the morpholino-mediated knockdown of proprotein convertases. © 2011 Springer Science+Business Media, LLC.
- Published
- 2011
16. Parthenogenic blastocysts cultured under in vivo conditions exhibit proliferation and differentiation expression genes similar to those of normal embryos
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Maria Desemparats Saenz-de-Juano, C. Naturil-Alfonso, David S. Peñaranda, Jose S. Vicente, and Francisco Marco-Jiménez
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Physiology ,Cellular differentiation ,Parthenogenesis ,Rabbit ,PRODUCCION ANIMAL ,Embryo development ,Animal embryo ,Embryo Culture Techniques ,Endocrinology ,Food Animals ,Cell differentiation ,Nidation ,Evaluation ,Cell proliferation ,Cells, Cultured ,Embryo ,General Medicine ,Gene expression profiling ,medicine.anatomical_structure ,BIOLOGIA ANIMAL ,Female ,Rabbits ,Stem cell ,Culture conditions ,Embryonic Development ,Biology ,Article ,Andrology ,In vitro ,In vivo ,medicine ,Genetics ,Animals ,Embryo culture ,Blastocyst ,Embryo Implantation ,Animal ,MRNA expression ,Embryogenesis ,Embryo, Mammalian ,Molecular biology ,Metabolism ,Cell culture ,Animal Science and Zoology ,Comparative study ,Gene expression ,Cytology ,Transforming growth factor - Abstract
Parthenote embryos offer multiple possibilities in biotechnological investigation, such as stem cell research. However, there is still a dearth of knowledge of this kind of embryo. In this study, development and ploidy were analysed in parthenotes under in vitro and in vivo culture conditions. Subsequently, using real-time PCR, the expressions of factor OCT-4, Vascular Endothelial Growth Factor, Epidermal Growth Factor Receptor 3 and Transforming Growth Factor ß2 genes were analysed to compare the embryo types at the blastocyst stage. Development and implantation of parthenote embryos were described after transfer at day 10 of pregnancy. Parthenotes showed similar blastocyst development for both culture conditions and most of the parthenotes produced were diploid. However, parthenotes developed under in vivo conditions showed similar mRNA expression of OCT-4, VEGF and TGF-ß2 to 5 and 6 day old blastocysts. In contrast, parthenotes developed under in vitro conditions had altered the expression pattern of these genes, except for erbB3 mRNA. Finally, transferred parthenotes had the ability to implant but showed severe growth retardation and lesser size. This is the first demonstration of the influence of culture conditions on parthenote mRNA expression. Our study highlights the importance of culture conditions in subsequent uses of parthenotes, such as the production of stem cell lines. © 2011 Elsevier B.V., This work was supported by Generalitat Valenciana research program (Prometeo 2009-8260: 125) and by the Spanish Research Projects (CICYT AGL2008-03274). The authors thank Neil Macowan Language Services for revising the English version of the manuscript.
- Published
- 2011
17. Imprinted gene expression in in vivo- And in vi/ro-produced bovine embryos and chorio-allantoic membranes
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Felipe Perecin, Flávio Vieira Meirelles, Giovana Krempel Fonseca Merighe, Christina R. Ferreira, Joaquim Mansano Garcia, W. Yamazaki, S. C. Méo, Universidade Estadual Paulista (Unesp), Universidade de São Paulo (USP), and Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA)
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Male ,Nuclear Transfer Techniques ,Genomic imprinting ,Unclassified drug ,Somatic cell ,Parthenogenesis ,Gene Expression ,Animal tissue ,Chorioallantoic Membrane ,Receptor, IGF Type 2 ,Animal embryo ,In vivo study ,Gene expression ,Gene activation ,Bos ,Somatomedin a receptor ,Chorioallantois ,Regulation of gene expression ,Growth factor receptor ,Genetic transcription ,Messenger RNA ,IGF2 ,Embryo ,General Medicine ,embryonic structures ,Somatic cell nuclear transfer ,Epigenetics ,Female ,Animal cell ,Nuclear transfer ,Glyceraldehyde 3 phosphate dehydrogenase ,animal structures ,Genome imprinting ,Fertilization in Vitro ,Biology ,Bovinae ,Andrology ,Genomic Imprinting ,Somatomedin A ,Somatomedin B ,In vivo ,Insulin-Like Growth Factor II ,Genetics ,Animals ,Animal experiment ,Molecular Biology ,Cloning ,Cell nucleus transplantation ,Animal ,Cow ,In vitro study ,Nonhuman ,Embryo, Mammalian ,Molecular biology ,Gene frequency ,F2 ,Metabolism ,Cattle ,Somatomedin B receptor ,Controlled study ,Nucleotide sequence - Abstract
Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:23:55Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:41:00Z : No. of bitstreams: 1 2-s2.0-66349090503.pdf: 575567 bytes, checksum: 75f313bd58f9a4d47c3c947901c00564 (MD5) Made available in DSpace on 2014-05-27T11:23:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-06-15 Item merged in doublecheck by Felipe Arakaki (arakaki@reitoria.unesp.br) on 2015-12-14T16:11:12Z Item was identical to item(s): 2502, 70521 at handle(s): http://hdl.handle.net/11449/2604, http://hdl.handle.net/11449/71039 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Cloning by nuclear transfer is often associated with poor results due to abnormal nuclear reprogramming of somatic donor cells and altered gene expression patterns. We investigated the expression patterns of imprinted genes IGF2 and IGF2R in 33- to 36-day bovine embryos and chorio-allantoic membranes derived from in vivo- and in vitro-produced embryos by somatic cell nuclear transfer (SCNT), parthenogenetic activation, and in vitro fertilization (IVF). There was a lower IGF2 expression rate in the SCNT (0.19) and parthenogenetic (0.02) groups when compared to in vivo and IVF embryos (2.01; P
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- 2009
18. Multiple Embryo Time-Lapse Imaging of Zebrafish Development
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Andrew C. Oates, Lola Bajard, Leah Herrgen, and Christian Schröter
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Time Factors ,Population ,embryo ,Growth ,animal embryo ,Article ,Time ,histology ,03 medical and health sciences ,Computer-Assisted ,0302 clinical medicine ,Image processing ,Developmental biology ,zebra fish ,Animals ,animal ,Time-Lapse Imaging ,education ,Zebrafish ,030304 developmental biology ,0303 health sciences ,education.field_of_study ,Nonmammalian ,biology ,development and aging ,Methodology ,temperature ,embryo development ,Embryo ,Anatomy ,zebrafish ,biology.organism_classification ,Temporal resolution ,Embryonic development ,Biological system ,030217 neurology & neurosurgery - Abstract
Understanding the dynamics of developmental and cellular processes requires documentation of their changes with appropriate temporal and spatial resolution. Furthermore, simultaneous recording from a population of embryos under identical conditions allows statistical estimates of precision and variability to be made. This chapter describes a protocol for time-lapse microscopy of multiple embryos in parallel developing under tightly controlled conditions. This method is currently best suited to follow tissue-scale morphogenetic movements with temporal resolution in the minute range, for hours or even days. Applications of the method include the comparison of the dynamics of a process of interest between groups of wild-type embryos and their mutant siblings or between embryos treated with different chemical compounds. Temperature control allows for the investigation of the temperature dependence of a process of interest.
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- 2009
19. Parthenogenic blastocysts cultured under in vivo conditions exhibit proliferation and differentiation expression genes similar to those of normal embryos
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Universitat Politècnica de València. Instituto de Ciencia y Tecnología Animal - Institut de Ciència i Tecnologia Animal, Universitat Politècnica de València. Departamento de Ciencia Animal - Departament de Ciència Animal, Ministerio de Ciencia e Innovación, Generalitat Valenciana, Naturil Alfonso, Carmen, Saenz de Juano Ribes, María de los Desamparados, Peñaranda, D.S., Vicente Antón, José Salvador, Marco Jiménez, Francisco, Universitat Politècnica de València. Instituto de Ciencia y Tecnología Animal - Institut de Ciència i Tecnologia Animal, Universitat Politècnica de València. Departamento de Ciencia Animal - Departament de Ciència Animal, Ministerio de Ciencia e Innovación, Generalitat Valenciana, Naturil Alfonso, Carmen, Saenz de Juano Ribes, María de los Desamparados, Peñaranda, D.S., Vicente Antón, José Salvador, and Marco Jiménez, Francisco
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Parthenote embryos offer multiple possibilities in biotechnological investigation, such as stem cell research. However, there is still a dearth of knowledge of this kind of embryo. In this study, development and ploidy were analysed in parthenotes under in vitro and in vivo culture conditions. Subsequently, using real-time PCR, the expressions of factor OCT-4, Vascular Endothelial Growth Factor, Epidermal Growth Factor Receptor 3 and Transforming Growth Factor ß2 genes were analysed to compare the embryo types at the blastocyst stage. Development and implantation of parthenote embryos were described after transfer at day 10 of pregnancy. Parthenotes showed similar blastocyst development for both culture conditions and most of the parthenotes produced were diploid. However, parthenotes developed under in vivo conditions showed similar mRNA expression of OCT-4, VEGF and TGF-ß2 to 5 and 6 day old blastocysts. In contrast, parthenotes developed under in vitro conditions had altered the expression pattern of these genes, except for erbB3 mRNA. Finally, transferred parthenotes had the ability to implant but showed severe growth retardation and lesser size. This is the first demonstration of the influence of culture conditions on parthenote mRNA expression. Our study highlights the importance of culture conditions in subsequent uses of parthenotes, such as the production of stem cell lines. © 2011 Elsevier B.V.
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- 2011
20. Wnt3a deficiency irreversibly impairs hematopoietic stem cell self-renewal and leads to defects in progenitor cell differentiation
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Luis, T.C. (Tiago), Weerkamp, F. (Floor), Naber, B.A. (Brigitta), Baert, M.R.M. (Miranda), Haas, E.F. (Edwin) de, Nikolic, T. (Tatjana), Heuvelmans, S. (Sjanneke), Krijger, R.R. (Ronald) de, Dongen, J.J.M. (Jacques) van, Staal, F.J.T. (Frank), Luis, T.C. (Tiago), Weerkamp, F. (Floor), Naber, B.A. (Brigitta), Baert, M.R.M. (Miranda), Haas, E.F. (Edwin) de, Nikolic, T. (Tatjana), Heuvelmans, S. (Sjanneke), Krijger, R.R. (Ronald) de, Dongen, J.J.M. (Jacques) van, and Staal, F.J.T. (Frank)
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Canonical Wnt signaling has been implicated in various aspects of hematopoiesis. Its role is controversial due to different outcomes between various inducible Wnt-signaling loss-of-function models and also compared with gain-of-function systems. We therefore studied a mouse deficient for a Wnt gene that seemed to play a nonredundant role in hematopoiesis. Mice lacking Wnt3a die prenatally around embryonic day (E) 12.5, allowing fetal hematopoiesis to be studied using in vitro assays and transplantation into irradiated recipient mice. Here we show that Wnt3a deficiency leads to a reduction in the numbers of hematopoietic stem cells (HSCs) and progenitor cells in the fetal liver (FL) and to severely reduced reconstitution capacity as measured in secondary transplantation assays. This deficiency is irreversible and cannot be restored by transplantation into Wnt3a competent mice. The impaired long-term repopulation capacity of Wnt3a-/- HSCs could not be explained by altered cell cycle or survival of primitive progenitors. Moreover, Wnt3a deficiency affected myeloid but not B-lymphoid development at the progenitor level, and affected immature thymocyte differentiation. Our results show that Wnt3a signaling not only provides proliferative stimuli, such as for immature thymocytes, but also regulates cell fate decisions of HSC during hematopoiesis.
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- 2009
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21. Mutations in SLC29A3, Encoding an Equilibrative Nucleoside Transporter ENT3, Cause a Familial Histiocytosis Syndrome (Faisalabad Histiocytosis) and Familial Rosai-Dorfman Disease
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Dean Gentle, Hans-Christoph Rossbach, Carlos Dalence, Richard C. Trembath, Anna Straatman-Iwanowska, Nicholas J. Davies, Hakan Cangul, Eamonn R. Maher, Serdar Ceylaner, Peter Devilee, Paul Gissen, Mark R. Morris, Fatimah Rahman, Maaike P.G. Vreeswijk, David Tannahill, Margaret A. Knowles, Erol Kismet, Diane Gleeson, Vedat Koseoglu, Stephen Keenan, Neil V. Morgan, Shanaz Pasha, Uludağ Üniversitesi/Tıp Fakültesi/Tıbbi Genetik Anabilim Dalı., and Cangül, Hakan
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Hypertrichosis ,Pathology ,Adenosine ,Mouse ,Lymphadenopathy ,Apoptosis ,Pathogenesis ,Faisalabad histiocytosis ,Embryo development ,Gene ,Animal embryo ,Chromosome 10 ,Mice ,Consanguinity ,Breast cancer ,0302 clinical medicine ,Cell proliferation ,Gene expression regulation ,0303 health sciences ,Physical chromosome mapping ,RNA, small interfering ,3. Good health ,Histiocytosis ,030220 oncology & carcinogenesis ,HeLa cell ,Human ,Protein slc29a3 ,medicine.medical_specialty ,Embryo, mammalian ,Cells ,Genetic loci ,Urinary bladder neoplasms ,Clinical article ,SLC29A3 protein ,Article ,03 medical and health sciences ,Genetics ,Humans ,Cancer cell culture ,Family ,Chromosome 10q ,Molecular Biology ,Alleles ,Gene mapping ,Ecology, Evolution, Behavior and Systematics ,Histiocyte ,Animal ,Chromosomes, human, pair 10 ,Tumor cell line ,ENT3 protein ,medicine.disease ,Rosai Dorfman disease ,Cell line, tumor ,Human cell ,Chromosome map ,Mutation ,Cell strain HEK293 ,Gene expression ,Breast neoplasms ,Nucleotide sequence ,Cancer Research ,Candidate gene ,Unclassified drug ,Growth ,Gene locus ,Colony-forming units assay ,Animal tissue ,DNA mutational analysis ,adenosine induces apoptosis sensorineural deafness cells lymphadenopathy activation brothers pathway growth cancer hent3 ,Histiocytosis, sinus ,Molecular genetics ,Genetics and Genomics/Genetics of Disease ,Genetics (clinical) ,Rosai–Dorfman disease ,Allele ,Genetics & heredity ,Bladder cancer ,Syndrome ,Small interfering RNA ,Phenotype ,Embryo ,Histiocytoses ,Genetics and Genomics/Gene Discovery ,Female ,Research Article ,lcsh:QH426-470 ,Breast tumor ,ADENOSINE INDUCES APOPTOSIS, SENSORINEURAL DEAFNESS, CELLS, LYMPHADENOPATHY, ACTIVATION, BROTHERS, PATHWAY, GROWTH, CANCER, HENT3 ,Nucleoside transporter ,Biology ,Germline mutation ,Molecular sequence data ,Bladder tumor ,medicine ,Animals ,Gene mutation ,Sinus Histiocytosis ,Emperipolesis ,Sinus histiocytosis ,030304 developmental biology ,Equilibrative nucleoside transporter 3 ,Nucleoside transport proteins ,Clonogenic assay ,Sinus Histiocytosis with Massive Lymphadenopathy ,Carrier proteins and binding proteins ,Mutational analysis ,Nonhuman ,Base sequence ,Autosomal recessive inheritance ,lcsh:Genetics ,Metabolism ,Clinical feature ,Protein protein interaction ,Equilibrative nucleoside transporter ,Controlled study - Abstract
The histiocytoses are a heterogeneous group of disorders characterised by an excessive number of histiocytes. In most cases the pathophysiology is unclear and treatment is nonspecific. Faisalabad histiocytosis (FHC) (MIM 602782) has been classed as an autosomal recessively inherited form of histiocytosis with similarities to Rosai-Dorfman disease (RDD) (also known as sinus histiocytosis with massive lymphadenopathy (SHML)). To elucidate the molecular basis of FHC, we performed autozygosity mapping studies in a large consanguineous family and identified a novel locus at chromosome 10q22.1. Mutation analysis of candidate genes within the target interval identified biallelic germline mutations in SLC29A3 in the FHC kindred and in two families reported to have familial RDD. Analysis of SLC29A3 expression during mouse embryogenesis revealed widespread expression by e14.5 with prominent expression in the central nervous system, eye, inner ear, and epithelial tissues including the gastrointestinal tract. SLC29A3 encodes an intracellular equilibrative nucleoside transporter (hENT3) with affinity for adenosine. Recently germline mutations in SLC29A3 were also described in two rare autosomal recessive disorders with overlapping phenotypes: (a) H syndrome (MIM 612391) that is characterised by cutaneous hyperpigmentation and hypertrichosis, hepatomegaly, heart anomalies, hearing loss, and hypogonadism; and (b) PHID (pigmented hypertrichosis with insulin-dependent diabetes mellitus) syndrome. Our findings suggest that a variety of clinical diagnoses (H and PHID syndromes, FHC, and familial RDD) can be included in a new diagnostic category of SLC29A3 spectrum disorder., Author Summary The histiocytoses are a group of systemic disorders usually confined to childhood and are caused by an excessive number of histiocytes which phagocytose other cells and process antigens. Although nearly a century has passed since histiocytic disorders were recognised, their pathophysiology remains largely unclear, and treatment is nonspecific. The identification of SLC29A3 mutations as the molecular basis for a familial form of syndromic histiocytosis (FHC/RDD) confirms a direct link between Faisalabad histiocytosis and Rosai-Dorfman disease and links these disorders to other SLC29A3-associated phenotypes.
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- 2010
22. Fibrillarin and U3 RNA expression during Xenopus oogenesis and embryo development
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Colette Mathieu, Michèle Caizergues-Ferrer, Francesco Amaldi, François Amalric, Paolo Mariottini, Caizerguesferrer, M, Mathieu, C, Mariottini, Paolo, Amalric, F, and Amaldi, F.
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Embryo, Nonmammalian ,Chromosomal Proteins, Non-Histone ,Messenger ,Xenopus ,Post-Transcriptional ,Non Histone ,Oogenesis ,Xenopus laevis ,Chromosomal Proteins, Non Histone ,fibrillarin ,messenger RNA ,nonhistone protein ,ribonucleoprotein ,small nuclear ribonucleoprotein ,small nuclear RNA ,animal ,animal embryo ,article ,biosynthesis ,embryology ,gene expression regulation ,genetics ,metabolism ,nucleolus ,oocyte ,oocyte development ,RNA processing ,Animal ,Cell Nucleolus ,Gene Expression Regulation ,Oocytes ,Ribonucleoproteins ,Ribonucleoproteins, Small Nuclear ,RNA Processing, Post-Transcriptional ,RNA, Messenger ,RNA, Small Nuclear ,Genetics ,Nonmammalian ,biology ,Settore BIO/11 ,General Medicine ,Chromosomal Proteins ,Rna expression ,Embryo ,RNA Processing ,Small Nuclear ,Animals ,Molecular Biology ,Fibrillarin ,Embryogenesis ,Non-Histone ,biology.organism_classification ,RNA - Published
- 1990
23. A doppler effect in embryonic pattern formation
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Soroldoni, D., Jörg, D. J., Morelli, L. G., Richmond, D. L., Schindelin, J., Jul̈icher, F., and Oates, A. C.
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Kinematics ,Physiology ,tissue level ,animal experiment ,biological rhythm ,embryo ,morphogenesis ,spike wave ,prenatal development ,genetic analysis ,periodicity ,animal embryo ,Article ,zebra fish ,Genetics ,Animals ,animal ,steady state ,germ layer ,Body Patterning ,embryonic structures ,nonhuman ,Nonmammalian ,transgenics ,embryo segmentation ,embryo development ,oscillation ,zebrafish ,Doppler effect ,physical parameters ,cyprinid ,priority journal ,Doppler flowmetry ,Embryonic development ,genetic variation ,Gene expression ,embryo pattern formation - Abstract
During embryonic development, temporal and spatial cues are coordinated to generate a segmented body axis. In sequentially segmenting animals, the rhythm of segmentation is reported to be controlled by the time scale of genetic oscillations that periodically trigger new segment formation. However, we present real-time measurements of genetic oscillations in zebrafish embryos showing that their time scale is not sufficient to explain the temporal period of segmentation. A second time scale, the rate of tissue shortening, contributes to the period of segmentation through a Doppler effect. This contribution is modulated by a gradual change in the oscillation profile across the tissue. We conclude that the rhythm of segmentation is an emergent property controlled by the time scale of genetic oscillations, the change of oscillation profile, and tissue shortening.
24. HairyE/(spl)-related (Her) genes are central components of the segmentation oscillator and display redundancy with the Delta/Notch signaling pathway in the formation of anterior segmental boundaries in the zebrafish
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Oates, A. C. and Ho, R. K.
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antisense oligonucleotide ,genotype ,Oligonucleotides ,her 1 protein ,trunk ,Caenorhabditis elegans protein ,somite ,tail ,Receptors ,zebra fish ,animal ,Developmental ,membrane protein ,transcription factor ,Nonmammalian ,gene expression regulation ,Helminth Proteins ,oscillation ,HER7 protein ,unclassified drug ,priority journal ,helminth protein ,cell cycle ,signal transduction ,Notch ,phenotype ,congenital malformation ,embryo ,morphogenesis ,prenatal development ,animal embryo ,Article ,her-1 protein ,C elegans ,Notch receptor ,Genetics ,Animalia ,Animals ,controlled study ,mutant ,Antisense ,Caenorhabditis elegans Proteins ,Body Patterning ,Vertebrata ,Danio rerio ,nonhuman ,protein her7 ,gene interaction ,embryo segmentation ,Membrane Proteins ,zebrafish protein ,Zebrafish Proteins ,zebrafish ,gene function ,Mutation ,genetic disorder ,Gene expression ,protein ,metabolism ,Transcription Factors - Abstract
We have examined the expression of a Hairy/E(spl)-related (Her) gene, her7, in the zebrafish and show that its expression in the PSM cycles similarly to her1 and deltaC. A decrease in her7 function generated by antisense oligonucleotides disrupts somite formation in the posterior trunk and tail, and disrupts the dynamic expression domains of her1 and deltaC, suggesting that her7 plays a role in coordinating the oscillations of neighboring cells in the presomitic mesoderm. This phenotype is reminiscent of zebrafish segmentation mutants with lesions in genes of the Delta/Notch signaling pathway, which also show a disruption of cyclic her7 expression. The interaction of HER genes with the Delta/Notch signaling system was investigated by introducing a loss of her7 function into mutant backgrounds. This leads to segmental defects more anterior than in either condition alone. Combining a decrease of her7 function with reduction of her7 function results in an enhanced phenotype that affects all the anterior segments, indicating that Her functions in the anterior segments are also partially redundant. In these animals, gene expression does not cycle at any time, suggesting that a complete loss of oscillator function had been achieved. Consistent with this, combining a reduction of her7 and her1 function with a Delta/Notch mutant genotype does not worsen the phenotype further. Thus, our results identify members of the Her family of transcription factors that together behave as a central component of the oscillator, and not as an output. This indicates, therefore, that the function of the segmentation oscillator is restricted to the positioning of segmental boundaries. Furthermore, our data suggest that redundancy between Her genes and genes of the Delta/Notch pathway is in part responsible for the robust formation of anterior somites in vertebrates.
25. A crucial interaction between embryonic red blood cell progenitors and paraxial mesoderm revealed in spadetail embryos
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Rohde, L. A., Oates, A. C., and Ho, R. K.
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Erythrocytes ,Time Factors ,T box transcription factor ,cell maturation ,animal cell ,protein binding ,Stem cells ,fluorescence microscopy ,trunk ,Computer-Assisted ,Models ,zebra fish ,animal ,Developmental ,cell interaction ,gene mutation ,Microscopy ,Nonmammalian ,tbx16 protein ,gene expression regulation ,priority journal ,cell disruption ,fluorescence ,endothelium ,regulatory mechanism ,animal experiment ,embryo ,microcirculation ,macrophage ,animal embryo ,Article ,animal tissue ,Time ,embryo cell ,Image processing ,mesoderm ,Genetics ,Animals ,Cell Lineage ,controlled study ,cell transplantation ,Danio rerio ,nonhuman ,zebrafish protein ,head ,Zebrafish Proteins ,Biological ,biological model ,Hematopoietic Stem Cells ,zebrafish ,microenvironment ,hematopoiesis ,stem cell ,cell differentiation ,gene function ,Mutation ,genetic disorder ,hematopoietic stem cell ,erythrocyte ,Gene expression ,in situ hybridization ,Cytology ,T-Box Domain Proteins ,metabolism ,erythropoiesis - Abstract
Zebrafish embryonic red blood cells (RBCs) develop in trunk intermediate mesoderm (IM), and early macrophages develop in the head, suggesting that local microenvironmental cues regulate differentiation of these two blood lineages. s padetail (spt) mutant embryos, which lack trunk paraxial mesoderm (PM) due to a cell-autonomous defect in tbx16, fail to produce embryonic RBCs but retain head macrophage development. In spt mutants, initial hematopoietic gene expression is absent in trunk IM, although endothelial and pronephric expression is retained, suggesting that early blood progenitor development is specifically disrupted. Using cell transplantation, we reveal that spt is required cell autonomously for early hematopoietic gene expression in trunk IM. Further, we uncover an interaction between embryonic trunk PM and blood progenitors that is essential for RBC development. Importantly, our data identify a hematopoietic microenvironment that allows embryonic RBC production in the zebrafish. © 2004 by Cell Press.
26. Split-Inteins for Simultaneous, site-specific conjugation of Quantum Dots to multiple protein targets In vivo
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Charalambous, Anna, Antoniades, Ioanna, Christodoulou, Neophytos, Skourides, Paris A., and Skourides, Paris A. [0000-0003-3502-5729]
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green fluorescent protein ,Intracellular compartments ,Embryo, Nonmammalian ,dnaE ,Xenopus ,Medicine (miscellaneous) ,Pharmaceutical Science ,enhanced green fluorescent protein ,animal cell ,Xenopus Proteins ,Applied Microbiology and Biotechnology ,C-terminus ,Temporal resolution ,Spatio-temporal dynamics ,biotin ,Target proteins ,Protein targets ,Semiconductor quantum dots ,animal ,genetics ,protein tag ,biotinylation ,focal adhesion ,article ,quantum dot ,protein processing ,Multiparameters ,Molecular complexes ,lcsh:R855-855.5 ,Focal adhesions ,material coating ,Site-specific ,Molecular Medicine ,Biological imaging ,DNA directed DNA polymerase gamma ,Xenopus protein ,DnaB Helicases ,signal transduction ,Dissolution ,conjugation ,DnaB helicase ,lcsh:Medical technology ,in vitro study ,lcsh:Biotechnology ,enzymology ,Split-inteins ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,Live cell ,Biomedical Engineering ,embryo ,Bio-imaging ,Nanotechnology ,prenatal development ,Bioengineering ,protein localization ,Biology ,intein ,animal embryo ,Inteins ,in vivo study ,Multiple targets ,In vivo ,Protein splicing ,In-vivo ,lcsh:TP248.13-248.65 ,complex formation ,Quantum Dots ,protein targeting ,streptavidin ,Animals ,Protein Splicing ,Unique features ,DNA Polymerase III ,hybrid protein ,Protein tagging ,carboxy terminal sequence ,Focal Adhesions ,nonhuman ,Research ,N-terminals ,technology, industry, and agriculture ,Proteins ,Multiple target ,equipment and supplies ,Quantum dot ,DNA polymerase III, alpha subunit ,amino terminal sequence ,RNA ,Paxillin ,Intein ,metabolism ,cell membrane ,Conjugate - Abstract
Background Proteins labelled with Quantum Dots (QDs) can be imaged over long periods of time with ultrahigh spatial and temporal resolution, yielding important information on the spatiotemporal dynamics of proteins within live cells or in vivo. However one of the major problems regarding the use of QDs for biological imaging is the difficulty of targeting QDs onto proteins. We have recently developed a DnaE split intein-based method to conjugate Quantum Dots (QDs) to the C-terminus of target proteins in vivo. In this study, we expand this approach to achieve site-specific conjugation of QDs to two or more proteins simultaneously with spectrally distinguishable QDs for multiparameter imaging of cellular functions. Results Using the DnaE split intein we target QDs to the C-terminus of paxillin and show that paxillin-QD conjugates become localized at focal adhesions allowing imaging of the formation and dissolution of these complexes. We go on to utilize a different split intein, namely Ssp DnaB mini-intein, to demonstrate N-terminal protein tagging with QDs. Combination of these two intein systems allowed us to simultaneously target two distinct proteins with spectrally distinguishable QDs, in vivo, without any cross talk between the two intein systems. Conclusions Multiple target labeling is a unique feature of the intein based methodology which sets it apart from existing tagging methodologies in that, given the large number of characterized split inteins, the number of individual targets that can be simultaneously tagged is only limited by the number of QDs that can be spectrally distinguished within the cell. Therefore, the intein-mediated approach for simultaneous, in vivo, site-specific (N- and C-terminus) conjugation of Quantum Dots to multiple protein targets opens up new possibilities for bioimaging applications and offers an effective system to target QDs and other nanostructures to intracellular compartments as well as specific molecular complexes.
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