Microfluidic-based patterning of embryonic stem cells for in vitro development studies

PubMed Central

Suri, Shalu; Singh, Ankur; Nguyen, Anh H.; Bratt-Leal, Andres M.; McDevitt, Todd C.

2013-01-01

In vitro recapitulation of mammalian embryogenesis and examination of the emerging behaviours of embryonic structures require both the means to engineer complexity and accurately assess phenotypes of multicellular aggregates. Current approaches to study multicellular populations in 3D configurations are limited by the inability to create complex (i.e. spatially heterogeneous) environments in a reproducible manner with high fidelity thus impeding the ability to engineer microenvironments and combinations of cells with similar complexity to that found during morphogenic processes such as development, remodelling and wound healing. Here, we develop a multicellular embryoid body (EB) fusion technique as a higher-throughput in vitro tool, compared to a manual assembly, to generate developmentally relevant embryonic patterns. We describe the physical principles of the EB fusion microfluidic device design; we demonstrate that >60 conjoined EBs can be generated overnight and emulate a development process analogous to mouse gastrulation during early embryogenesis. Using temporal delivery of bone morphogenic protein 4 (BMP4) to embryoid bodies, we recapitulate embryonic day 6.5 (E6.5) during mouse embryo development with induced mesoderm differentiation in murine embryonic stem cells leading to expression of Brachyury-T-green fluorescent protein (T-GFP), an indicator of primitive streak development and mesoderm differentiation during gastrulation. The proposed microfluidic approach could be used to manipulate hundreds or more of individual embryonic cell aggregates in a rapid fashion, thereby allowing controlled differentiation patterns in fused multicellular assemblies to generate complex yet spatially controlled microenvironments. PMID:24113509

Raman-based noninvasive metabolic profile evaluation of in vitro bovine embryos

NASA Astrophysics Data System (ADS)

dos Santos, Érika Cristina; Martinho, Herculano; Annes, Kelly; da Silva, Thais; Soares, Carlos Alexandre; Leite, Roberta Ferreira; Milazzotto, Marcella Pecora

2016-07-01

The timing of the first embryonic cell divisions may predict the ability of an embryo to establish pregnancy. Similarly, metabolic profiles may be markers of embryonic viability. However, in bovine, data about the metabolomics profile of these embryos are still not available. In the present work, we describe Raman-based metabolomic profiles of culture media of bovine embryos with different developmental kinetics (fast x slow) throughout the in vitro culture. The principal component analysis enabled us to classify embryos with different developmental kinetics since they presented specific spectroscopic profiles for each evaluated time point. We noticed that bands at 1076 cm-1 (lipids), 1300 cm-1 (Amide III), and 2719 cm-1 (DNA nitrogen bases) gave the most relevant spectral features, enabling the separation between fast and slow groups. Bands at 1001 cm-1 (phenylalanine) and 2892 cm-1 (methylene group of the polymethylene chain) presented specific patterns related to embryonic stage and can be considered as biomarkers of embryonic development by Raman spectroscopy. The culture media analysis by Raman spectroscopy proved to be a simple and sensitive technique that can be applied with high efficiency to characterize the profiles of in vitro produced bovine embryos with different development kinetics and different stages of development.

[Cell therapy for Parkinson's disease: IV. Risks and future trends].

PubMed

Anisimov, S V

2009-01-01

Motor dysfunctions in Parkinson's disease are believed to be primarily due to the degeneration of dopaminergic neurons located in the substantia nigra pars compacta. Numerous cell replacement therapy approaches have been developed and tested, including these based on donor cell transplantation (embryonic and adult tissue-derived), adult mesenchymal stem cells (hMSCs)-, neural stem cells (hNSCs)- and finally human embryonic stem cells (hESCs)-based. Despite the progress achieved, numerous difficulties prevent wider practical application of stem cell-based therapy approaches for the treatment of Parkinson's disease. Among the latter, ethical, safety and technical issues stand out. Current series of reviews (Cell therapy for Parkinson's disease: I. Embryonic and adult donor tissue-based applications; II. Adult stem cell-based applications; III. Neonatal, fetal and embryonic stem cell-based applications; IV. Risks and future trends) aims providing a balanced and updated view on various issues associated with cell types (including stem cells) in regards to their potential in the treatment of Parkinson's disease. Essential features of the individual cell subtypes, principles of available cell handling protocols, transplantation, and safety issues are discussed extensively.

Role of adiponectin in delayed embryonic development of the short-nosed fruit bat, Cynopterus sphinx.

PubMed

Anuradha; Krishna, Amitabh

2014-12-01

The aim of this study was to evaluate the role of adiponectin in the delayed embryonic development of Cynopterus sphinx. Adiponectin receptor (ADIPOR1) abundance was first observed to be lower during the delayed versus non-delayed periods of utero-embryonic unit development. The effects of adiponectin treatment on embryonic development were then evaluated during the period of delayed development. Exogenous treatment increased the in vivo rate of embryonic development, as indicated by an increase in weight, ADIPOR1 levels in the utero-embryonic unit, and histological changes in embryonic development. Treatment with adiponectin during embryonic diapause showed a significant increase in circulating progesterone and estradiol concentrations, and in production of their receptors in the utero-embryonic unit. The adiponectin-induced increase in estradiol synthesis was correlated with increased cell survival (BCL2 protein levels) and cell proliferation (PCNA protein levels) in the utero-embryonic unit, suggesting an indirect effect of adiponectin via estradiol synthesis by the ovary. An in vitro study further confirmed the in vivo findings that adiponectin treatment increases PCNA levels together with increased uptake of glucose by increasing the abundance of glucose transporter 8 (GLUT8) in the utero-embryonic unit. The in vitro study also revealed that adiponectin, together with estradiol but not alone, significantly increased ADIPOR1 protein levels. Thus, adiponectin works in concert with estradiol to increase glucose transport to the utero-embryonic unit and promote cell proliferation, which together accelerate embryonic development. © 2014 Wiley Periodicals, Inc.

An integrated miRNA functional screening and target validation method for organ morphogenesis.

PubMed

Rebustini, Ivan T; Vlahos, Maryann; Packer, Trevor; Kukuruzinska, Maria A; Maas, Richard L

2016-03-16

The relative ease of identifying microRNAs and their increasing recognition as important regulators of organogenesis motivate the development of methods to efficiently assess microRNA function during organ morphogenesis. In this context, embryonic organ explants provide a reliable and reproducible system that recapitulates some of the important early morphogenetic processes during organ development. Here we present a method to target microRNA function in explanted mouse embryonic organs. Our method combines the use of peptide-based nanoparticles to transfect specific microRNA inhibitors or activators into embryonic organ explants, with a microRNA pulldown assay that allows direct identification of microRNA targets. This method provides effective assessment of microRNA function during organ morphogenesis, allows prioritization of multiple microRNAs in parallel for subsequent genetic approaches, and can be applied to a variety of embryonic organs.

Promotion of human early embryonic development and blastocyst outgrowth in vitro using autocrine/paracrine growth factors.

PubMed

Kawamura, Kazuhiro; Chen, Yuan; Shu, Yimin; Cheng, Yuan; Qiao, Jie; Behr, Barry; Pera, Renee A Reijo; Hsueh, Aaron J W

2012-01-01

Studies using animal models demonstrated the importance of autocrine/paracrine factors secreted by preimplantation embryos and reproductive tracts for embryonic development and implantation. Although in vitro fertilization-embryo transfer (IVF-ET) is an established procedure, there is no evidence that present culture conditions are optimal for human early embryonic development. In this study, key polypeptide ligands known to be important for early embryonic development in animal models were tested for their ability to improve human early embryo development and blastocyst outgrowth in vitro. We confirmed the expression of key ligand/receptor pairs in cleavage embryos derived from discarded human tri-pronuclear zygotes and in human endometrium. Combined treatment with key embryonic growth factors (brain-derived neurotrophic factor, colony-stimulating factor, epidermal growth factor, granulocyte macrophage colony-stimulating factor, insulin-like growth factor-1, glial cell-line derived neurotrophic factor, and artemin) in serum-free media promoted >2.5-fold the development of tri-pronuclear zygotes to blastocysts. For normally fertilized embryos, day 3 surplus embryos cultured individually with the key growth factors showed >3-fold increases in the development of 6-8 cell stage embryos to blastocysts and >7-fold increase in the proportion of high quality blastocysts based on Gardner's criteria. Growth factor treatment also led to a 2-fold promotion of blastocyst outgrowth in vitro when day 7 surplus hatching blastocysts were used. When failed-to-be-fertilized oocytes were used to perform somatic cell nuclear transfer (SCNT) using fibroblasts as donor karyoplasts, inclusion of growth factors increased the progression of reconstructed SCNT embryos to >4-cell stage embryos. Growth factor supplementation of serum-free cultures could promote optimal early embryonic development and implantation in IVF-ET and SCNT procedures. This approach is valuable for infertility treatment and future derivation of patient-specific embryonic stem cells.

Long-term in vivo study of vertebrate embryonic development using noninvasive harmonics optical microscopy

NASA Astrophysics Data System (ADS)

Chen, Szu-Yu; Hsieh, C.-S.; Chu, S.-W.; Lin, Cheng-Yung; Ko, C.-Y.; Chen, Y.-C.; Tsai, Huai-Jen; Hu, C.-H.; Sun, Chi-Kuang

2005-03-01

Harmonics optical microscopy (HOM) provides a truly "noninvasive" tool for in vivo and long-term study of vertebrate embryonic development. Based on the nonlinear natures, it provides sub-micrometer 3D spatial resolution and high 3D optical-sectioning power (~1μm axial resolution) without using invasive and toxic fluorophores. Since only virtual-level-transition is involved, HOM is known to leave no energy deposition and no photodamages. Combined with second harmonic generation, which is sensitive to specific structure such as nerve and muscle fibers, HOM can be used to do functional studies of early developmental dynamics of many vertebrate physiological systems. Recently, zebrafish has become a standard model for many biological and medical studies of vertebrates, due to the similarity between embryonic development of zebrafish and human being. Zebrafish embryos now have been used to study many vertebrate physiological systems. We have demonstrated an in vivo HOM study of developmental dynamics of several embryonic physiological systems in live zebrafish embryos, with focuses on the developments of brains, eyes, ears, and hearts. Based on a femtosecond Cr:forsterite laser, which provides the deepest penetration (~1.5mm) and least photodamage in the zebrafish embryo, complete developing processes of different physiological systems within a period of time longer than 20 hours can be non-invasively observed inside the same embryo.

Measurement of wall shear stress in chick embryonic heart using optical coherence tomography

NASA Astrophysics Data System (ADS)

Ma, Zhenhe; Dou, Shidan; Zhao, Yuqian; Wang, Yi; Suo, Yanyan; Wang, Fengwen

2015-03-01

The cardiac development is a complicated process affected by genetic and environmental factors. Wall shear stress (WSS) is one of the components which have been proved to influence the morphogenesis during early stages of cardiac development. To study the mechanism, WSS measurement is a step with significant importance. WSS is caused by blood flow imposed on the inner surface of the heart wall and it can be determined by calculating velocity gradients of blood flow in a direction perpendicular to the wall. However, the WSS of the early stage embryonic heart is difficult to measure since the embryonic heart is tiny and beating fast. Optical coherence tomography (OCT) is a non-invasive imaging modality with high spatial and temporal resolution, which is uniquely suitable for the study of early stage embryonic heart development. In this paper, we introduce a method to measure the WSS of early stage chick embryonic heart based on high speed spectral domain optical coherence tomography (SDOCT). 4D (x,y,z,t) scan was performed on the outflow tract (OFT) of HH18 (~3 days of incubation) chick embryonic heart. After phase synchronization, OFT boundary segmentation, and OFT center line calculation, Doppler angle of the blood flow in the OFT can be achieved (This method has been described in previous publications). Combining with the Doppler OCT results, we calculate absolute blood flow velocity distribution in the OFT. The boundary of the OFT was segmented at each cross-sectional structural image, then geometrical center of the OFT can be calculated. Thus, the gradients of blood flow in radial direction can be calculated. This velocity gradient near the wall is termed wall shear rate and the WSS value is proportional to the wall shear rate. Based on this method, the WSS at different heart beating phase are compare. The result demonstrates that OCT is capable of early stage chicken embryonic heart WSS study.

Comparative ovicidal activity of Moringa oleifera leaf extracts on Fasciola gigantica eggs

PubMed Central

Hegazi, Ahmed G.; Megeed, Kadria N. Abdel; Hassan, Soad E.; Abdelaziz, M. M.; Toaleb, Nagwa I.; Shanawany, Eman E. El; Aboelsoued, Dina

2018-01-01

Background: Fasciolosis is an important zoonotic disease affecting the productive performance of farm animals in Egypt. Aim: The aim of the present study was comparing the ovicidal effect of different extracts as an alcoholic (Methanolic and Ethanolic) and aqueous Moringa oleifera leaf extracts on Fasciola gigantica non-embryonated and developed eggs. Materials and Methods: Tested concentrations of extracts ranged from 12.5 to 800 mg/ml. Nitroxynil was used as reference drug with a dose of 100 mg/ml. Results: M. oleifera alcoholic and aqueous extracts showed a concentration-dependent ovicidal effect on F. gigantica non-embryonated and developed eggs. Based on LC50 values, water extract showed the highest ovicidal activity since it registered the lowest values of 2.6 mg/ml on non-embryonated eggs. Non-embryonated eggs were more susceptible to aqueous extract than developed eggs. On the other hand, the developed eggs were more susceptible to ethanolic extract than non-embryonated eggs even the lowest LC50 (12.38 mg/ml). Conclusion: M. oleifera leaf extracts especially aqueous extract could be a promising step in the field of controlling fascioliasis. Further, in vivo studies are needed to enlighten the therapeutic potential of M. oleifera extracts in treating F. gigantica infection. PMID:29657406

WAVE2 deficiency reveals distinct roles in embryogenesis and Rac-mediated actin-based motility

PubMed Central

Yan, Catherine; Martinez-Quiles, Narcisa; Eden, Sharon; Shibata, Tomoyuki; Takeshima, Fuminao; Shinkura, Reiko; Fujiwara, Yuko; Bronson, Roderick; Snapper, Scott B.; Kirschner, Marc W.; Geha, Raif; Rosen, Fred S.; Alt, Frederick W.

2003-01-01

The Wiskott–Aldrich syndrome related protein WAVE2 is implicated in the regulation of actin-cytoskeletal reorganization downstream of the small Rho GTPase, Rac. We inactivated the WAVE2 gene by gene-targeted mutation to examine its role in murine development and in actin assembly. WAVE2-deficient embryos survived until approximately embryonic day 12.5 and displayed growth retardation and certain morphological defects, including malformations of the ventricles in the developing brain. WAVE2-deficient embryonic stem cells displayed normal proliferation, whereas WAVE2-deficient embryonic fibroblasts exhibited severe growth defects, as well as defective cell motility in response to PDGF, lamellipodium formation and Rac-mediated actin polymerization. These results imply a non-redundant role for WAVE2 in murine embryogenesis and a critical role for WAVE2 in actin-based processes downstream of Rac that are essential for cell movement. PMID:12853475

WAVE2 deficiency reveals distinct roles in embryogenesis and Rac-mediated actin-based motility.

PubMed

Yan, Catherine; Martinez-Quiles, Narcisa; Eden, Sharon; Shibata, Tomoyuki; Takeshima, Fuminao; Shinkura, Reiko; Fujiwara, Yuko; Bronson, Roderick; Snapper, Scott B; Kirschner, Marc W; Geha, Raif; Rosen, Fred S; Alt, Frederick W

2003-07-15

The Wiskott-Aldrich syndrome related protein WAVE2 is implicated in the regulation of actin-cytoskeletal reorganization downstream of the small Rho GTPase, Rac. We inactivated the WAVE2 gene by gene-targeted mutation to examine its role in murine development and in actin assembly. WAVE2-deficient embryos survived until approximately embryonic day 12.5 and displayed growth retardation and certain morphological defects, including malformations of the ventricles in the developing brain. WAVE2-deficient embryonic stem cells displayed normal proliferation, whereas WAVE2-deficient embryonic fibroblasts exhibited severe growth defects, as well as defective cell motility in response to PDGF, lamellipodium formation and Rac-mediated actin polymerization. These results imply a non-redundant role for WAVE2 in murine embryogenesis and a critical role for WAVE2 in actin-based processes downstream of Rac that are essential for cell movement.

Importance of the pluripotency factor LIN28 in the mammalian nucleolus during early embryonic development.

PubMed

Vogt, Edgar J; Meglicki, Maciej; Hartung, Kristina Ilka; Borsuk, Ewa; Behr, Rüdiger

2012-12-01

The maternal nucleolus is required for proper activation of the embryonic genome (EGA) and early embryonic development. Nucleologenesis is characterized by the transformation of a nucleolar precursor body (NPB) to a mature nucleolus during preimplantation development. However, the function of NPBs and the involved molecular factors are unknown. We uncover a novel role for the pluripotency factor LIN28, the biological significance of which was previously demonstrated in the reprogramming of human somatic cells to induced pluripotent stem (iPS) cells. Here, we show that LIN28 accumulates at the NPB and the mature nucleolus in mouse preimplantation embryos and embryonic stem cells (ESCs), where it colocalizes with the nucleolar marker B23 (nucleophosmin 1). LIN28 has nucleolar localization in non-human primate (NHP) preimplantation embryos, but is cytoplasmic in NHP ESCs. Lin28 transcripts show a striking decline before mouse EGA, whereas LIN28 protein localizes to NPBs at the time of EGA. Following knockdown with a Lin28 morpholino, the majority of embryos arrest between the 2- and 4-cell stages and never develop to morula or blastocyst. Lin28 morpholino-injected embryos arrested at the 2-cell stage were not enriched with nucleophosmin at presumptive NPB sites, indicating that functional NPBs were not assembled. Based on these results, we propose that LIN28 is an essential factor of nucleologenesis during early embryonic development.

Somatic cell nuclear transfer in Oregon: expanding the pluripotent space and informing research ethics.

PubMed

Lomax, Geoffrey P; DeWitt, Natalie D

2013-12-01

In May, Oregon Health and Science University (OHSU) announced the successful derivation, by the Mitalipov laboratory, of embryonic stem cells by somatic cell nuclear transfer. This experiment was recognized as a "formidable technical feat" and potentially a key step toward developing cell-based therapies. The OHSU report is also an example of how a scientific breakthrough can inform research ethics. This article suggests ways that nuclear transfer embryonic stem cell lines may contribute to research ethics by adding rigor to studies addressing pressing research questions important to the development of cell-based therapies.

Embryonic domains of the aorta derived from diverse origins exhibit distinct properties that converge into a common phenotype in the adult

PubMed Central

Pfaltzgraff, Elise R.; Shelton, Elaine L.; Galindo, Cristi L.; Nelms, Brian L.; Hooper, Christopher W.; Poole, Stanley D.; Labosky, Patricia A.; Bader, David M.; Reese, Jeff

2014-01-01

Vascular smooth muscle cells (VSMCs) are derived from distinct embryonic origins. Vessels originating from differing smooth muscle cell populations have distinct vascular and pathological properties involving calcification, atherosclerosis, and structural defects such as aneurysm and coarctation. We hypothesized that domains within a single vessel, such as the aorta, vary in phenotype based on embryonic origin. Gene profiling and myographic analyses demonstrated that embryonic ascending and descending aortic domains exhibited distinct phenotypes. In vitro analyses demonstrated that VSMCs from each region were dissimilar in terms of cytoskeletal and migratory properties, and retention of different gene expression patterns. Using the same analysis, we found that these same two domains are indistinguishable in the adult vessel. Our data demonstrate that VSMCs from different embryonic origins are functionally distinct in the embryonic mouse, but converge to assume a common phenotype in the aorta of healthy adults. These findings have fundamental implications for aortic development, function and disease progression. PMID:24508561

Gene expression dynamics during embryonic development in rainbow trout

USDA-ARS?s Scientific Manuscript database

The supply of maternal RNAs in fertilized egg and activation of embryonic genome during maternal-zygotic transition (MZT) are important for normal embryonic development. In order to identify genes and gene products that are essential in the regulation of embryonic development in rainbow trout, RNA-S...

Multidisciplinary Inquiry-Based Investigation Learning Using an Ex Ovo Chicken Culture Platform: Role of Vitamin A on Embryonic Morphogenesis

ERIC Educational Resources Information Center

Buskohl, Philip R.; Gould, Russell A.; Curran, Susan; Archer, Shivaun D.; Butcher, Jonathan T.

2012-01-01

Embryonic development offers a unique perspective on the function of many biological processes because of embryos' heightened sensitivity to environmental factors. This hands-on lesson investigates the effects of elevated vitamin A on the morphogenesis of chicken embryos. The active form of vitamin A (retinoic acid) is applied to shell-less (ex…

Altered glucose transport to utero-embryonic unit in relation to delayed embryonic development in the Indian short-nosed fruit bat, Cynopterus sphinx.

PubMed

Arnab, Banerjee; Amitabh, Krishna

2011-02-10

The aim of this study was to compare the changes in concentration of glucose and glucose transporters (GLUTs) in the utero-embryonic unit, consisting of decidua, trophoblast and embryo, during delayed and non-delayed periods to understand the possible cause of delayed embryonic development in Cynopterus sphinx. The results showed a significantly decreased concentration of glucose in the utero-embryonic unit due to decline in the expression of insulin receptor (IR) and GLUT 3, 4 and 8 proteins in the utero-embryonic unit during delayed period. The in vitro study showed suppressive effect of insulin on expression of GLUTs 4 and 8 in the utero-embryonic unit and a significant positive correlation between the decreased amount of glucose consumed by the utero-embryonic unit and decreased expression of GLUTs 4 (r=0.99; p<0.05) and 8 (r=0.98; p<0.05). The in vivo study showed expression of IR and GLUT 4 proteins in adipose tissue during November suggesting increased transport of glucose to adipose tissue for adipogenesis. This study showed increased expression of HSL and OCTN2 and increased availability of l-carnitine to utero-embryonic unit suggesting increased transport of fatty acid to utero-embryonic unit during the period of delayed embryonic development. Hence it appears that due to increased transport of glucose for adipogenesis prior to winter, glucose utilization by utero-embryonic unit declines and this may be responsible for delayed embryonic development in C. sphinx. Increased supply of fatty acid to the delayed embryo may be responsible for its survival under low glucose condition but unable to promote embryonic development in C. sphinx. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Case Study: Organotypic human in vitro models of embryonic ...

EPA Pesticide Factsheets

Morphogenetic fusion of tissues is a common event in embryonic development and disruption of fusion is associated with birth defects of the eye, heart, neural tube, phallus, palate, and other organ systems. Embryonic tissue fusion requires precise regulation of cell-cell and cell-matrix interactions that drive proliferation, differentiation, and morphogenesis. Chemical low-dose exposures can disrupt morphogenesis across space and time by interfering with key embryonic fusion events. The Morphogenetic Fusion Task uses computer and in vitro models to elucidate consequences of developmental exposures. The Morphogenetic Fusion Task integrates multiple approaches to model responses to chemicals that leaad to birth defects, including integrative mining on ToxCast DB, ToxRefDB, and chemical structures, advanced computer agent-based models, and human cell-based cultures that model disruption of cellular and molecular behaviors including mechanisms predicted from integrative data mining and agent-based models. The purpose of the poster is to indicate progress on the CSS 17.02 Virtual Tissue Models Morphogenesis Task 1 products for the Board of Scientific Counselors meeting on Nov 16-17.

Stability of citrate-capped silver nanoparticles in exposure media and their effects on the development of embryonic zebrafish (Danio rerio)

PubMed Central

Park, Kwangsik; Tuttle, George; Sinche, Federico; Harper, Stace L.

2014-01-01

The stability of citrate-capped silver nanoparticles (AgNPs) and the embryonic developmental toxicity were evaluated in the fish test water. Serious aggregation of AgNPs was observed in undiluted fish water (DM-100) in which high concentration of ionic salts exist. However, AgNPs were found to be stable for 7 days in DM-10, prepared by diluting the original fish water (DM-100) with deionized water to 10%. The normal physiology of zebrafish embryos were evaluated in DM-10 to see if DM-10 can be used as a control vehicle for the embryonic fish toxicity test. As results, DM-10 without AgNPs did not induce any significant adverse effects on embryonic development of zebrafish determined by mortality, hatching, malformations and heart rate. When embryonic toxicity of AgNPs was tested in both DM-10 and in DM-100, AgNPs showed higher toxicity in DM-10 than in DM-100. This means that the big-sized aggregates of AgNPs were low toxic compared to the nano-sized AgNPs. AgNPs induced delayed hatching, decreased heart rate, pericardial edema, and embryo death. Accumulation of AgNPs in the embryo bodies was also observed. Based on this study, citrate-capped AgNPs are not aggregated in DM-10 and it can be used as a control vehicle in the toxicity test of fish embryonic development. PMID:23325492

Melatonin regulates delayed embryonic development in the short-nosed fruit bat, Cynopterus sphinx.

PubMed

Banerjee, Arnab; Meenakumari, K J; Udin, S; Krishna, A

2009-12-01

The aim of the present study was to evaluate the seasonal variation in serum melatonin levels and their relationship to the changes in the serum progesterone level, ovarian steroidogenesis, and embryonic development during two successive pregnancies of Cynopterus sphinx. Circulating melatonin concentrations showed two peaks; one coincided with the period of low progesterone synthesis and delayed embryonic development, whereas the second peak coincided with regressing corpus luteum. This finding suggests that increased serum melatonin level during November-December may be responsible for delayed embryonic development by suppressing progesterone synthesis. The study showed increased melatonin receptors (MTNR1A and MTNR1B) in the corpus luteum and in the utero-embryonic unit during the period of delayed embryonic development. The in vitro study showed that a high dose of melatonin suppressed progesterone synthesis, whereas a lower dose of melatonin increased progesterone synthesis by the ovary. The effects of melatonin on ovarian steroidogenesis are mediated through changes in the expression of peripheral-type benzodiazepine receptor, P450 side chain cleavage enzyme, and LH receptor proteins. This study further showed a suppressive impact of melatonin on the progesterone receptor (PGR) in the utero-embryonic unit; this effect might contribute to delayed embryonic development in C. sphinx. The results of the present study thus suggest that a high circulating melatonin level has a dual contribution in retarding embryonic development in C. sphinx by impairing progesterone synthesis as well as by inhibiting progesterone action by reducing expression of PGR in the utero-embryonic unit.

Evidence of local adaptation in westslope cutthroat trout

USGS Publications Warehouse

Drinan, Daniel P.; Zale, Alexander V.; Webb, Molly A.H.; Taper, Mark L.; Shepard, Bradley B.; Kalinowski, Steven T.

2012-01-01

An understanding of the process of local adaptation would allow managers to better protect and conserve species. Many salmonids are in need of such efforts, and because they often persist in differing, isolated environments, they are useful organisms for studying local adaptation. In addition, the temperature sensitivity of salmonids provides a likely target for natural selection. We studied thermal adaptation in four wild populations and one hatchery stock of westslope cutthroat trout Oncorhynchus clarkii lewisi . The mean summer temperatures of source streams ranged from 6.7°C to 11.2°C. Embryos were collected from the wild, and embryonic development, embryonic survival, and juvenile growth were determined. A significant relationship between median embryonic survival and source stream temperature was detected. Based on a rank test, populations from colder streams had a greater decline in median embryonic survival at warm temperatures than populations from warmer streams. Embryonic development and juvenile growth did not appear to be influenced by source. These findings suggest that populations are thermally adapted to their source streams and this should be considered by managers. However, further study is necessary to sort out the potential confounding factors, whether genetic or epigenetic.

Directed differentiation of embryonic stem cells using a bead-based combinatorial screening method.

PubMed

Tarunina, Marina; Hernandez, Diana; Johnson, Christopher J; Rybtsov, Stanislav; Ramathas, Vidya; Jeyakumar, Mylvaganam; Watson, Thomas; Hook, Lilian; Medvinsky, Alexander; Mason, Chris; Choo, Yen

2014-01-01

We have developed a rapid, bead-based combinatorial screening method to determine optimal combinations of variables that direct stem cell differentiation to produce known or novel cell types having pre-determined characteristics. Here we describe three experiments comprising stepwise exposure of mouse or human embryonic cells to 10,000 combinations of serum-free differentiation media, through which we discovered multiple novel, efficient and robust protocols to generate a number of specific hematopoietic and neural lineages. We further demonstrate that the technology can be used to optimize existing protocols in order to substitute costly growth factors with bioactive small molecules and/or increase cell yield, and to identify in vitro conditions for the production of rare developmental intermediates such as an embryonic lymphoid progenitor cell that has not previously been reported.

Pipette-based Method to Study Embryoid Body Formation Derived from Mouse and Human Pluripotent Stem Cells Partially Recapitulating Early Embryonic Development Under Simulated Microgravity Conditions

NASA Astrophysics Data System (ADS)

Shinde, Vaibhav; Brungs, Sonja; Hescheler, Jürgen; Hemmersbach, Ruth; Sachinidis, Agapios

2016-06-01

The in vitro differentiation of pluripotent stem cells partially recapitulates early in vivo embryonic development. More recently, embryonic development under the influence of microgravity has become a primary focus of space life sciences. In order to integrate the technique of pluripotent stem cell differentiation with simulated microgravity approaches, the 2-D clinostat compatible pipette-based method was experimentally investigated and adapted for investigating stem cell differentiation processes under simulated microgravity conditions. In order to keep residual accelerations as low as possible during clinorotation, while also guaranteeing enough material for further analysis, stem cells were exposed in 1-mL pipettes with a diameter of 3.5 mm. The differentiation of mouse and human pluripotent stem cells inside the pipettes resulted in the formation of embryoid bodies at normal gravity (1 g) after 24 h and 3 days. Differentiation of the mouse pluripotent stem cells on a 2-D pipette-clinostat for 3 days also resulted in the formation of embryoid bodies. Interestingly, the expression of myosin heavy chain was downregulated when cultivation was continued for an additional 7 days at normal gravity. This paper describes the techniques for culturing and differentiation of pluripotent stem cells and exposure to simulated microgravity during culturing or differentiation on a 2-D pipette clinostat. The implementation of these methodologies along with -omics technologies will contribute to understand the mechanisms regulating how microgravity influences early embryonic development.

Blastocyst-like structures generated solely from stem cells.

PubMed

Rivron, Nicolas C; Frias-Aldeguer, Javier; Vrij, Erik J; Boisset, Jean-Charles; Korving, Jeroen; Vivié, Judith; Truckenmüller, Roman K; van Oudenaarden, Alexander; van Blitterswijk, Clemens A; Geijsen, Niels

2018-05-01

The blastocyst (the early mammalian embryo) forms all embryonic and extra-embryonic tissues, including the placenta. It consists of a spherical thin-walled layer, known as the trophectoderm, that surrounds a fluid-filled cavity sheltering the embryonic cells 1 . From mouse blastocysts, it is possible to derive both trophoblast 2 and embryonic stem-cell lines 3 , which are in vitro analogues of the trophectoderm and embryonic compartments, respectively. Here we report that trophoblast and embryonic stem cells cooperate in vitro to form structures that morphologically and transcriptionally resemble embryonic day 3.5 blastocysts, termed blastoids. Like blastocysts, blastoids form from inductive signals that originate from the inner embryonic cells and drive the development of the outer trophectoderm. The nature and function of these signals have been largely unexplored. Genetically and physically uncoupling the embryonic and trophectoderm compartments, along with single-cell transcriptomics, reveals the extensive inventory of embryonic inductions. We specifically show that the embryonic cells maintain trophoblast proliferation and self-renewal, while fine-tuning trophoblast epithelial morphogenesis in part via a BMP4/Nodal-KLF6 axis. Although blastoids do not support the development of bona fide embryos, we demonstrate that embryonic inductions are crucial to form a trophectoderm state that robustly implants and triggers decidualization in utero. Thus, at this stage, the nascent embryo fuels trophectoderm development and implantation.

Live dynamic imaging and analysis of developmental cardiac defects in mouse models with optical coherence tomography

NASA Astrophysics Data System (ADS)

Lopez, Andrew L.; Wang, Shang; Garcia, Monica; Valladolid, Christian; Larin, Kirill V.; Larina, Irina V.

2015-03-01

Understanding mouse embryonic development is an invaluable resource for our interpretation of normal human embryology and congenital defects. Our research focuses on developing methods for live imaging and dynamic characterization of early embryonic development in mouse models of human diseases. Using multidisciplinary methods: optical coherence tomography (OCT), live mouse embryo manipulations and static embryo culture, molecular biology, advanced image processing and computational modeling we aim to understand developmental processes. We have developed an OCT based approach to image live early mouse embryos (E8.5 - E9.5) cultured on an imaging stage and visualize developmental events with a spatial resolution of a few micrometers (less than the size of an individual cell) and a frame rate of up to hundreds of frames per second and reconstruct cardiodynamics in 4D (3D+time). We are now using these methods to study how specific embryonic lethal mutations affect cardiac morphology and function during early development.

Female parthenogenetic apomixis and androsporogenetic parthenogenesis in embryonal cells of Araucaria angustifolia: interpolation of progenesis and asexual heterospory in an artificial sporangium.

PubMed

Durzan, Don J

2012-09-01

Cell fate, development timing and occurrence of reproductive versus apomictic development in gymnosperms are shown to be influenced by culture conditions in vitro. In this study, female parthenogenetic apomixis (fPA), androsporogenetic parthenogenesis (mAP) and progenesis were demonstrated using embryonal initials of Araucaria angustifolia in scaled-up cell suspensions passing through a single-cell bottleneck in darkness and in an artificial sporangium (AS). Expression was based on defined nutrition, hormones and feedforward-adaptive feedback process controls at 23-25 °C and in darkness. In fPA, the nucleus of an embryonal initial undergoes endomitosis and amitosis, forming a diploid egg-equivalent and an apoptotic ventral canal nucleus in a transdifferentiated archegonial tube. Discharge of egg-equivalent cells as parthenospores and their dispersal into the aqueous culture medium were followed by free-nuclear conifer-type proembryogenesis. This replaced the plesiomorphic and central features of proembryogenesis in Araucariaceae. Protoplasmic fusions of embryonal initials were used to reconstruct heterokaryotic expressions of fPA in multiwell plates. In mAP, restitutional meiosis (automixis) was responsible for androsporogenesis and the discharge of monads, dyads, tetrads and polyads. In a display of progenesis, reproductive development was brought to an earlier ontogenetic stage and expressed by embryonal initials. Colchicine increased polyploidy, but androspore formation became aberrant and fragmented. Aberrant automixis led to the formation of chromosomal bouquets, which contributed to genomic silencing in embryonal initials, cytomixis and the formation of pycnotic micronucleated cells. Dispersal of female and male parthenospores displayed heteromorphic asexual heterospory in an aqueous environment.

Directed Differentiation of Embryonic Stem Cells Using a Bead-Based Combinatorial Screening Method

PubMed Central

Tarunina, Marina; Hernandez, Diana; Johnson, Christopher J.; Rybtsov, Stanislav; Ramathas, Vidya; Jeyakumar, Mylvaganam; Watson, Thomas; Hook, Lilian; Medvinsky, Alexander; Mason, Chris; Choo, Yen

2014-01-01

We have developed a rapid, bead-based combinatorial screening method to determine optimal combinations of variables that direct stem cell differentiation to produce known or novel cell types having pre-determined characteristics. Here we describe three experiments comprising stepwise exposure of mouse or human embryonic cells to 10,000 combinations of serum-free differentiation media, through which we discovered multiple novel, efficient and robust protocols to generate a number of specific hematopoietic and neural lineages. We further demonstrate that the technology can be used to optimize existing protocols in order to substitute costly growth factors with bioactive small molecules and/or increase cell yield, and to identify in vitro conditions for the production of rare developmental intermediates such as an embryonic lymphoid progenitor cell that has not previously been reported. PMID:25251366

The roles of ERAS during cell lineage specification of mouse early embryonic development.

PubMed

Zhao, Zhen-Ao; Yu, Yang; Ma, Huai-Xiao; Wang, Xiao-Xiao; Lu, Xukun; Zhai, Yanhua; Zhang, Xiaoxin; Wang, Haibin; Li, Lei

2015-08-01

Eras encodes a Ras-like GTPase protein that was originally identified as an embryonic stem cell-specific Ras. ERAS has been known to be required for the growth of embryonic stem cells and stimulates somatic cell reprogramming, suggesting its roles on mouse early embryonic development. We now report a dynamic expression pattern of Eras during mouse peri-implantation development: its expression increases at the blastocyst stage, and specifically decreases in E7.5 mesoderm. In accordance with its expression pattern, the increased expression of Eras promotes cell proliferation through controlling AKT activation and the commitment from ground to primed state through ERK activation in mouse embryonic stem cells; and the reduced expression of Eras facilitates primitive streak and mesoderm formation through AKT inhibition during gastrulation. The expression of Eras is finely regulated to match its roles in mouse early embryonic development during which Eras expression is negatively regulated by the β-catenin pathway. Thus, beyond its well-known role on cell proliferation, ERAS may also play important roles in cell lineage specification during mouse early embryonic development. © 2015 The Authors.

Embryonic mammary signature subsets are activated in Brca1-/- and basal-like breast cancers

PubMed Central

2013-01-01

Introduction Cancer is often suggested to result from development gone awry. Links between normal embryonic development and cancer biology have been postulated, but no defined genetic basis has been established. We recently published the first transcriptomic analysis of embryonic mammary cell populations. Embryonic mammary epithelial cells are an immature progenitor cell population, lacking differentiation markers, which is reflected in their very distinct genetic profiles when compared with those of their postnatal descendents. Methods We defined an embryonic mammary epithelial signature that incorporates the most highly expressed genes from embryonic mammary epithelium when compared with the postnatal mammary epithelial cells. We looked for activation of the embryonic mammary epithelial signature in mouse mammary tumors that formed in mice in which Brca1 had been conditionally deleted from the mammary epithelium and in human breast cancers to determine whether any genetic links exist between embryonic mammary cells and breast cancers. Results Small subsets of the embryonic mammary epithelial signature were consistently activated in mouse Brca1-/- tumors and human basal-like breast cancers, which encoded predominantly transcriptional regulators, cell-cycle, and actin cytoskeleton components. Other embryonic gene subsets were found activated in non-basal-like tumor subtypes and repressed in basal-like tumors, including regulators of neuronal differentiation, transcription, and cell biosynthesis. Several embryonic genes showed significant upregulation in estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and/or grade 3 breast cancers. Among them, the transcription factor, SOX11, a progenitor cell and lineage regulator of nonmammary cell types, is found highly expressed in some Brca1-/- mammary tumors. By using RNA interference to silence SOX11 expression in breast cancer cells, we found evidence that SOX11 regulates breast cancer cell proliferation and cell survival. Conclusions Specific subsets of embryonic mammary genes, rather than the entire embryonic development transcriptomic program, are activated in tumorigenesis. Genes involved in embryonic mammary development are consistently upregulated in some breast cancers and warrant further investigation, potentially in drug-discovery research endeavors. PMID:23506684

Identification of microRNAs controlling hepatic mRNA levels for metabolic genes during the metabolic transition from embryonic to posthatch development in the chicken.

PubMed

Hicks, Julie A; Porter, Tom E; Liu, Hsiao-Ching

2017-09-05

The transition from embryonic to posthatch development in the chicken represents a massive metabolic switch from primarily lipolytic to primarily lipogenic metabolism. This metabolic switch is essential for the chick to successfully transition from the metabolism of stored egg yolk to the utilization of carbohydrate-based feed. However, regulation of this metabolic switch is not well understood. We hypothesized that microRNAs (miRNAs) play an important role in the metabolic switch that is essential to efficient growth of chickens. We used high-throughput RNA sequencing to characterize expression profiles of mRNA and miRNA in liver during late embryonic and early posthatch development of the chicken. This extensive data set was used to define the contributions of microRNAs to the metabolic switch during development that is critical to growth and nutrient utilization in chickens. We found that expression of over 800 mRNAs and 30 miRNAs was altered in the embryonic liver between embryonic day 18 and posthatch day 3, and many of these differentially expressed mRNAs and miRNAs are associated with metabolic processes. We confirmed the regulation of some of these mRNAs by miRNAs expressed in a reciprocal pattern using luciferase reporter assays. Finally, through the use of yeast one-hybrid screens, we identified several proteins that likely regulate expression of one of these important miRNAs. Integration of the upstream regulatory mechanisms governing miRNA expression along with monitoring the downstream effects of this expression will ultimately allow for the construction of complete miRNA regulatory networks associated with the hepatic metabolic switch in chickens. Our findings support a key role for miRNAs in controlling the metabolic switch that occurs between embryonic and posthatch development in the chicken.

Medical student retention of embryonic development: impact of the dimensions added by multimedia tutorials.

PubMed

Marsh, Karen R; Giffin, Bruce F; Lowrie, Donald J

2008-01-01

The purpose of this project was to develop Web-based learning modules that combine (1) animated 3D graphics; (2) 3D models that a student can manipulate independently; (3) passage of time in embryonic development; and (4) animated 2D graphics, including 2D cross-sections that represent different "slices" of the embryo, and animate in parallel. These elements were presented in two tutorials, one depicting embryonic folding and the other showing development of the nervous system after neural tube formation. The goal was to enhance the traditional teaching format-lecture combined with printed diagrams, text, and existing computer animations-with customized, guided, Web-based learning modules that surpassed existing resources. To assess module effectiveness, we compared quiz performance of control groups who attended lecture and did not use a supporting module, with study groups who used a module in addition to attending lecture. We also assessed our students' long-term retention of the material, comparing classes who had used the module with students from a previous year that had not seen the module. Our data analysis suggests that students who used a module performed better than those given only traditional resources if they used the module after they were already somewhat familiar with the material. The findings suggest that our modules-and possibly computer-assisted-instruction modules in general-are more useful if used toward the later stages of learning, rather than as an initial resource. Furthermore, our data suggest that the animation aids in long-term retention. Both medical students at the University of Cincinnati and medical faculty from across the country commented favorably on their experiences with the embryonic development modules. Copyright 2008 American Association of Anatomists

Cell differentiation: therapeutical challenges in diabetes.

PubMed

Roche, Enrique; Vicente-Salar, Nestor; Arribas, Maribel; Paredes, Beatriz

2012-01-01

Stem cells, derived from either embryonic or adult tissues, are considered to be potential sources of insulin-secreting cells to be transplanted into type 1 and advanced stages of type 2 diabetic patients. Many laboratories have considered this possibility, resulting in a large amount of published protocols, with a wide degree of complexity among them. Our group was the first to report that it was possible to obtain insulin-secreting cells from mouse embryonic stem cells, proving the feasibility of this new challenge. The same observation was immediately reported using human embryonic stem cells. However, the resulting cell product was not properly characterised, affecting the reproducibility of the protocol by other groups. A more elaborated protocol was developed by Lumelsky and co-workers, demonstrating that neuroectodermal cells could be an alternative source for insulin-producing cells. However, the resulting cells of this protocol produced low amounts of the hormone. This aimed other groups to perform key changes in order to improve the insulin content of the resulting cells. Recently, Baetge's group has published a new protocol based on the knowledge accumulated in pancreatic development. In this protocol, human embryonic stem cells were differentiated into islet-like structures through a five step protocol, emulating the key steps during embryonic development of the endocrine pancreas. The final cell product, however, seemed to be in an immature state, thus further improvement is required. Despite this drawback, the protocol represents the culmination of work performed by different groups and offers new research challenges for the investigators in this exciting field. Concerning adult stem cells, the possibility of identifying pancreatic precursors or of reprogramming extrapancreatic derived cells are key possibilities that may circumvent the problems that appear when using embryonic stem cells, such as immune rejection and tumour formation.

Developmental staging of male murine embryonic gonad by SAGE analysis

PubMed Central

Lee, Tin-Lap; Li, Yunmin; Alba, Diana; Vong, Queenie P.; Wu, Shao-Ming; Baxendale, Vanessa; Rennert, Owen M.; Lau, Yun-Fai Chris; Chan, Wai-Yee

2012-01-01

Despite the identification of key genes such as Sry integral to embryonic gonadal development, the genomic classification and identification of chromosomal activation of this process is still poorly understood. To better understand the genetic regulation of gonadal development, we performed Serial Analysis of Gene Expression (SAGE) to profile the genes and novel transcripts, and an average of 152,000 tags from male embryonic gonads at E10.5 (embryonic day 10.5), E11.5, E12.5, E13.5, E15.5 and E17.5 were analyzed. A total of 275,583 non-singleton tags that do not map to any annotated sequence were identified in the six gonad libraries, and 47,255 tags were mapped to 24,975 annotated sequences, among which 987 sequences were uncharacterized. Utilizing an unsupervised pattern identification technique, we established molecular staging of male gonadal development. Rather than providing a static descriptive analysis, we developed algorithms to cluster the SAGE data and assign SAGE tags to a corresponding chromosomal position; these data are displayed in chromosome graphic format. A prominent increase in global genomic activity from E10.5 to E17.5 was observed. Important chromosomal regions related to the developmental processes were identified and validated based on established mouse models with developmental disorders. These regions may represent markers for early diagnosis for disorders of male gonad development as well as potential treatment targets. PMID:19376482

The plurennial life cycles of the European Tettigoniidae (Insecta: Orthoptera) : 1. The effect of temperature on embryonic development and hatching.

PubMed

Ingrisch, Sigfrid

1986-11-01

The effect of temperature on embryonic development, voltinism, and hatching was studied in the laboratory in eggs of 21 Central and Southeastern European Tettigoniidae species. In most species, the embryo has to arrive at a postkatatrepsis stage prior to the onset of cold to be able to hatch in the following spring. The rate of embryonic development differs: quickly developing species need 4 weeks at 24°C (prior to cold) and almost all eggs hatch after the first cold treatment, slowly developing species would need 8-12 weeks to do the same. In Central Europe, warmth is not enough for the slowly developing species to have an univoltine life cycle, but they could have it in southern Europe. Most species make use of a dormancy sequence to pass successive winters as follows: an initial embryonic dormancy (either quiscence or diapause in embryonic stage 4) and a final diapause in embryonic stage 23/24. Additionally, 3 forms of aestivation or summer dormancy were observed facultatively: an initial diapause in embryonic stage 4 (induced and terminated at 30°C), a median dormancy shortly before or after katatrepsis (at 30°C), and a penultimate diapause in embryonic stage 20 (at 24°C).The life cycles of the European Tettigoniidae species can follow one of 3 types: 1. annual life cycle (no initial embryonic dormancy); 2. annual or biennial depending on whether laid early or late; 3. biennial or many year life cycle (up to 8 years due to a prolonged initial diapause).

Arrested embryonic development: a review of strategies to delay hatching in egg-laying reptiles

PubMed Central

Rafferty, Anthony R.; Reina, Richard D.

2012-01-01

Arrested embryonic development involves the downregulation or cessation of active cell division and metabolic activity, and the capability of an animal to arrest embryonic development results in temporal plasticity of the duration of embryonic period. Arrested embryonic development is an important reproductive strategy for egg-laying animals that provide no parental care after oviposition. In this review, we discuss each type of embryonic developmental arrest used by oviparous reptiles. Environmental pressures that might have directed the evolution of arrest are addressed and we present previously undiscussed environmentally dependent physiological processes that may occur in the egg to bring about arrest. Areas for future research are proposed to clarify how ecology affects the phenotype of developing embryos. We hypothesize that oviparous reptilian mothers are capable of providing their embryos with a level of phenotypic adaptation to local environmental conditions by incorporating maternal factors into the internal environment of the egg that result in different levels of developmental sensitivity to environmental conditions after they are laid. PMID:22438503

Comparison of a teratogenic transcriptome-based predictive test based on human embryonic versus inducible pluripotent stem cells.

PubMed

Shinde, Vaibhav; Perumal Srinivasan, Sureshkumar; Henry, Margit; Rotshteyn, Tamara; Hescheler, Jürgen; Rahnenführer, Jörg; Grinberg, Marianna; Meisig, Johannes; Blüthgen, Nils; Waldmann, Tanja; Leist, Marcel; Hengstler, Jan Georg; Sachinidis, Agapios

2016-12-30

Human embryonic stem cells (hESCs) partially recapitulate early embryonic three germ layer development, allowing testing of potential teratogenic hazards. Because use of hESCs is ethically debated, we investigated the potential for human induced pluripotent stem cells (hiPSCs) to replace hESCs in such tests. Three cell lines, comprising hiPSCs (foreskin and IMR90) and hESCs (H9) were differentiated for 14 days. Their transcriptome profiles were obtained on day 0 and day 14 and analyzed by comprehensive bioinformatics tools. The transcriptomes on day 14 showed that more than 70% of the "developmental genes" (regulated genes with > 2-fold change on day 14 compared to day 0) exhibited variability among cell lines. The developmental genes belonging to all three cell lines captured biological processes and KEGG pathways related to all three germ layer embryonic development. In addition, transcriptome profiles were obtained after 14 days of exposure to teratogenic valproic acid (VPA) during differentiation. Although the differentially regulated genes between treated and untreated samples showed more than 90% variability among cell lines, VPA clearly antagonized the expression of developmental genes in all cell lines: suppressing upregulated developmental genes, while inducing downregulated ones. To quantify VPA-disturbed development based on developmental genes, we estimated the "developmental potency" (D p ) and "developmental index" (D i ). Despite differences in genes deregulated by VPA, uniform D i values were obtained for all three cell lines. Given that the D i values for VPA were similar for hESCs and hiPSCs, D i can be used for robust hazard identification, irrespective of whether hESCs or hiPSCs are used in the test systems.

The thyroid hormone receptor-associated protein TRAP220 is required at distinct embryonic stages in placental, cardiac, and hepatic development.

PubMed

Landles, Christian; Chalk, Sara; Steel, Jennifer H; Rosewell, Ian; Spencer-Dene, Bradley; Lalani, El-Nasir; Parker, Malcolm G

2003-12-01

Recent work indicates that thyroid hormone receptor-associated protein 220 (TRAP220), a subunit of the multiprotein TRAP coactivator complex, is essential for embryonic survival. We have generated TRAP220 conditional null mice that are hypomorphic and express the gene at reduced levels. In contrast to TRAP220 null mice, which die at embryonic d 11.5 (E11.5), hypomorphic mice survive until E13.5. The reduced expression in hypomorphs results in hepatic necrosis, defects in hematopoiesis, and hypoplasia of the ventricular myocardium, similar to that observed in TRAP220 null embryos at an earlier stage. The embryonic lethality of null embryos at E11.5 is due to placental insufficiency. Tetraploid aggregation assays partially rescues embryonic development until E13.5, when embryonic loss occurs due to hepatic necrosis coupled with poor myocardial development as observed in hypomorphs. These findings demonstrate that, for normal placental function, there is an absolute requirement for TRAP220 in extraembryonic tissues at E11.5, with an additional requirement in embryonic tissues for hepatic and cardiovascular development thereafter.

GLUT3 gene expression is critical for embryonic growth, brain development and survival.

PubMed

Carayannopoulos, Mary O; Xiong, Fuxia; Jensen, Penny; Rios-Galdamez, Yesenia; Huang, Haigen; Lin, Shuo; Devaskar, Sherin U

2014-04-01

Glucose is the primary energy source for eukaryotic cells and the predominant substrate for the brain. GLUT3 is essential for trans-placental glucose transport and highly expressed in the mammalian brain. To further elucidate the role of GLUT3 in embryonic development, we utilized the vertebrate whole animal model system of Danio rerio as a tractable system for defining the cellular and molecular mechanisms altered by impaired glucose transport and metabolism related to perturbed expression of GLUT3. The comparable orthologue of human GLUT3 was identified and the expression of this gene abrogated during early embryonic development. In a dose-dependent manner embryonic brain development was disrupted resulting in a phenotype of aberrant brain organogenesis, associated with embryonic growth restriction and increased cellular apoptosis. Rescue of the morphant phenotype was achieved by providing exogenous GLUT3 mRNA. We conclude that GLUT3 is critically important for brain organogenesis and embryonic growth. Disruption of GLUT3 is responsible for the phenotypic spectrum of embryonic growth restriction to demise and neural apoptosis with microcephaly. Copyright © 2014 Elsevier Inc. All rights reserved.

GLUT3 Gene Expression is Critical for Embryonic Growth, Brain Development and Survival

PubMed Central

Carayannopoulos, Mary O.; Xiong, Fuxia; Jensen, Penny; Rios-Galdamez, Yesenia; Huang, Haigen; Lin, Shuo; Devaskar, Sherin U.

2015-01-01

Glucose is the primary energy source for eukaryotic cells and the predominant substrate for the brain. GLUT3 is essential for trans-placental glucose transport and highly expressed in the mammalian brain. To further elucidate the role of GLUT3 in embryonic development, we utilized the vertebrate whole animal model system of Danio rerio as a tractable system for defining the cellular and molecular mechanisms altered by impaired glucose transport and metabolism related to perturbed expression of GLUT3. The comparable orthologue of human GLUT3 was identified and the expression of this gene abrogated during early embryonic development. In a dose-dependent manner embryonic brain development was disrupted resulting in a phenotype of aberrant brain organogenesis, associated with embryonic growth restriction and increased cellular apoptosis. Rescue of the morphant phenotype was achieved by providing exogenous GLUT3 mRNA. We conclude that GLUT3 is critically important for brain organogenesis and embryonic growth. Disruption of GLUT3 is responsible for the phenotypic spectrum of embryonic growth restriction to demise and neural apoptosis with microcephaly. PMID:24529979

Four-hour quantitative real-time polymerase chain reaction-based comprehensive chromosome screening and accumulating evidence of accuracy, safety, predictive value, and clinical efficacy.

PubMed

Treff, Nathan R; Scott, Richard T

2013-03-15

Embryonic comprehensive chromosomal euploidy may represent a powerful biomarker to improve the success of IVF. However, there are a number of aneuploidy screening strategies to consider, including different technologic platforms with which to interrogate the embryonic DNA, and different embryonic developmental stages from which DNA can be analyzed. Although there are advantages and disadvantages associated with each strategy, a series of experiments producing evidence of accuracy, safety, clinical predictive value, and clinical efficacy indicate that trophectoderm biopsy and quantitative real-time polymerase chain reaction (qPCR)-based comprehensive chromosome screening (CCS) may represent a useful strategy to improve the success of IVF. This Biomarkers in Reproductive Medicine special issue review summarizes the accumulated experience with the development and clinical application of a 4-hour blastocyst qPCR-based CCS technology. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Establishment of stem cell lines from nuclear transferred and parthenogenetically activated mouse oocytes for therapeutic cloning.

PubMed

Ju, Jin Young; Park, Chun Young; Gupta, Mukesh Kumar; Uhm, Sang Jun; Paik, Eun Chan; Ryoo, Zae Young; Cho, Youl Hee; Chung, Kil Saeng; Lee, Hoon Taek

2008-05-01

To establish embryonic stem cell lines from nuclear transfer of somatic cell nuclei isolated from the same oocyte donor and from parthenogenetic activation. The study also evaluated the effect of the micromanipulation procedure on the outcome of somatic cell nuclear transfer in mice. Randomized, prospective study. Hospital-based assisted reproductive technology laboratory. F(1) (C57BL/6 x 129P3/J) mice. Metaphase II-stage oocytes were either parthenogenetically activated or nuclear transferred with cumulus cell nuclei or parthenogenetically activated after a sham-manipulation procedure. Embryogenesis and embryonic stem cell establishment. The development rate to morula/blastocyst of nuclear transferred oocytes (27.9% +/- 5.9%) was significantly lower than that of the sham-manipulated (84.1% +/- 5.6%) or parthenogenetic (98.6% +/- 1.4%) groups. A sharp decrease in cleavage potential was obvious in the two- to four-cell transition for the nuclear transferred embryos (79.0% +/- 4.6% and 43.3% +/- 5.0%), implying incomplete nuclear reprogramming in arrested oocytes. However, the cleavage, as well as the development rate, of parthenogenetic and sham-manipulated groups did not differ significantly. The embryonic stem cell line establishment rate was higher from parthenogenetically activated oocytes (15.7%) than nuclear transferred (4.3%) or sham-manipulated oocytes (12.5%). Cell colonies from all groups displayed typical morphology of mice embryonic stem cells and could be maintained successfully with undifferentiated morphology after continuous proliferation for more than 120 passages still maintaining normal karyotype. All these cells were positive for mice embryonic stem cell markers such as Oct-4 and SSEA-1 based on immunocytochemistry and reverse transcriptase-polymerase chain reaction. The clonal origin of the ntES cell line and the parthenogenetic embryonic stem cell lines were confirmed by polymerase chain reaction analysis of the polymorphic markers. Blastocyst injection experiments demonstrated that these lines contributed to resulting chimeras and are germ-line competent. We report the establishment of ntES cell lines from somatic cells isolated from same individual. Our data also suggest that embryo micromanipulation procedure during the nuclear transfer procedure influences the developmental ability and embryonic stem cell establishment rate of nuclear transferred embryos.

VE-cadherin expression allows identification of a new class of hematopoietic stem cells within human embryonic liver.

PubMed

Oberlin, Estelle; Fleury, Maud; Clay, Denis; Petit-Cocault, Laurence; Candelier, Jean-Jacques; Mennesson, Benoît; Jaffredo, Thierry; Souyri, Michèle

2010-11-25

Edification of the human hematopoietic system during development is characterized by the production of waves of hematopoietic cells separated in time, formed in distinct embryonic sites (ie, yolk sac, truncal arteries including the aorta, and placenta). The embryonic liver is a major hematopoietic organ wherein hematopoietic stem cells (HSCs) expand, and the future, adult-type, hematopoietic cell hierarchy becomes established. We report herein the identification of a new, transient, and rare cell population in the human embryonic liver, which coexpresses VE-cadherin, an endothelial marker, CD45, a pan-hematopoietic marker, and CD34, a common endothelial and hematopoietic marker. This population displays an outstanding self-renewal, proliferation, and differentiation potential, as detected by in vitro and in vivo hematopoietic assays compared with its VE-cadherin negative counterpart. Based on VE-cadherin expression, our data demonstrate the existence of 2 phenotypically and functionally separable populations of multipotent HSCs in the human embryo, the VE-cadherin(+) one being more primitive than the VE-cadherin(-) one, and shed a new light on the hierarchical organization of the embryonic liver HSC compartment.

[Tones and being tuned. Suggestions for the common origins of music therapy and hypnotherapy].

PubMed

Vas, József Pál

2013-01-01

Sound vibrations are viewed to play an important role in embryonic development. Before the cochlea evolves, the haptic and mechanic skin-receptors detect the amniotic fluid's pressure-waves produced by sounds in uterus. Touching and hearing are seen as primordial and the most relevant stimuli both of mother-fetus attunement and development of fetal nervous system. Man is attuned to environmental stimuli, mainly to human speaking since the embryonic period. Attunement is secured by energy zones (chakras) circling around body. It is considered to be base of our music capacity. Origin of hypnotic susceptibility is viewed as being in embryonic period as well. Movements, experiences supposed, bonding and communication patterns of both of fetus and hypnotized person are suggested to show similarities. Prenatal audio-somatosensory stimulating program facilitates newborn babies' cognitive, emotional and bonding capacities. As a matter of fact, by virtue of regressive fetus-like experiences, hypnotherapy contributes to the restart of personality development halted by trauma.

Stem cell maintenance by manipulating signaling pathways: past, current and future

PubMed Central

Chen, Xi; Ye, Shoudong; Ying, Qi-Long

2015-01-01

Pluripotent stem cells only exist in a narrow window during early embryonic development, whereas multipotent stem cells are abundant throughout embryonic development and are retainedin various adult tissues and organs. While pluripotent stem cell lines have been established from several species, including mouse, rat, and human, it is still challenging to establish stable multipotent stem cell lines from embryonic or adult tissues. Based on current knowledge, we anticipate that by manipulating extrinsic and intrinsic signaling pathways, most if not all types of stem cells can be maintained in a long-term culture. In this article, we summarize current culture conditions established for the long-term maintenance of authentic pluripotent and multipotent stem cells and the signaling pathways involved. We also discuss the general principles of stem cell maintenance and propose several strategies on the establishment of novel stem cell lines through manipulation of signaling pathways. [BMB Reports 2015; 48(12): 668-676] PMID:26497581

Brief Embryonic Strychnine Exposure in Zebrafish Causes Long-Term Adult Behavioral Impairment with Indications of Embryonic Synaptic Changes

PubMed Central

Roy, Nicole M.; Arpie, Brianna; Lugo, Joseph; Linney, Elwood; Levin, Edward D.; Cerutti, Daniel

2015-01-01

Zebrafish provide a powerful model of the impacts of embryonic toxicant exposure on neural development that may result in long-term behavioral dysfunction. In this study, zebrafish embryos were treated with 1.5 mM strychnine for short embryonic time windows to induce transient changes in inhibitory neural signaling, and were subsequently raised in untreated water until adulthood. PCR analysis showed indications that strychnine exposure altered expression of some genes related to glycinergic, GABAergic and glutamatergic neuronal synapses during embryonic development. In adulthood, treated fish showed significant changes in swimming speed and tank diving behavior compared to controls. Taken together, these data show that a short embryonic exposure to a neurotoxicant can alter development of neural synapses and lead to changes in adult behavior. PMID:23022260

Brief embryonic strychnine exposure in zebrafish causes long-term adult behavioral impairment with indications of embryonic synaptic changes.

PubMed

Roy, Nicole M; Arpie, Brianna; Lugo, Joseph; Linney, Elwood; Levin, Edward D; Cerutti, Daniel

2012-01-01

Zebrafish provide a powerful model of the impacts of embryonic toxicant exposure on neural development that may result in long-term behavioral dysfunction. In this study, zebrafish embryos were treated with 1.5mM strychnine for short embryonic time windows to induce transient changes in inhibitory neural signaling, and were subsequently raised in untreated water until adulthood. PCR analysis showed indications that strychnine exposure altered expression of some genes related to glycinergic, GABAergic and glutamatergic neuronal synapses during embryonic development. In adulthood, treated fish showed significant changes in swimming speed and tank diving behavior compared to controls. Taken together, these data show that a short embryonic exposure to a neurotoxicant can alter development of neural synapses and lead to changes in adult behavior. Copyright © 2012 Elsevier Inc. All rights reserved.

First trimester size charts of embryonic brain structures.

PubMed

Gijtenbeek, M; Bogers, H; Groenenberg, I A L; Exalto, N; Willemsen, S P; Steegers, E A P; Eilers, P H C; Steegers-Theunissen, R P M

2014-02-01

Can reliable size charts of human embryonic brain structures be created from three-dimensional ultrasound (3D-US) visualizations? Reliable size charts of human embryonic brain structures can be created from high-quality images. Previous studies on the visualization of both the cavities and the walls of the brain compartments were performed using 2D-US, 3D-US or invasive intrauterine sonography. However, the walls of the diencephalon, mesencephalon and telencephalon have not been measured non-invasively before. Last-decade improvements in transvaginal ultrasound techniques allow a better visualization and offer the tools to measure these human embryonic brain structures with precision. This study is embedded in a prospective periconceptional cohort study. A total of 141 pregnancies were included before the sixth week of gestation and were monitored until delivery to assess complications and adverse outcomes. For the analysis of embryonic growth, 596 3D-US scans encompassing the entire embryo were obtained from 106 singleton non-malformed live birth pregnancies between 7(+0) and 12(+6) weeks' gestational age (GA). Using 4D View (3D software) the measured embryonic brain structures comprised thickness of the diencephalon, mesencephalon and telencephalon, and the total diameter of the diencephalon and mesencephalon. Of 596 3D scans, 161 (27%) high-quality scans of 79 pregnancies were eligible for analysis. The reliability of all embryonic brain structure measurements, based on the intra-class correlation coefficients (ICCs) (all above 0.98), was excellent. Bland-Altman plots showed moderate agreement for measurements of the telencephalon, but for all other measurements the agreement was good. Size charts were constructed according to crown-rump length (CRL). The percentage of high-quality scans suitable for analysis of these brain structures was low (27%).  The size charts of human embryonic brain structures can be used to study normal and abnormal development of brain development in future. Also, the effects of periconceptional maternal exposures, such as folic acid supplement use and smoking, on human embryonic brain development can be a topic of future research. This study was supported by the Department of Obstetrics and Gynaecology of the Erasmus University Medical Center. M.G. was supported by an additional grant from the Sophia Foundation for Medical Research (SSWO grant number 644). No competing interests are declared.

Diploid, but not haploid, human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes

PubMed Central

Fan, Yong; Li, Rong; Huang, Jin; Yu, Yang; Qiao, Jie

2013-01-01

Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells. PMID:23255130

Three-dimensional microCT imaging of murine embryonic development from immediate post-implantation to organogenesis: application for phenotyping analysis of early embryonic lethality in mutant animals.

PubMed

Ermakova, Olga; Orsini, Tiziana; Gambadoro, Alessia; Chiani, Francesco; Tocchini-Valentini, Glauco P

2018-04-01

In this work, we applied three-dimensional microCT imaging to study murine embryogenesis in the range from immediate post-implantation period (embryonic day 5.5) to mid-gestation (embryonic day 12.5) with the resolution up to 1.4 µm/voxel. Also, we introduce an imaging procedure for non-invasive volumetric estimation of an entire litter of embryos within the maternal uterine structures. This method allows for an accurate, detailed and systematic morphometric analysis of both embryonic and extra-embryonic components during embryogenesis. Three-dimensional imaging of unperturbed embryos was performed to visualize the egg cylinder, primitive streak, gastrulation and early organogenesis stages of murine development in the C57Bl6/N mouse reference strain. Further, we applied our microCT imaging protocol to determine the earliest point when embryonic development is arrested in a mouse line with knockout for tRNA splicing endonuclease subunit Tsen54 gene. Our analysis determined that the embryonic development in Tsen54 null embryos does not proceed beyond implantation. We demonstrated that application of microCT imaging to entire litter of non-perturbed embryos greatly facilitate studies to unravel gene function during early embryogenesis and to determine the precise point at which embryonic development is arrested in mutant animals. The described method is inexpensive, does not require lengthy embryos dissection and can be applicable for detailed analysis of mutant mice at laboratory scale as well as for high-throughput projects.

Delayed embryonic development in the Indian short-nosed fruit bat, Cynopterus sphinx.

PubMed

Meenakumari, Karukayil J; Krishna, Amitabh

2005-01-01

The unusual feature of the breeding cycle of Cynopterus sphinx at Varanasi is the significant variation in gestation length of the two successive pregnancies of the year. The aim of this study was to investigate whether the prolongation of the first pregnancy in C. sphinx is due to delayed embryonic development. The first (winter) pregnancy commences in late October and lasts until late March and has a gestation period of about 150 days. The second (summer) pregnancy commences in April and lasts until the end of July or early August with a gestation period of about 125 days. Changes in the size and weight of uterine cornua during the two successive pregnancies suggest retarded embryonic growth during November and December. Histological analysis during the period of retarded embryonic development in November and December showed a slow gastrulation process. The process of amniogenesis was particularly slow. When the embryos attained the early primitive streak stage, their developmental rate suddenly increased considerably. During the summer pregnancy, on the other hand, the process of gastrulation was much faster and proceeded quickly. A comparison of the pattern of embryonic development for 4 consecutive years consistently showed retarded or delayed embryonic development during November and December. The time of parturition and post-partum oestrus showed only a limited variation from 1 year to another. This suggests that delayed embryonic development in C. sphinx may function to synchronize parturition among females. The period of delayed embryonic development in this species clearly coincides with the period of fat deposition. The significance of this correlation warrants further investigation.

Expression pattern of pluripotent markers in different embryonic developmental stages of buffalo (Bubalus bubalis) embryos and putative embryonic stem cells generated by parthenogenetic activation.

PubMed

Singh, Karn P; Kaushik, Ramakant; Garg, Veena; Sharma, Ruchi; George, Aman; Singh, Manoj K; Manik, Radhey S; Palta, Prabhat; Singla, Suresh K; Chauhan, Manmohan S

2012-12-01

In this study, we describe the production of buffalo parthenogenetic blastocysts and subsequent isolation of parthenogenetic embryonic stem cell (PGESC)-like cells. PGESC colonies exhibited dome-shaped morphology and were clearly distinguishable from the feeder layer cells. Different stages of development of parthenogenetic embryos and derived embryonic stem cell (ESC)-like cells expressed key ESC-specific markers, including OCT-4, NANOG, SOX-2, FOXD3, REX-1, STAT-3, TELOMERASE, NUCLEOSTEMIN, and cMYC. Immunofluorescence-based studies revealed that the PGESCs were positive for surface-based pluripotent markers, viz., SSEA-3, SSEA-4, TRA 1-80, TRA 1-60, CD-9, and CD-90 and exhibited high alkaline phosphatase (ALP) activity. PGEC cell-like cells formed embryoid body (EB)-like structures in hanging drop cultures and when cultured for extended period of time spontaneously differentiated into derivatives of three embryonic germ layers as confirmed by RT-PCR for ectodermal (CYTOKERATIN8, NF-68), mesodermal (MSX1, BMP-4, ASA), and endodermal markers (AFP, HNF-4, GATA-4). Differentiation of PGESCs toward the neuronal lineage was successfully directed by supplementation of serum-containing media with retinoic acid. Our results indicate that the isolated ESC-like cells from parthenogenetic blastocyst hold properties of ESCs and express markers of pluripotency. The pluripotency markers were also expressed by early cleavage-stage of buffalo embryos.

Mitochondrial functionality in reproduction: from gonads and gametes to embryos and embryonic stem cells.

PubMed

Ramalho-Santos, João; Varum, Sandra; Amaral, Sandra; Mota, Paula C; Sousa, Ana Paula; Amaral, Alexandra

2009-01-01

Mitochondria are multitasking organelles involved in ATP synthesis, reactive oxygen species (ROS) production, calcium signalling and apoptosis; and mitochondrial defects are known to cause physiological dysfunction, including infertility. The goal of this review was to identify and discuss common themes in mitochondrial function related to mammalian reproduction. The scientific literature was searched for studies reporting on the several aspects of mitochondrial activity in mammalian testis, sperm, oocytes, early embryos and embryonic stem cells. ATP synthesis and ROS production are the most discussed aspects of mitochondrial function. Metabolic shifts from mitochondria-produced ATP to glycolysis occur at several stages, notably during gametogenesis and early embryo development, either reflecting developmental switches or substrate availability. The exact role of sperm mitochondria is especially controversial. Mitochondria-generated ROS function in signalling but are mostly described when produced under pathological conditions. Mitochondria-based calcium signalling is primarily important in embryo activation and embryonic stem cell differentiation. Besides pathologically triggered apoptosis, mitochondria participate in apoptotic events related to the regulation of spermatogonial cell number, as well as gamete, embryo and embryonic stem cell quality. Interestingly, data from knock-out (KO) mice is not always straightforward in terms of expected phenotypes. Finally, recent data suggests that mitochondrial activity can modulate embryonic stem cell pluripotency as well as differentiation into distinct cellular fates. Mitochondria-based events regulate different aspects of reproductive function, but these are not uniform throughout the several systems reviewed. Low mitochondrial activity seems a feature of 'stemness', being described in spermatogonia, early embryo, inner cell mass cells and embryonic stem cells.

GLUCOCORTICOID RECEPTOR EXPRESSION DURING THE DEVELOPMENT OF THE EMBRYONIC MOUSE SECONDARY PALATE

EPA Science Inventory

Glucocorticoids are important regulators of embryonic growth and development. hese effects are mediated through glucocorticoid receptors (GR) which bind to glucocorticoid response elements upstream of regulated genes. his study examines the expression of GR and GR mRNA in embryon...

Luteal cell steroidogenesis in relation to delayed embryonic development in the Indian short-nosed fruit bat, Cynopterus sphinx.

PubMed

Meenakumari, Karukayil J; Banerjee, Arnab; Krishna, Amitabh

2009-01-01

The primary aim of this study was to determine the possible cause of slow or delayed embryonic development in Cynopterus sphinx by investigating morphological and steroidogenic changes in the corpus luteum (CL) and circulating hormone concentrations during two pregnancies of a year. This species showed delayed post-implantational embryonic development during gastrulation of the first pregnancy. Morphological features of the CL showed normal luteinization during both pregnancies. The CL did not change significantly in luteal cell size during the delay period of the first pregnancy as compared with the second pregnancy. The circulating progesterone and 17beta-estradiol concentrations were significantly lower during the period of delayed embryonic development as compared with the same stage of embryonic development during the second pregnancy. We also showed a marked decline in the activity of 3beta-hydroxysteroid dehydrogenase, P450 side chain cleavage enzyme, and steroidogenic acute regulatory peptide in the CL during the delay period. This may cause low circulating progesterone and estradiol synthesis and consequently delay embryonic development. What causes the decrease in steroidogenic factors in the CL during the period of delayed development in C. sphinx is under investigation.

Jaw muscle development as evidence for embryonic repatterning in direct-developing frogs.

PubMed Central

Hanken, J; Klymkowsky, M W; Alley, K E; Jennings, D H

1997-01-01

The Puerto Rican direct-developing frog Eleutherodactylus coqui (Leptodactylidae) displays a novel mode of jaw muscle development for anuran amphibians. Unlike metamorphosing species, several larval-specific features never form in E. coqui; embryonic muscle primordia initially assume an abbreviated, mid-metamorphic configuration that is soon remodelled to form the adult morphology before hatching. Also lacking are both the distinct population of larval myofibres and the conspicuous, larval-to-adult myofibre turnover that are characteristic of muscle development in metamorphosing species. These modifications are part of a comprehensive alteration in embryonic cranial patterning that has accompanied life history evolution in this highly speciose lineage. Embryonic 'repatterning' in Eleutherodactylus may reflect underlying developmental mechanisms that mediate the integrated evolution of complex structures. Such mechanisms may also facilitate, in organisms with a primitively complex life cycle, the evolutionary dissociation of embryonic, larval, and adult features. PMID:9332017

Based serum metabolomics analysis reveals simultaneous interconnecting changes during chicken embryonic development.

PubMed

Peng, M L; Li, S N; He, Q Q; Zhao, J L; Li, L L; Ma, H T

2018-05-28

Metabolic disorder is a major health problem and is associated with a number of metabolic diseases. Due to native hyperglycaemia and resistance to exogenous insulin, chickens as a model had used in the studies of adipose tissue biology, metabolism and obesity. But no detailed information is available about the comprehensive changes of serum metabolites at different stages of chicken embryonic development. This study employed LC/MS-QTOF to determine the changes of major functional metabolites at incubation day 14 (E14d), 19 (E19d) and hatching day 1 (H1d), and the associated pathways of differential metabolites during chicken embryonic development were analysed using Metabolite Set Enrichment Analysis method. Results showed that 39 metabolites were significantly changed from E14d to E19d and 68 metabolites were significantly altered from E19d to H1d in chicken embryos. Protein synthesis was promoted by increasing the concentrations of L-glutamine and threonine, and gonadal development was promoted through increasing oestrone content from E14d to E19d in chicken embryos, which indicated that serum glutamine, threonine and oestrone contents may be considered as the candidate indicators for assessment of early embryonic development. 2-oxoglutaric acid mainly contributed to enhancing the citric cycle, and it plays an important role in improving the growth of chicken embryos at the late development; the decreasing of L-glutamine, L-isoleucine and L-leucine contents from E19d to H1d in chicken embryonic development implied their possible functions as the feed additive during early posthatch period of broiler chickens to satisfy the growth. These results provided insights into understand the roles of serum metabolites at different developmental stages of chicken embryos, it also provides available information for chicken as a model to study metabolic disease or human obesity. © 2018 Blackwell Verlag GmbH.

Parthenogenesis in unfertilized eggs of Coturnix chinensis, the Chinese painted quail, and the effect of egg clutch position on embryonic development.

PubMed

Parker, H M; McDaniel, C D

2009-04-01

Parthenogenesis, embryonic development of an unfertilized egg, was studied for many years in turkeys. In fact, as many as 49% of unfertilized Beltsville Small White turkey eggs develop embryos. However, no research exists on parthenogenesis in quail. The Chinese painted quail is a close relative of the more common Japanese quail and, unlike turkeys or chickens, the small Chinese painted quail reaches sexual maturity rapidly, making it a great candidate for further research on parthenogenesis. Obviously, a better understanding of avian parthenogenesis should increase our knowledge of avian fertilization and early embryonic development. Therefore, we determined if unfertilized Chinese painted quail hens produce embryos. Second, we explored the possibility that position of the egg within the clutch influences parthenogenesis. When initial secondary sexual plumage was apparent at 4 wk of age, male chicks were separated from females to prevent fertilization. Hens were placed in individual cages near sexual maturity, at approximately 6 wk of age. Individual eggs were collected daily and labeled with hen number and date. Eggs were stored for 0 to 3 d at 20 degrees C before incubation at 37.5 degrees C. After 10 d of incubation, approximately 4,000 eggs from 300 laying hens were examined for embryonic development under a magnifying lamp. On average, 4.8% of the unfertilized eggs contained an abortive form of embryonic development consisting of undifferentiated cells and unorganized membranes. Approximately 27% of the laying hens produced at least 1 egg with parthenogenic development. However, about 10% (30) of these hens exhibited a predisposition for parthenogenesis by producing 2 or more unfertilized eggs with embryonic development. Twenty percent of the eggs from 2 hens produced embryonic development. Additionally, the first egg laid in a clutch was most likely to produce embryonic development, with a steady decline in the percentage of eggs with embryonic development as position in the clutch increased. In conclusion, the Chinese painted quail does exhibit parthenogenesis and clutch position influences the rate of naturally occurring parthenogenesis.

Evaluation of 309 environmental chemicals using a mouse embryonic stem cell adherent cell differentiation and cytotoxicity assay

EPA Science Inventory

The vast landscape of environmental chemicals has motivated the need for alternative methods to traditional whole-animal bioassays in toxicity testing. Embryonic stem (ES) cells provide an in vitro model of embryonic development and an alternative method for assessing development...

Relationship between delayed embryonic development and metabolic factors and fat deposition in fruit bat Cynopterus sphinx.

PubMed

Banerjee, Arnab; Meenakumari, K J; Krishna, Amitabh

2007-01-01

The present study was undertaken in the fruit bat Cynopterus sphinx, which breeds twice in quick succession at Varanasi, India. Its gestation period varies significantly in the two successive pregnancies of the year owing to delayed embryonic development during the first (winter) pregnancy. The primary aim of the present study was to determine the role of metabolic factors in delayed embryonic development in the fruit bat C. sphinx. Variation in bodyweight, fat deposition, oxygen (O(2)) consumption rate, basal metabolic rate (BMR), body temperature (Tb) and hepatic succinate dehydrogenase (SDH) activity, along with circulating levels of thyroid hormones (tri-iodothyronine and thyroxine), were examined as metabolic factors during the two successive pregnancies in C. sphinx. The increase in bodyweight observed in November was due to accumulation of white adipose tissue in the posterior abdominal region. A significant decline in O(2) consumption rate, BMR, Tb and SDH activity was found in early winter in November-December, which coincides closely with the period of fat accumulation and with the period of delayed embryonic development in C. sphinx. A significantly higher O(2) consumption rate, BMR, Tb and SDH activity was noted during the second pregnancy in, when embryonic development was relatively faster. Thyroid hormone levels were high during the period of embryonic delay compared with levels during the remaining months. The results of the present study suggest that the delayed embryonic development in C. sphinx during early winter may be due to a low O(2) consumption rate, BMR, Tb and SDH activity in November-December. The energy saved by suppressing embryonic development in this species may be advantageous for fat accumulation. Increased thyroid hormone levels during the early winter period might facilitate fat accumulation in C. sphinx.

Znrg, a novel gene expressed mainly in the developing notochord of zebrafish.

PubMed

Zhou, Yaping; Xu, Yan; Li, Jianzhen; Liu, Yao; Zhang, Zhe; Deng, Fengjiao

2010-06-01

The notochord, a defining characteristic of the chordate embryo is a critical midline structure required for axial skeletal formation in vertebrates, and acts as a signaling center throughout embryonic development. We utilized the digital differential display program of the National Center for Biotechnology Information, and identified a contig of expressed sequence tags (no. Dr. 83747) from the zebrafish ovary library in Genbank. Full-length cDNA of the identified gene was cloned by 5'- and 3'- RACE, and the resulting sequence was confirmed by polymerase chain reaction and sequencing. The cDNA clone contains 2,505 base pairs and encodes a novel protein of 707 amino acids that shares no significant homology with any known proteins. This gene was expressed in mature oocytes and at the one-cell stage, and persisted until the 5th day of development, as determined by RT-PCR. Transcripts were detected by whole-mount RNA in situ hybridization from the two-cell stage to 72 h of embryonic development. This gene was uniformly distributed from the cleavage stage up to the blastula stage. During early gastrulation, it was present in the dorsal region, and became restricted to the notochord and pectoral fin at 48 and 72 h of embryonic development. Based on its abundance in the notochord, we hypothesized that the novel gene may play an important role in notochord development in zebrafish; we named this gene, zebrafish notochord-related gene, or znrg.

Embryonic development rates of northern grasshoppers (Orthoptera: Acrididae): implications for climate change and habitat management

USDA-ARS?s Scientific Manuscript database

Temperature-dependent rates of embryonic development are a primary determinant of the life cycle of many species of grasshoppers which, in cold climates, spend two winters in the egg stage. Knowledge of embryonic developmental rates is important for an assessment of the effects of climate change and...

Role of leptin in delayed embryonic development in the Indian short-nosed fruit bat, Cynopterus sphinx.

PubMed

Banerjee, A; Meenakumari, K J; Krishna, A

2010-08-01

An adiposity-associated rise in leptin occurs at the time of delayed embryonic development in Cynopterus sphinx. The aim of present study was to examine the mechanism by which leptin may inhibit progesterone, and therefore could be responsible for delayed development. The study showed a significant increase in circulating leptin level during the period of increased fat accumulation, which coincided with significant decrease in serum progesterone level and delayed embryonic development in C. sphinx. The study showed increased Ob-R expression in the corpus luteum and in the utero-embryonic unit during the period of delayed embryonic development. The in vitro study showed suppressive effect of leptin on progesterone synthesis. The effect of high dose of leptin on ovarian steroidogenesis was found to be mediated through decreased expression of StAR and LH-R proteins in the ovary. The treatment with leptin caused increased expression of STAT 3 and iNOS proteins in the ovary, which correlated with decreased expression of StAR protein in the ovary. The inhibitory effects of leptin on progesterone synthesis in the ovary are thus mediated through STAT 3 and iNOS-NO signaling pathways. This study further demonstrated low expression of PCNA coinciding with the increased concentration of the leptin receptor in the utero-embryonic unit and high circulating leptin level during November. In conclusion, adiposity associated increased leptin level during November-December might play role in suppressing progesterone synthesis in the corpus luteum as well as suppressing the rate of cell-proliferation in the utero-embryonic unit thereby causing delayed embryonic development in C. sphinx. Copyright 2010 Elsevier Inc. All rights reserved.

Periods and stages of the prenatal development of the domestic cat.

PubMed

Knospe, C

2002-02-01

Twenty-two stages of the prenatal development of the domestic cat are described for intraspecies comparison in embryological studies. These are assigned to the 15 embryonal periods based on the Nomina Embryologica Veterinaria to make the interspecies comparison possible.

The business of human embryonic stem cell research and an international analysis of relevant laws.

PubMed

De Trizio, Ella; Brennan, Christopher S

2004-01-01

Few sciences have held out such therapeutic promise and correspondingly stirred so much controversy in countries throughout the world as the developing science surrounding human embryonic stem cells. Since the first reported development of several lines of human embryonic stem cells in 1988, many governments around the world have attempted to address the thorny ethical issues raised by human embryonic stem cell research by the passage of laws. In some cases these laws have directly regulated governmental funding of the science; in other cases they have created a legal environment that has either encouraged or discouraged both governmental and private funding of the science. This article first differentiates human embryonic stem cells from other types of stem cells and frames the ethical controversy surrounding human embryonic stem cell research, then surveys laws governing human embryonic stem cell research in various scientifically advanced countries located throughout the Pacific Rim, Europe and North America and explains the impact these laws have had on governmental and private funding of human embryonic stem cell research.

Non-invasive long-term fluorescence live imaging of Tribolium castaneum embryos.

PubMed

Strobl, Frederic; Stelzer, Ernst H K

2014-06-01

Insect development has contributed significantly to our understanding of metazoan development. However, most information has been obtained by analyzing a single species, the fruit fly Drosophila melanogaster. Embryonic development of the red flour beetle Tribolium castaneum differs fundamentally from that of Drosophila in aspects such as short-germ development, embryonic leg development, extensive extra-embryonic membrane formation and non-involuted head development. Although Tribolium has become the second most important insect model organism, previous live imaging attempts have addressed only specific questions and no long-term live imaging data of Tribolium embryogenesis have been available. By combining light sheet-based fluorescence microscopy with a novel mounting method, we achieved complete, continuous and non-invasive fluorescence live imaging of Tribolium embryogenesis at high spatiotemporal resolution. The embryos survived the 2-day or longer imaging process, developed into adults and produced fertile progeny. Our data document all morphogenetic processes from the rearrangement of the uniform blastoderm to the onset of regular muscular movement in the same embryo and in four orientations, contributing significantly to the understanding of Tribolium development. Furthermore, we created a comprehensive chronological table of Tribolium embryogenesis, integrating most previous work and providing a reference for future studies. Based on our observations, we provide evidence that serosa window closure and serosa opening, although deferred by more than 1 day, are linked. All our long-term imaging datasets are available as a resource for the community. Tribolium is only the second insect species, after Drosophila, for which non-invasive long-term fluorescence live imaging has been achieved. © 2014. Published by The Company of Biologists Ltd.

Effect of temperature on embryonic development of Melanotaenia boesemani (Allen and Cross, 1982).

PubMed

Radael, Marcella Costa; Cardoso, Leonardo Demier; de Andrade, Dalcio Ricardo; Ferreira, André Veloso; da Cruz Mattos, Douglas; Vidal, Manuel Vazquez

2016-04-01

The present study aimed to provide data on the time required for Melanotaenia boesemani to complete embryonic development, and to investigate the influence that incubation at different temperatures caused in this species. The effects of temperature on the time and hatching rate are presented, as well as information related to embryonic development stages. After fertilization, the eggs were kept in incubators at 23, 26, 29 or 32°C and observed at predetermined times until the moment of hatching. Stages of development were identified and classified according to morphological and physiological characteristics. Oil droplets were visualized inside the eggs as well as filament adhesion present at the chorion. Embryonic development was similar to that observed in other species of the genus Melanotaenia with hatching and faster development in higher temperatures.

Cardiac Progenitor Cells and Bone Marrow-Derived Very Small Embryonic-Like Stem Cells for Cardiac Repair After Myocardial Infarction

PubMed Central

Tang, Xian-Liang; Rokosh, D. Gregg; Guo, Yiru; Bolli, Roberto

2010-01-01

Heart failure after myocardial infarction (MI) continues to be the most prevalent cause of morbidity and mortality worldwide. Although pharmaceutical agents and interventional strategies have contributed greatly to therapy, new and superior treatment modalities are urgently needed given the overall disease burden. Stem cell-based therapy is potentially a promising strategy to lead to cardiac repair after MI. An array of cell types has been explored in this respect, including skeletal myoblasts, bone marrow (BM)-derived stem cells, embryonic stem cells, and more recently, cardiac progenitor cells (CPCs). Recently studies have obtained evidence that transplantation of CPCs or BM-derived very small embryonic-like stem cells can improve cardiac function and alleviate cardiac remodeling, supporting the potential therapeutic utility of these cells for cardiac repair. This report summarizes the current data from those studies and discusses the potential implication of these cells in developing clinically-relevant stem cell-based therapeutic strategies for cardiac regeneration. PMID:20081317

Embryonic blood-cerebrospinal fluid barrier formation and function

PubMed Central

Bueno, David; Parvas, Maryam; Hermelo, Ismaïl; Garcia-Fernàndez, Jordi

2014-01-01

During embryonic development and adult life, brain cavities and ventricles are filled with cerebrospinal fluid (CSF). CSF has attracted interest as an active signaling medium that regulates brain development, homeostasis and disease. CSF is a complex protein-rich fluid containing growth factors and signaling molecules that regulate multiple cell functions in the central nervous system (CNS). The composition and substance concentrations of CSF are tightly controlled. In recent years, it has been demonstrated that embryonic CSF (eCSF) has a key function as a fluid pathway for delivering diffusible signals to the developing brain, thus contributing to the proliferation, differentiation and survival of neural progenitor cells, and to the expansion and patterning of the brain. From fetal stages through to adult life, CSF is primarily produced by the choroid plexus. The development and functional activities of the choroid plexus and other blood–brain barrier (BBB) systems in adults and fetuses have been extensively analyzed. However, eCSF production and control of its homeostasis in embryos, from the closure of the anterior neuropore when the brain cavities become physiologically sealed, to the formation of the functional fetal choroid plexus, has not been studied in as much depth and remains open to debate. This review brings together the existing literature, some of which is based on experiments conducted by our research group, concerning the formation and function of a temporary embryonic blood–CSF barrier in the context of the crucial roles played by the molecules in eCSF. PMID:25389383

Morphological and histomorphological structures of testes and ovaries in early developmental stages of the silkworm, Bombyx mori.

PubMed

Sakai, Hiroki; Kirino, Yohei; Katsuma, Susumu; Aoki, Fugaku; Suzuki, Masataka G

2016-01-01

The gonad develops as a testis in male or an ovary in female. In the silkworm, B. mori , little is known about testis and ovary in the embryonic stages and early larval stages. In this study, we performed morphological and histomorphological observations of ovaries and testes from the late embryonic stage to the 1st instar larval stage. Results obtained with lack of accurate information on sex of examined individuals may be misleading, thus we performed phenotypic observations of gonads by utilizing sex-limited strain that enables us to easily discriminate female embryos from male ones based on those egg colors. In testis, four testicular follicles were clearly observed in the testis at the first instar larval stage, and boundary layers were formed between the testicular follicles. At the late embryonic stage, the testis consisted of four testicular follicles, while the boundary layers were still obscure. In ovary, four ovarioles were easily recognizable in the ovary at the first instar larval stage, and boundary layers were formed between the ovarioles. However, in the late embryonic stage, it was quite difficult to identify four ovarioles. Morphological characteristics were almost similar between testis and ovary in early developmental stages. Our present study demonstrates that the most reliable difference between testis and ovary in early developmental stages is the attaching point of the duct. Formation and development of the duct may be sensitive to the sex-determining signal and display sexual dimorphism in early embryonic stages.

Derivation of Multipotent Mesenchymal Precursors from Human Embryonic Stem Cells

PubMed Central

Barberi, Tiziano; Willis, Lucy M; Socci, Nicholas D; Studer, Lorenz

2005-01-01

Background Human embryonic stem cells provide access to the earliest stages of human development and may serve as a source of specialized cells for regenerative medicine. Thus, it becomes crucial to develop protocols for the directed differentiation of embryonic stem cells into tissue-restricted precursors. Methods and Findings Here, we present culture conditions for the derivation of unlimited numbers of pure mesenchymal precursors from human embryonic stem cells and demonstrate multilineage differentiation into fat, cartilage, bone, and skeletal muscle cells. Conclusion Our findings will help to elucidate the mechanism of mesoderm specification during embryonic stem cell differentiation and provide a platform to efficiently generate specialized human mesenchymal cell types for future clinical applications. PMID:15971941

Elevated temperature enhances normal early embryonic development in the coral Platygyra acuta under low salinity conditions

NASA Astrophysics Data System (ADS)

Chui, Apple Pui Yi; Ang, Put

2015-06-01

To better understand the possible consequences of climate change on reef building scleractinian corals in a marginal environment, laboratory experiments were conducted to examine the interactive effects of changes in salinity and temperature on percent fertilization success and early embryonic development of the coral Platygyra acuta. In the present study, a salinity of 24 psu (ambient 32 psu) reduced fertilization success by 60 %. Normal embryonic development was reduced by >80 % at 26 psu (ambient 33 psu) with 100 % abnormal development at 22 psu under ambient temperature. Elevated temperature (+3 °C) above the ambient spawning temperature did not show any negative effects on fertilization success. However, there was a trend for more abnormal embryos to develop at elevated temperature in the 2 d of the spawning event. The interactive effects between salinity and temperature are statistically significant only on normal embryonic development of P. acuta, but not on its fertilization success. Salinity was revealed to be the main factor affecting both fertilization success and normal embryonic development. Interestingly, the much lower fertilization success (76 %) observed in the second day of spawning (Trial 2) under ambient temperature recovered to 99 % success under elevated (+3 °C) temperature conditions. Moreover, elevated temperature enhanced normal early embryonic development under lowered salinity (26 psu). This antagonistic interactive effect was consistently observed in two successive nights of spawning. Overall, our results indicate that, in terms of its fertilization success and embryonic development, P. acuta is the most tolerant coral species to reduced salinity thus far reported in the literature. Elevated temperature, at least that within the tolerable range of the corals, could apparently alleviate the potential negative effects from salinity stresses. This mitigating role of elevated temperature appears not to have been reported on corals before.

The promise of human embryonic stem cells in aging-associated diseases

PubMed Central

Yabut, Odessa; Bernstein, Harold S.

2011-01-01

Aging-associated diseases are often caused by progressive loss or dysfunction of cells that ultimately affect the overall function of tissues and organs. Successful treatment of these diseases could benefit from cell-based therapy that would regenerate lost cells or otherwise restore tissue function. Human embryonic stem cells (hESCs) promise to be an important therapeutic candidate in treating aging-associated diseases due to their unique capacity for self-renewal and pluripotency. To date, there are numerous hESC lines that have been developed and characterized. We will discuss how hESC lines are derived, their molecular and cellular properties, and how their ability to differentiate into all three embryonic germ layers is determined. We will also outline the methods currently employed to direct their differentiation into populations of tissue-specific, functional cells. Finally, we will highlight the general challenges that must be overcome and the strategies being developed to generate highly-purified hESC-derived cell populations that can safely be used for clinical applications. PMID:21566262

Development of an invitro technique to use mouse embryonic stem cell in evaluating effects of xenobiotics

EPA Science Inventory

Our goal has been to develop a high-throughput, in vitro technique for evaluating the effects of xenobiotics using mouse embryonic stem cells (mESCs). We began with the Embryonic Stem Cell Test (EST), which is used to predict the embryotoxic potential of a test compound by combin...

High- and low-temperature manipulation during late incubation: effects on embryonic development, the hatching process, and metabolism in broilers.

PubMed

Willemsen, H; Kamers, B; Dahlke, F; Han, H; Song, Z; Ansari Pirsaraei, Z; Tona, K; Decuypere, E; Everaert, N

2010-12-01

Temperatures continuously higher and lower than the standard incubation temperature by 3°C from embryonic d 16 until embryonic d 18.5 result in differential effects on embryonic development, the hatching process, and embryonic metabolism. Embryos in the high-temperature group were forced into a state of malnutrition by the temperature treatment, as reflected by reduced embryo growth and yolk consumption, resulting in a significantly lower chick weight at hatch. In addition, altered air cell and blood gases as well as a retarded hatching process further indicated reduced growth of embryos exposed to higher incubation temperatures during the latter part of incubation. In addition, hatchability was significantly reduced by the high-temperature treatment due to higher embryonic mortality during the treatment period and the hatching process. Levels of blood glucose, lactate, liver glycogen, plasma triglycerides, and nonesterified fatty acids indicated an altered carbohydrate and lipid metabolism for the high-temperature group. Although the hatching process of embryos exposed to lower incubation temperatures was also significantly retarded, their embryonic development and growth were strikingly similar to those of the control group.

[Embryos and embryo-like entities: problem of definition in the draft of the Swiss embryonic research law].

PubMed

Bürgin, M T; Bürkli, P

2002-11-01

At the end of May 2002, the draft of the Swiss "Federal Act on Research on Surplus Embryos and Embryonic Stem Cells" (EFG, Embryonic Research Act) reached the pre-legislative consultation stage. Under certain conditions, it would allow research on "surplus" embryos from in-vitro fertilization, and the derivation of embryonic stem cells from surplus embryos for research purposes. The EFG draft defines an embryo as "the developing organism from the point of nuclear fusion until the completion of organ development". New technological developments show that embryo-like entities can also be created without nuclear fusion having taken place. It remains unclear how to treat embryonic entities that don't fall under the draft's narrow definition of an embryo. Expanding this definition would be a welcome improvement.

A novel approach for studying the temporal modulation of embryonic skeletal development using organotypic bone cultures and microcomputed tomography.

PubMed

Kanczler, Janos M; Smith, Emma L; Roberts, Carol A; Oreffo, Richard O C

2012-10-01

Understanding the structural development of embryonic bone in a three dimensional framework is fundamental to developing new strategies for the recapitulation of bone tissue in latter life. We present an innovative combined approach of an organotypic embryonic femur culture model, microcomputed tomography (μCT) and immunohistochemistry to examine the development and modulation of the three dimensional structures of the developing embryonic femur. Isolated embryonic chick femurs were organotypic (air/liquid interface) cultured for 10 days in either basal, chondrogenic, or osteogenic supplemented culture conditions. The growth development and modulating effects of basal, chondrogenic, or osteogenic culture media of the embryonic chick femurs was investigated using μCT, immunohistochemistry, and histology. The growth and development of noncultured embryonic chick femur stages E10, E11, E12, E13, E15, and E17 were very closely correlated with increased morphometric indices of bone formation as determined by μCT. After 10 days in the organotpyic culture set up, the early aged femurs (E10 and E11) demonstrated a dramatic response to the chondrogenic or osteogenic culture conditions compared to the basal cultured femurs as determined by a change in μCT morphometric indices and modified expression of chondrogenic and osteogenic markers. Although the later aged femurs (E12 and E13) increased in size and structure after 10 days organotpypic culture, the effects of the osteogenic and chondrogenic organotypic cultures on these femurs were not significantly altered compared to basal conditions. We have demonstrated that the embryonic chick femur organotpyic culture model combined with the μCT and immunohistochemical analysis can provide an integral methodology for investigating the modulation of bone development in an ex vivo culture setting. Hence, these interdisciplinary techniques of μCT and whole organ bone cultures will enable us to delineate some of the temporal, structural developmental paradigms and modulation of bone tissue formation to underpin innovative skeletal regenerative technology for clinical therapeutic strategies in musculoskeletal trauma and diseases.

Microrna expression signatures predict patient progression and disease outcome in pediatric embryonal central nervous system neoplasms.

PubMed

Braoudaki, Maria; Lambrou, George I; Giannikou, Krinio; Milionis, Vasilis; Stefanaki, Kalliopi; Birks, Diane K; Prodromou, Neophytos; Kolialexi, Aggeliki; Kattamis, Antonis; Spiliopoulou, Chara A; Tzortzatou-Stathopoulou, Fotini; Kanavakis, Emmanouel

2014-12-31

Although, substantial experimental evidence related to diagnosis and treatment of pediatric central nervous system (CNS) neoplasms have been demonstrated, the understanding of the etiology and pathogenesis of the disease remains scarce. Recent microRNA (miRNA)-based research reveals the involvement of miRNAs in various aspects of CNS development and proposes that they might compose key molecules underlying oncogenesis. The current study evaluated miRNA differential expression detected between pediatric embryonal brain tumors and normal controls to characterize candidate biomarkers related to diagnosis, prognosis and therapy. Overall, 19 embryonal brain tumors; 15 Medulloblastomas (MBs) and 4 Atypical Teratoid/Rabdoid Tumors (AT/RTs) were studied. As controls, 13 samples were used; The First-Choice Human Brain Reference RNA and 12 samples from deceased children who underwent autopsy and were not present with any brain malignancy. RNA extraction was carried out using the Trizol method, whilst miRNA extraction was performed with the mirVANA miRNA isolation kit. The experimental approach included miRNA microarrays covering 1211 miRNAs. Quantitative Real-Time Polymerase Chain Reaction was performed to validate the expression profiles of miR-34a and miR-601 in all 32 samples initially screened with miRNA microarrays and in an additional independent cohort of 30 patients (21MBs and 9 AT/RTs). Moreover, meta-analyses was performed in total 27 embryonal tumor samples; 19 MBs, 8 ATRTs and 121 control samples. Twelve germinomas were also used as an independent validation cohort. All deregulated miRNAs were correlated to patients' clinical characteristics and pathological measures. In several cases, there was a positive correlation between individual miRNA expression levels and laboratory or clinical characteristics. Based on that, miR-601 could serve as a putative tumor suppressor gene, whilst miR-34a as an oncogene. In general, miR-34a demonstrated oncogenic roles in all pediatric embryonal CNS neoplasms studied. Deeper understanding of the aberrant miRNA expression in pediatric embryonal brain tumors might aid in the development of tumor-specific miRNA signatures, which could potentially afford promising biomarkers related to diagnosis, prognosis and patient targeted therapy.

8-Oxoguanine DNA glycosylase 1 (ogg1) maintains the function of cardiac progenitor cells during heart formation in zebrafish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yan, Lifeng; Key Laboratory of Modern Toxicology of Ministry of Education, School of Public Health, Nanjing Medical University, Nanjing 210029; Zhou, Yong

Genomic damage may devastate the potential of progenitor cells and consequently impair early organogenesis. We found that ogg1, a key enzyme initiating the base-excision repair, was enriched in the embryonic heart in zebrafish. So far, little is known about DNA repair in cardiogenesis. Here, we addressed the critical role of ogg1 in cardiogenesis for the first time. ogg1 mainly expressed in the anterior lateral plate mesoderm (ALPM), the primary heart tube, and subsequently the embryonic myocardium by in situ hybridisation. Loss of ogg1 resulted in severe cardiac morphogenesis and functional abnormalities, including the short heart length, arrhythmia, decreased cardiomyocytes andmore » nkx2.5{sup +} cardiac progenitor cells. Moreover, the increased apoptosis and repressed proliferation of progenitor cells caused by ogg1 deficiency might contribute to the heart phenotype. The microarray analysis showed that the expression of genes involved in embryonic heart tube morphogenesis and heart structure were significantly changed due to the lack of ogg1. Among those, foxh1 is an important partner of ogg1 in the cardiac development in response to DNA damage. Our work demonstrates the requirement of ogg1 in cardiac progenitors and heart development in zebrafish. These findings may be helpful for understanding the aetiology of congenital cardiac deficits. - Highlights: • A key DNA repair enzyme ogg1 is expressed in the embryonic heart in zebrafish. • We found that ogg1 is essential for normal cardiac morphogenesis in zebrafish. • The production of embryonic cardiomyocytes requires appropriate ogg1 expression. • Ogg1 critically regulated proliferation of cardiac progenitor cells in zebrafish. • foxh1 is a partner of ogg1 in the cardiac development in response to DNA damage.« less

Proximate effects of temperature versus evolved intrinsic constraints for embryonic development times among temperate and tropical songbirds

USGS Publications Warehouse

Ton, Riccardo; Martin, Thomas E.

2017-01-01

The relative importance of intrinsic constraints imposed by evolved physiological trade-offs versus the proximate effects of temperature for interspecific variation in embryonic development time remains unclear. Understanding this distinction is important because slow development due to evolved trade-offs can yield phenotypic benefits, whereas slow development from low temperature can yield costs. We experimentally increased embryonic temperature in free-living tropical and north temperate songbird species to test these alternatives. Warmer temperatures consistently shortened development time without costs to embryo mass or metabolism. However, proximate effects of temperature played an increasingly stronger role than intrinsic constraints for development time among species with colder natural incubation temperatures. Long development times of tropical birds have been thought to primarily reflect evolved physiological trade-offs that facilitate their greater longevity. In contrast, our results indicate a much stronger role of temperature in embryonic development time than currently thought.

Developments in stem cell research and therapeutic cloning: Islamic ethical positions, a review.

PubMed

Fadel, Hossam E

2012-03-01

Stem cell research is very promising. The use of human embryos has been confronted with objections based on ethical and religious positions. The recent production of reprogrammed adult (induced pluripotent) cells does not - in the opinion of scientists - reduce the need to continue human embryonic stem cell research. So the debate continues. Islam always encouraged scientific research, particularly research directed toward finding cures for human disease. Based on the expectation of potential benefits, Islamic teachings permit and support human embryonic stem cell research. The majority of Muslim scholars also support therapeutic cloning. This permissibility is conditional on the use of supernumerary early pre-embryos which are obtained during infertility treatment in vitro fertilization (IVF) clinics. The early pre-embryos are considered in Islamic jurisprudence as worthy of respect but do not have the full sanctity offered to the embryo after implantation in the uterus and especially after ensoulment. In this paper the Islamic positions regarding human embryonic stem cell research and therapeutic cloning are reviewed in some detail, whereas positions in other religious traditions are mentioned only briefly. The status of human embryonic stem cell research and therapeutic cloning in different countries, including the USA and especially in Muslim countries, is discussed. © 2010 Blackwell Publishing Ltd.

Transforming Growth Factor Beta (TGFβ1, TGFβ2 and TGFβ3) Null-Mutant Phenotypes in Embryonic Gonadal Development

PubMed Central

Memon, Mushtaq A.; Anway, Matthew D.; Covert, Trevor R.; Uzumcu, Mehmet; Skinner, Michael K.

2008-01-01

The role transforming growth factor beta (TGFb) isoforms TGFb1, TGFb2 and TGFb3 have in the regulation of embryonic gonadal development was investigated with the use of null-mutant (i.e. knockout) mice for each of the TGFb isoforms. Late embryonic gonadal development was investigated because homozygote TGFb null-mutant mice generally die around birth, with some embryonic loss as well. In the testis, the TGFb1 null-mutant mice had a decrease in the number of germ cells at birth, postnatal day 0 (P0). In the testis, the TGFb2 null-mutant mice had a decrease in the number of seminiferous cords at embryonic day 15 (E15). In the ovary, the TGFb2 null-mutant mice had an increase in the number of germ cells at P0. TGFb isoforms appear to have a role in gonadal development, but interactions between the isoforms is speculated to compensate in the different TGFb isoform null-mutant mice. PMID:18790002

TEAD and YAP regulate the enhancer network of human embryonic pancreatic progenitors.

PubMed

Cebola, Inês; Rodríguez-Seguí, Santiago A; Cho, Candy H-H; Bessa, José; Rovira, Meritxell; Luengo, Mario; Chhatriwala, Mariya; Berry, Andrew; Ponsa-Cobas, Joan; Maestro, Miguel Angel; Jennings, Rachel E; Pasquali, Lorenzo; Morán, Ignasi; Castro, Natalia; Hanley, Neil A; Gomez-Skarmeta, Jose Luis; Vallier, Ludovic; Ferrer, Jorge

2015-05-01

The genomic regulatory programmes that underlie human organogenesis are poorly understood. Pancreas development, in particular, has pivotal implications for pancreatic regeneration, cancer and diabetes. We have now characterized the regulatory landscape of embryonic multipotent progenitor cells that give rise to all pancreatic epithelial lineages. Using human embryonic pancreas and embryonic-stem-cell-derived progenitors we identify stage-specific transcripts and associated enhancers, many of which are co-occupied by transcription factors that are essential for pancreas development. We further show that TEAD1, a Hippo signalling effector, is an integral component of the transcription factor combinatorial code of pancreatic progenitor enhancers. TEAD and its coactivator YAP activate key pancreatic signalling mediators and transcription factors, and regulate the expansion of pancreatic progenitors. This work therefore uncovers a central role for TEAD and YAP as signal-responsive regulators of multipotent pancreatic progenitors, and provides a resource for the study of embryonic development of the human pancreas.

Utilization of ketone bodies by chick brain and spinal cord during embryonic and postnatal development.

PubMed

Linares, A; Caamaño, G J; Diaz, R; Gonzalez, F J; Garcia-Peregrin, E

1993-10-01

Lipid synthesis from acetoacetate and 3-hydroxybutyrate was studied in chick embryo from 15 to 21 days and in chick neonate from 1 to 21 days. Embryonic spinal cord showed higher ability than brain to incorporate acetoacetate into total lipids, although a sharp decrease was found at hatching. 3-Hydroxybutyrate incorporation into total lipids was also higher in spinal cord than in brain, especially during the embryonic period. Phospholipids were the main lipids formed in both tissues from both precursors. An appreciable percentage of radioactivity was also recovered as free cholesterol, especially during the embryonic phase. The developmental patterns of amino acid synthesis from acetoacetate and 3-hydroxybutyrate were similar in both tissues: a clear increase after hatching was followed by a decrease at day 4 of neonatal life. Acetoacetate was a better substrate for amino acid synthesis than 3-hydroxybutyrate during the embryonic development in both tissues. Oxidation of both precursors to CO2 strongly decreased between 15 and 21 days of embryonic development both in brain and spinal cord.

PTBP1 Is Required for Embryonic Development before Gastrulation

PubMed Central

Suckale, Jakob; Wendling, Olivia; Masjkur, Jimmy; Jäger, Melanie; Münster, Carla; Anastassiadis, Konstantinos; Stewart, A. Francis; Solimena, Michele

2011-01-01

Polypyrimidine-tract binding protein 1 (PTBP1) is an important cellular regulator of messenger RNAs influencing the alternative splicing profile of a cell as well as its mRNA stability, location and translation. In addition, it is diverted by some viruses to facilitate their replication. Here, we used a novel PTBP1 knockout mouse to analyse the tissue expression pattern of PTBP1 as well as the effect of its complete removal during development. We found evidence of strong PTBP1 expression in embryonic stem cells and throughout embryonic development, especially in the developing brain and spinal cord, the olfactory and auditory systems, the heart, the liver, the kidney, the brown fat and cartilage primordia. This widespread distribution points towards a role of PTBP1 during embryonic development. Homozygous offspring, identified by PCR and immunofluorescence, were able to implant but were arrested or retarded in growth. At day 7.5 of embryonic development (E7.5) the null mutants were about 5x smaller than the control littermates and the gap in body size widened with time. At mid-gestation, all homozygous embryos were resorbed/degraded. No homozygous mice were genotyped at E12 and the age of weaning. Embryos lacking PTBP1 did not display differentiation into the 3 germ layers and cavitation of the epiblast, which are hallmarks of gastrulation. In addition, homozygous mutants displayed malformed ectoplacental cones and yolk sacs, both early supportive structure of the embryo proper. We conclude that PTBP1 is not required for the earliest isovolumetric divisions and differentiation steps of the zygote up to the formation of the blastocyst. However, further post-implantation development requires PTBP1 and stalls in homozygous null animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures. PMID:21423341

PTBP1 is required for embryonic development before gastrulation.

PubMed

Suckale, Jakob; Wendling, Olivia; Masjkur, Jimmy; Jäger, Melanie; Münster, Carla; Anastassiadis, Konstantinos; Stewart, A Francis; Solimena, Michele

2011-02-17

Polypyrimidine-tract binding protein 1 (PTBP1) is an important cellular regulator of messenger RNAs influencing the alternative splicing profile of a cell as well as its mRNA stability, location and translation. In addition, it is diverted by some viruses to facilitate their replication. Here, we used a novel PTBP1 knockout mouse to analyse the tissue expression pattern of PTBP1 as well as the effect of its complete removal during development. We found evidence of strong PTBP1 expression in embryonic stem cells and throughout embryonic development, especially in the developing brain and spinal cord, the olfactory and auditory systems, the heart, the liver, the kidney, the brown fat and cartilage primordia. This widespread distribution points towards a role of PTBP1 during embryonic development. Homozygous offspring, identified by PCR and immunofluorescence, were able to implant but were arrested or retarded in growth. At day 7.5 of embryonic development (E7.5) the null mutants were about 5x smaller than the control littermates and the gap in body size widened with time. At mid-gestation, all homozygous embryos were resorbed/degraded. No homozygous mice were genotyped at E12 and the age of weaning. Embryos lacking PTBP1 did not display differentiation into the 3 germ layers and cavitation of the epiblast, which are hallmarks of gastrulation. In addition, homozygous mutants displayed malformed ectoplacental cones and yolk sacs, both early supportive structure of the embryo proper. We conclude that PTBP1 is not required for the earliest isovolumetric divisions and differentiation steps of the zygote up to the formation of the blastocyst. However, further post-implantation development requires PTBP1 and stalls in homozygous null animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.

Fucoidan promotes early step of cardiac differentiation from human embryonic stem cells and long-term maintenance of beating areas.

PubMed

Hamidi, Sofiane; Letourneur, Didier; Aid-Launais, Rachida; Di Stefano, Antonio; Vainchenker, William; Norol, Françoise; Le Visage, Catherine

2014-04-01

Somatic stem cells require specific niches and three-dimensional scaffolds provide ways to mimic this microenvironment. Here, we studied a scaffold based on Fucoidan, a sulfated polysaccharide known to influence morphogen gradients during embryonic development, to support human embryonic stem cells (hESCs) differentiation toward the cardiac lineage. A macroporous (pore 200 μm) Fucoidan scaffold was selected to support hESCs attachment and proliferation. Using a protocol based on the cardiogenic morphogen bone morphogenic protein 2 (BMP2) and transforming growth factor (TGFβ) followed by tumor necrosis factor (TNFα), an effector of cardiopoietic priming, we examined the cardiac differentiation in the scaffold compared to culture dishes and embryoid bodies (EBs). At day 8, Fucoidan scaffolds supported a significantly higher expression of the 3 genes encoding for transcription factors marking the early step of embryonic cardiac differentiation NKX2.5 (p<0.05), MEF2C (p<0.01), and GATA4 (p<0.01), confirmed by flow cytometry analysis for MEF2C and NKX2.5. The ability of Fucoidan scaffolds to locally concentrate and slowly release TGFβ and TNFα was confirmed by Luminex technology. We also found that Fucoidan scaffolds supported the late stage of embryonic cardiac differentiation marked by a significantly higher atrial natriuretic factor (ANF) expression (p<0.001), although only rare beating areas were observed. We postulated that absence of mechanical stress in the soft hydrogel impaired sarcomere formation, as confirmed by molecular analysis of the cardiac muscle myosin MYH6 and immunohistological staining of sarcomeric α-actinin. Nevertheless, Fucoidan scaffolds contributed to the development of thin filaments connecting beating areas through promotion of smooth muscle cells, thus enabling maintenance of beating areas for up to 6 months. In conclusion, Fucoidan scaffolds appear as a very promising biomaterial to control cardiac differentiation from hESCs that could be further combined with mechanical stress to promote sarcomere formation at terminal stages of differentiation.

Tyrosine pathway regulation is host-mediated in the pea aphid symbiosis during late embryonic and early larval development.

PubMed

Rabatel, Andréane; Febvay, Gérard; Gaget, Karen; Duport, Gabrielle; Baa-Puyoulet, Patrice; Sapountzis, Panagiotis; Bendridi, Nadia; Rey, Marjolaine; Rahbé, Yvan; Charles, Hubert; Calevro, Federica; Colella, Stefano

2013-04-10

Nutritional symbioses play a central role in insects' adaptation to specialized diets and in their evolutionary success. The obligatory symbiosis between the pea aphid, Acyrthosiphon pisum, and the bacterium, Buchnera aphidicola, is no exception as it enables this important agricultural pest insect to develop on a diet exclusively based on plant phloem sap. The symbiotic bacteria provide the host with essential amino acids lacking in its diet but necessary for the rapid embryonic growth seen in the parthenogenetic viviparous reproduction of aphids. The aphid furnishes, in exchange, non-essential amino acids and other important metabolites. Understanding the regulations acting on this integrated metabolic system during the development of this insect is essential in elucidating aphid biology. We used a microarray-based approach to analyse gene expression in the late embryonic and the early larval stages of the pea aphid, characterizing, for the first time, the transcriptional profiles in these developmental phases. Our analyses allowed us to identify key genes in the phenylalanine, tyrosine and dopamine pathways and we identified ACYPI004243, one of the four genes encoding for the aspartate transaminase (E.C. 2.6.1.1), as specifically regulated during development. Indeed, the tyrosine biosynthetic pathway is crucial for the symbiotic metabolism as it is shared between the two partners, all the precursors being produced by B. aphidicola. Our microarray data are supported by HPLC amino acid analyses demonstrating an accumulation of tyrosine at the same developmental stages, with an up-regulation of the tyrosine biosynthetic genes. Tyrosine is also essential for the synthesis of cuticular proteins and it is an important precursor for cuticle maturation: together with the up-regulation of tyrosine biosynthesis, we observed an up-regulation of cuticular genes expression. We were also able to identify some amino acid transporter genes which are essential for the switch over to the late embryonic stages in pea aphid development. Our data show that, in the development of A. pisum, a specific host gene set regulates the biosynthetic pathways of amino acids, demonstrating how the regulation of gene expression enables an insect to control the production of metabolites crucial for its own development and symbiotic metabolism.

The Maternal to Zygotic Transition in Mammals

PubMed Central

Li, Lei; Lu, Xukun; Dean, Jurrien

2013-01-01

Prior to activation of the embryonic genome, the initiating events of mammalian development are under maternal control and include fertilization, the block to polyspermy and processing sperm DNA. Following gamete union, the transcriptionally inert sperm DNA is repackaged into the male pronucleus which fuses with the female pronucleus to form a 1-cell zygote. Embryonic transcription begins during the maternal to zygotic transfer of control in directing development. This transition occurs at species-specific times after one or several rounds of blastomere cleavage and is essential for normal development. However, even after activation of the embryonic genome, successful development relies on stored maternal components without which embryos fail to progress beyond initial cell divisions. Better understanding of the molecular basis of maternal to zygotic transition including fertilization, the activation of the embryonic genome and cleavage-stage development will provide insight into early human development that should translate into clinical applications for regenerative medicine and assisted reproductive technologies. PMID:23352575

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Xiugong, E-mail: xiugong.gao@fda.hhs.gov; Sprando, Robert L.; Yourick, Jeffrey J.

Developmental toxicity testing has traditionally relied on animal models which are costly, time consuming, and require the sacrifice of large numbers of animals. In addition, there are significant disparities between human beings and animals in their responses to chemicals. Thalidomide is a species-specific developmental toxicant that causes severe limb malformations in humans but not in mice. Here, we used microarrays to study transcriptomic changes induced by thalidomide in an in vitro model based on differentiation of mouse embryonic stem cells (mESCs). C57BL/6 mESCs were allowed to differentiate spontaneously and RNA was collected at 24, 48, and 72 h after exposuremore » to 0.25 mM thalidomide. Global gene expression analysis using microarrays revealed hundreds of differentially expressed genes upon thalidomide exposure that were enriched in gene ontology (GO) terms and canonical pathways associated with embryonic development and differentiation. In addition, many genes were found to be involved in small GTPases-mediated signal transduction, heart development, and inflammatory responses, which coincide with clinical evidences and may represent critical embryotoxicities of thalidomide. These results demonstrate that transcriptomics in combination with mouse embryonic stem cell differentiation is a promising alternative model for developmental toxicity assessment. - Highlights: • Studied genomic changes in mouse embryonic stem cells upon thalidomide exposure • Identified gene expression changes that may represent thalidomide embryotoxicity • The toxicogenomic changes coincide well with known thalidomide clinical outcomes. • The mouse embryonic stem cell model is suitable for developmental toxicity testing. • The model has the potential for high-throughput screening of a multitude of compounds.« less

A review of factors influencing the availability of dissolved oxygen to incubating salmonid embryos

NASA Astrophysics Data System (ADS)

Greig, S. M.; Sear, D. A.; Carling, P. A.

2007-01-01

Previous investigations into factors influencing incubation success of salmonid progeny have largely been limited to the development of empirical relationships between characteristics of the incubation environment and survival to emergence. It is suggested that adopting a process-based approach to assessing incubation success aids identification of the precise causes of embryonic mortalities, and provides a robust framework for developing and implementing managerial responses.Identifying oxygen availability within the incubation environment as a limiting factor, a comprehensive review of trends in embryonic respiration, and processes influencing the flux of oxygenated water through gravel riverbeds is provided. The availability of oxygen to incubating salmonid embryos is dependent on the exchange of oxygenated water with the riverbed, and the ability of the riverbed gravel medium to transport this water at a rate and concentration appropriate to support embryonic respiratory requirements. Embryonic respiratory trends indicate that oxygen consumption varies with stage of development, ambient water temperature and oxygen availability. The flux of oxygenated water through the incubation environment is controlled by a complex interaction of intragravel and extragravel processes and factors. The processes driving the exchange of channel water with gravel riverbeds include bed topography, bed permeability, and surface roughness effects. The flux of oxygenated water through riverbed gravels is controlled by gravel permeability, coupling of surface-subsurface flow and oxygen demands imposed by materials infiltrating riverbed gravels. Temporally and spatially variable inputs of groundwater can also influence the oxygen concentration of interstitial water. Copyright

A Novel Approach for Studying the Temporal Modulation of Embryonic Skeletal Development Using Organotypic Bone Cultures and Microcomputed Tomography

PubMed Central

Smith, Emma L.; Roberts, Carol A.

2012-01-01

Understanding the structural development of embryonic bone in a three dimensional framework is fundamental to developing new strategies for the recapitulation of bone tissue in latter life. We present an innovative combined approach of an organotypic embryonic femur culture model, microcomputed tomography (μCT) and immunohistochemistry to examine the development and modulation of the three dimensional structures of the developing embryonic femur. Isolated embryonic chick femurs were organotypic (air/liquid interface) cultured for 10 days in either basal, chondrogenic, or osteogenic supplemented culture conditions. The growth development and modulating effects of basal, chondrogenic, or osteogenic culture media of the embryonic chick femurs was investigated using μCT, immunohistochemistry, and histology. The growth and development of noncultured embryonic chick femur stages E10, E11, E12, E13, E15, and E17 were very closely correlated with increased morphometric indices of bone formation as determined by μCT. After 10 days in the organotpyic culture set up, the early aged femurs (E10 and E11) demonstrated a dramatic response to the chondrogenic or osteogenic culture conditions compared to the basal cultured femurs as determined by a change in μCT morphometric indices and modified expression of chondrogenic and osteogenic markers. Although the later aged femurs (E12 and E13) increased in size and structure after 10 days organotpypic culture, the effects of the osteogenic and chondrogenic organotypic cultures on these femurs were not significantly altered compared to basal conditions. We have demonstrated that the embryonic chick femur organotpyic culture model combined with the μCT and immunohistochemical analysis can provide an integral methodology for investigating the modulation of bone development in an ex vivo culture setting. Hence, these interdisciplinary techniques of μCT and whole organ bone cultures will enable us to delineate some of the temporal, structural developmental paradigms and modulation of bone tissue formation to underpin innovative skeletal regenerative technology for clinical therapeutic strategies in musculoskeletal trauma and diseases. PMID:22472170

Development and maintenance of a telescoping debris flow fan in response to human-induced fan surface channelization, Chalk Creek Valley Natural Debris Flow Laboratory, Colorado, USA

NASA Astrophysics Data System (ADS)

Wasklewicz, T.; Scheinert, C.

2016-01-01

Channel change has been a constant theme throughout William L. Graf's research career. Graf's work has examined channel changes in the context of natural environmental fluctuations, but more often has focused on quantifying channel change in the context of anthropogenic modifications. Here, we consider how channelization of a debris flows along a bajada has perpetuated and sustained the development of 'telescoping' alluvial fan. Two-dimensional debris-flow modeling shows the importance of the deeply entrenched channelized flow in the development of a telescoping alluvial fan. GIS analyses of repeat (five different debris flows), high-resolution (5 cm) digital elevation models (DEMs) generated from repeat terrestrial laser scanning (TLS) data elucidate sediment and topographic dynamics of the new telescoping portion of the alluvial fan (the embryonic fan). Flow constriction from channelization helps to perpetuate debris-flow runout and to maintain the embryonic fan and telescoping nature of the alluvial fan complex. Embryonic fan development, in response to five debris flows, proceeds with a major portion of the flows depositing on the southern portion of the embryonic fan. The third through the fifth debris flows also begin to shift some deposition to the northern portion of the embryonic. The transfer of sediment from a higher portion of the embryonic fan to a lower portion continues currently on the embryonic fan. While channelized flow has been shown to be critical to the maintenance of the telescoping fan, the flow constriction has led to higher than background levels of sediment deposition in Chalk Creek, a tributary of the Arkansas River. A majority of the sediment from each debris flow is incorporated into Chalk Creek as opposed to being stored on the embryonic fan.

Transcriptional profiles of bovine in vivo pre-implantation development.

PubMed

Jiang, Zongliang; Sun, Jiangwen; Dong, Hong; Luo, Oscar; Zheng, Xinbao; Obergfell, Craig; Tang, Yong; Bi, Jinbo; O'Neill, Rachel; Ruan, Yijun; Chen, Jingbo; Tian, Xiuchun Cindy

2014-09-04

During mammalian pre-implantation embryonic development dramatic and orchestrated changes occur in gene transcription. The identification of the complete changes has not been possible until the development of the Next Generation Sequencing Technology. Here we report comprehensive transcriptome dynamics of single matured bovine oocytes and pre-implantation embryos developed in vivo. Surprisingly, more than half of the estimated 22,000 bovine genes, 11,488 to 12,729 involved in more than 100 pathways, is expressed in oocytes and early embryos. Despite the similarity in the total numbers of genes expressed across stages, the nature of the expressed genes is dramatically different. A total of 2,845 genes were differentially expressed among different stages, of which the largest change was observed between the 4- and 8-cell stages, demonstrating that the bovine embryonic genome is activated at this transition. Additionally, 774 genes were identified as only expressed/highly enriched in particular stages of development, suggesting their stage-specific roles in embryogenesis. Using weighted gene co-expression network analysis, we found 12 stage-specific modules of co-expressed genes that can be used to represent the corresponding stage of development. Furthermore, we identified conserved key members (or hub genes) of the bovine expressed gene networks. Their vast association with other embryonic genes suggests that they may have important regulatory roles in embryo development; yet, the majority of the hub genes are relatively unknown/under-studied in embryos. We also conducted the first comparison of embryonic expression profiles across three mammalian species, human, mouse and bovine, for which RNA-seq data are available. We found that the three species share more maternally deposited genes than embryonic genome activated genes. More importantly, there are more similarities in embryonic transcriptomes between bovine and humans than between humans and mice, demonstrating that bovine embryos are better models for human embryonic development. This study provides a comprehensive examination of gene activities in bovine embryos and identified little-known potential master regulators of pre-implantation development.

Endothelin-1 signalling controls early embryonic heart rate in vitro and in vivo.

PubMed

Karppinen, S; Rapila, R; Mäkikallio, K; Hänninen, S L; Rysä, J; Vuolteenaho, O; Tavi, P

2014-02-01

Spontaneous activity of embryonic cardiomyocytes originates from sarcoplasmic reticulum (SR) Ca(2+) release during early cardiogenesis. However, the regulation of heart rate during embryonic development is still not clear. The aim of this study was to determine how endothelin-1 (ET-1) affects the heart rate of embryonic mice, as well as the pathway through which it exerts its effects. The effects of ET-1 and ET-1 receptor inhibition on cardiac contraction were studied using confocal Ca(2+) imaging of isolated mouse embryonic ventricular cardiomyocytes and ultrasonographic examination of embryonic cardiac contractions in utero. In addition, the amount of ET-1 peptide and ET receptor a (ETa) and b (ETb) mRNA levels were measured during different stages of development of the cardiac muscle. High ET-1 concentration and expression of both ETa and ETb receptors was observed in early cardiac tissue. ET-1 was found to increase the frequency of spontaneous Ca(2+) oscillations in E10.5 embryonic cardiomyocytes in vitro. Non-specific inhibition of ET receptors with tezosentan caused arrhythmia and bradycardia in isolated embryonic cardiomyocytes and in whole embryonic hearts both in vitro (E10.5) and in utero (E12.5). ET-1-mediated stimulation of early heart rate was found to occur via ETb receptors and subsequent inositol trisphosphate receptor activation and increased SR Ca(2+) leak. Endothelin-1 is required to maintain a sufficient heart rate, as well as to prevent arrhythmia during early development of the mouse heart. This is achieved through ETb receptor, which stimulates Ca(2+) leak through IP3 receptors. © 2013 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Identification and Characterization of Long Non-Coding RNAs Related to Mouse Embryonic Brain Development from Available Transcriptomic Data

PubMed Central

He, Hongjuan; Xiu, Youcheng; Guo, Jing; Liu, Hui; Liu, Qi; Zeng, Tiebo; Chen, Yan; Zhang, Yan; Wu, Qiong

2013-01-01

Long non-coding RNAs (lncRNAs) as a key group of non-coding RNAs have gained widely attention. Though lncRNAs have been functionally annotated and systematic explored in higher mammals, few are under systematical identification and annotation. Owing to the expression specificity, known lncRNAs expressed in embryonic brain tissues remain still limited. Considering a large number of lncRNAs are only transcribed in brain tissues, studies of lncRNAs in developmental brain are therefore of special interest. Here, publicly available RNA-sequencing (RNA-seq) data in embryonic brain are integrated to identify thousands of embryonic brain lncRNAs by a customized pipeline. A significant proportion of novel transcripts have not been annotated by available genomic resources. The putative embryonic brain lncRNAs are shorter in length, less spliced and show less conservation than known genes. The expression of putative lncRNAs is in one tenth on average of known coding genes, while comparable with known lncRNAs. From chromatin data, putative embryonic brain lncRNAs are associated with active chromatin marks, comparable with known lncRNAs. Embryonic brain expressed lncRNAs are also indicated to have expression though not evident in adult brain. Gene Ontology analysis of putative embryonic brain lncRNAs suggests that they are associated with brain development. The putative lncRNAs are shown to be related to possible cis-regulatory roles in imprinting even themselves are deemed to be imprinted lncRNAs. Re-analysis of one knockdown data suggests that four regulators are associated with lncRNAs. Taken together, the identification and systematic analysis of putative lncRNAs would provide novel insights into uncharacterized mouse non-coding regions and the relationships with mammalian embryonic brain development. PMID:23967161

The ‘Ventral Organs’ of Pycnogonida (Arthropoda) Are Neurogenic Niches of Late Embryonic and Post-Embryonic Nervous System Development

PubMed Central

Brenneis, Georg; Scholtz, Gerhard

2014-01-01

Early neurogenesis in arthropods has been in the focus of numerous studies, its cellular basis, spatio-temporal dynamics and underlying genetic network being by now comparably well characterized for representatives of chelicerates, myriapods, hexapods and crustaceans. By contrast, neurogenesis during late embryonic and/or post-embryonic development has received less attention, especially in myriapods and chelicerates. Here, we apply (i) immunolabeling, (ii) histology and (iii) scanning electron microscopy to study post-embryonic ventral nerve cord development in Pseudopallene sp., a representative of the sea spiders (Pycnogonida), the presumable sister group of the remaining chelicerates. During early post-embryonic development, large neural stem cells give rise to additional ganglion cell material in segmentally paired invaginations in the ventral ectoderm. These ectodermal cell regions – traditionally designated as ‘ventral organs’ – detach from the surface into the interior and persist as apical cell clusters on the ventral ganglion side. Each cluster is a post-embryonic neurogenic niche that features a tiny central cavity and initially still houses larger neural stem cells. The cluster stays connected to the underlying ganglionic somata cortex via an anterior and a posterior cell stream. Cell proliferation remains restricted to the cluster and streams, and migration of newly produced cells along the streams seems to account for increasing ganglion cell numbers in the cortex. The pycnogonid cluster-stream-systems show striking similarities to the life-long neurogenic system of decapod crustaceans, and due to their close vicinity to glomerulus-like neuropils, we consider their possible involvement in post-embryonic (perhaps even adult) replenishment of olfactory neurons – as in decapods. An instance of a potentially similar post-embryonic/adult neurogenic system in the arthropod outgroup Onychophora is discussed. Additionally, we document two transient posterior ganglia in the ventral nerve cord of Pseudopallene sp. and evaluate this finding in light of the often discussed reduction of a segmented ‘opisthosoma’ during pycnogonid evolution. PMID:24736377

Virtual reality imaging techniques in the study of embryonic and early placental health.

PubMed

Rousian, Melek; Koster, Maria P H; Mulders, Annemarie G M G J; Koning, Anton H J; Steegers-Theunissen, Régine P M; Steegers, Eric A P

2018-04-01

Embryonic and placental growth and development in the first trimester of pregnancy have impact on the health of the fetus, newborn, child and even the adult. This emphasizes the importance of this often neglected period in life. The development of three-dimensional transvaginal ultrasonography in combination with virtual reality (VR) opens the possibility of accurate and reliable visualization of embryonic and placental structures with real depth perception. These techniques enable new biometry and volumetry measurements that contribute to the knowledge of the (patho)physiology of embryonic and early placental health. Examples of such measurements are the length of complex structures like the umbilical cord, vitelline duct, limbs and cerebellum or the volume of the whole embryo and brain cavities. Moreover, for the first time, embryos can now be staged in vivo (Carnegie stages) and vasculature volumes of both the embryo and the early placenta can be measured when VR is combined with power Doppler signals. These innovative developments have already been used to study associations between periconceptional maternal factors, such as age, smoking, alcohol use, diet and vitamin status, and embryonic and early placental growth and development. Future studies will also focus on the identification of abnormal embryonic and early placental development already in the earliest weeks of pregnancy, which provides opportunities for early prevention of pregnancy complications. Copyright © 2018 IFPA, Elsevier Ltd. Published by Elsevier Ltd.. All rights reserved.

Metabolome analysis of Drosophila melanogaster during embryogenesis.

PubMed

An, Phan Nguyen Thuy; Yamaguchi, Masamitsu; Bamba, Takeshi; Fukusaki, Eiichiro

2014-01-01

The Drosophila melanogaster embryo has been widely utilized as a model for genetics and developmental biology due to its small size, short generation time, and large brood size. Information on embryonic metabolism during developmental progression is important for further understanding the mechanisms of Drosophila embryogenesis. Therefore, the aim of this study is to assess the changes in embryos' metabolome that occur at different stages of the Drosophila embryonic development. Time course samples of Drosophila embryos were subjected to GC/MS-based metabolome analysis for profiling of low molecular weight hydrophilic metabolites, including sugars, amino acids, and organic acids. The results showed that the metabolic profiles of Drosophila embryo varied during the course of development and there was a strong correlation between the metabolome and different embryonic stages. Using the metabolome information, we were able to establish a prediction model for developmental stages of embryos starting from their high-resolution quantitative metabolite composition. Among the important metabolites revealed from our model, we suggest that different amino acids appear to play distinct roles in different developmental stages and an appropriate balance in trehalose-glucose ratio is crucial to supply the carbohydrate source for the development of Drosophila embryo.

Metabolome Analysis of Drosophila melanogaster during Embryogenesis

PubMed Central

An, Phan Nguyen Thuy; Yamaguchi, Masamitsu; Bamba, Takeshi; Fukusaki, Eiichiro

2014-01-01

The Drosophila melanogaster embryo has been widely utilized as a model for genetics and developmental biology due to its small size, short generation time, and large brood size. Information on embryonic metabolism during developmental progression is important for further understanding the mechanisms of Drosophila embryogenesis. Therefore, the aim of this study is to assess the changes in embryos’ metabolome that occur at different stages of the Drosophila embryonic development. Time course samples of Drosophila embryos were subjected to GC/MS-based metabolome analysis for profiling of low molecular weight hydrophilic metabolites, including sugars, amino acids, and organic acids. The results showed that the metabolic profiles of Drosophila embryo varied during the course of development and there was a strong correlation between the metabolome and different embryonic stages. Using the metabolome information, we were able to establish a prediction model for developmental stages of embryos starting from their high-resolution quantitative metabolite composition. Among the important metabolites revealed from our model, we suggest that different amino acids appear to play distinct roles in different developmental stages and an appropriate balance in trehalose-glucose ratio is crucial to supply the carbohydrate source for the development of Drosophila embryo. PMID:25121768

Discovery, characterization and expression of a novel zebrafish gene, znfr, important for notochord formation.

PubMed

Xu, Yan; Zou, Peng; Liu, Yao; Deng, Fengjiao

2010-06-01

Genes specifically expressed in the notochord may be crucial for proper notochord development. Using the digital differential display program offered by the National Center for Biotechnology Information, we identified a novel EST sequence from a zebrafish ovary library (No. XM_701450). The full-length cDNA of this transcript was cloned by performing 3' and 5'-RACE and was further confirmed by PCR and sequencing. The resulting 614 bp gene was found to encode a novel 94 amino acid protein that did not share significant homology with any other known protein. Characterization of the genomic sequence revealed that the gene spanned 4.9 kb and was composed of four exons and three introns. RT-PCR gene expression analysis revealed that our gene of interest was expressed in ovary, kidney, brain, mature oocytes and during the early stages of embryogenesis. During embryonic development, znfr mRNA was found to be expressed in the embryonic shield, chordamesoderm and the vacuolated notochord cells by in situ hybridization. Based on this information, we hypothesize that this novel gene is an important maternal factor required for zebrafish notochord formation during early embryonic development. We have thus named this gene znfr (zebrafish notochord formation related).

Growth enhancement by embryonic fibroblasts upon cotransplantation of noncommitted pig embryonic tissues with fully committed organs.

PubMed

Cohen, Sivan; Tchorsh-Yutsis, Dalit; Aronovich, Anna; Tal, Orna; Eventov-Friedman, Smadar; Katchman, Helena; Klionsky, Yael; Shezen, Elias; Reisner, Yair

2010-05-27

We recently defined the optimal gestational time windows for the transplantation of several embryonic tissues. We showed that the liver and kidney obtained from E28 pig embryos can grow and differentiate normally after transplantation, whereas 1 week earlier in gestation, these tissues develop into teratoma-like structures or fibrotic mass. In this study, we investigated whether cotransplantation of E28 with E21 tissue could control its tumorogenic potential, or alternatively whether the stem cells derived from the earlier tissue contribute to the growth of the more committed one. Pig embryonic precursors from E21 and E28 gestational age were transplanted alone or together, into nonobese diabetic/severe combined immunodeficiency mice, and their growth and differentiation was evaluated by immunohistology. In situ analysis, based on sex disparity between the E21 and E28 tissues, was used to identify the tissue source. In some experiments, mouse embryonic fibroblasts (MEF) were cotransplanted with E28 liver, and their effect was evaluated. E28 tissues could not abrogate the propensity of the cells within the undifferentiated tissue to form teratoma-like structures. However, E21 kidney or liver tissue markedly enhanced the growth and function of E28 kidney, liver, and heart grafts. Moreover, similar growth enhancement was observed on coimplantation of E28 liver tissue with MEF or on infusion of MEF culture medium, indicating that this enhancement is likely mediated through soluble factors secreted by the fibroblasts. Our results suggest a novel approach for the enhancement of growth and differentiation of transplanted embryonic tissues by the use of soluble factors secreted by embryonic fibroblasts.

Can physics help to explain embryonic development? An overview.

PubMed

Fleury, V

2013-10-01

Recent technical advances including digital imaging and particle image velocimetry can be used to extract the full range of embryonic movements that constitute the instantaneous 'morphogenetic fields' of a developing animal. The final shape of the animal results from the sum over time (integral) of the movements that make up the velocity fields of all the tissue constituents. In vivo microscopy can be used to capture the details of vertebrate development at the earliest embryonic stages. The movements thus observed can be quantitatively compared to physical models that provide velocity fields based on simple hypotheses about the nature of living matter (a visco-elastic gel). This approach has cast new light on the interpretation of embryonic movement, folding, and organisation. It has established that several major discontinuities in development are simple physical changes in boundary conditions. In other words, with no change in biology, the physical consequences of collisions between folds largely explain the morphogenesis of the major structures (such as the head). Other discontinuities result from changes in physical conditions, such as bifurcations (changes in physical behaviour beyond specific yield points). For instance, beyond a certain level of stress, a tissue folds, without any new gene being involved. An understanding of the physical features of movement provides insights into the levers that drive evolution; the origin of animals is seen more clearly when viewed under the light of the fundamental physical laws (Newton's principle, action-reaction law, changes in symmetry breaking scale). This article describes the genesis of a vertebrate embryo from the shapeless stage (round mass of tissue) to the development of a small, elongated, bilaterally symmetric structure containing vertebral precursors, hip and shoulder enlarges, and a head. Copyright © 2013. Published by Elsevier Masson SAS.

Dietary genistein supplementation in laying broiler breeder hens alters the development and metabolism of offspring embryos as revealed by hepatic transcriptome analysis.

PubMed

Lv, Zengpeng; Fan, Hao; Zhang, Beibei; Ning, Chao; Xing, Kun; Guo, Yuming

2018-03-08

Genistein (GEN) is a type of isoflavone mainly derived from soy products. In this experiment, we added 40 and 400 mg/kg GEN to the diet of laying broiler breeder hens to clarify the maternal effects of GEN on the development and metabolism of chick embryos. GEN treatment at 40 mg/kg increased embryonic length, weight, and liver index, as well as the width of the proliferative zone in the tibial growth plate of chick embryos. Gene ontology (GO) cluster analysis of the hepatic transcriptome showed that GEN treatment promoted embryonic development and cell proliferation. Low-dose GEN treatment increased insulin growth factor-binding protein (IGFBP)3 mRNA expression in the embryonic liver, whereas high-dose GEN treatment increased IGFBP5 expression and activated the apoptosis and protein tyrosine kinase signaling pathways. Furthermore, adding supplemental GEN to the diet of hens promoted the glycolysis process in the embryonic liver through the insulin-signaling pathway, upregulated target genes (phosphoglucomutase-2, hexokinase 1, dihydroxyacetone phosphate by aldolase, phosphofructokinase, platelet, and enolase 2), and enhanced the transport of carboxylic acids and cholesterol and the synthesis of unsaturated fatty acid (arachidonic acid) in the embryonic liver through upregulation of liver X receptor, sterol regulatory element-binding protein 1, and patatin-like phospholipase A. Additionally, GEN treatment increased fatty acid β-oxidation and Na + /K + -ATPase activity in the embryonic liver through activation of peroxisome proliferator-activated receptors (PPARs; PPARα and PPARδ) and the AMPK signaling pathway, which could provide energy for embryonic development. In addition, GEN treatment in hens increased superoxide dismutase activity and metallothionein expression in the chick embryonic liver and promoted lymphocyte proliferation through upregulation of mRNA expression of CDKN1A, IL12RB1, Sox11, PRKAR1A, PRKCQ, and TCF3. The improved immunity and antioxidant capacity, as a result of maternal GEN effects, was conducive to embryonic development. In conclusion, the addition of GEN to the diet of laying broiler breeder hens significantly promoted the development and metabolism of chick embryos.-Lv, Z., Fan, H., Zhang, B., Ning, C., Xing, K., Guo, Y. Dietary genistein supplementation in laying broiler breeder hens alters the development and metabolism of offspring embryos as revealed by hepatic transcriptome analysis.

Embryonic Cerebrospinal Fluid Increases Neurogenic Activity in the Brain Ventricular-Subventricular Zone of Adult Mice.

PubMed

Alonso, Maria I; Lamus, Francisco; Carnicero, Estela; Moro, Jose A; de la Mano, Anibal; Fernández, Jose M F; Desmond, Mary E; Gato, Angel

2017-01-01

Neurogenesis is a very intensive process during early embryonic brain development, becoming dramatically restricted in the adult brain in terms of extension and intensity. We have previously demonstrated the key role of embryonic cerebrospinal fluid (CSF) in developing brain neurogenic activity. We also showed that cultured adult brain neural stem cells (NSCs) remain competent when responding to the neurogenic influence of embryonic CSF. However, adult CSF loses its neurogenic inductive properties. Here, by means of an organotypic culture of adult mouse brain sections, we show that local administration of embryonic CSF in the subventricular zone (SVZ) niche is able to trigger a neurogenic program in NSCs. This leads to a significant increase in the number of non-differentiated NSCs, and also in the number of new neurons which show normal migration, differentiation and maturation. These new data reveal that embryonic CSF activates adult brain NSCs, supporting the previous idea that it contains key instructive components which could be useful in adult brain neuroregenerative strategies.

Embryonic Cerebrospinal Fluid Increases Neurogenic Activity in the Brain Ventricular-Subventricular Zone of Adult Mice

PubMed Central

Alonso, Maria I.; Lamus, Francisco; Carnicero, Estela; Moro, Jose A.; de la Mano, Anibal; Fernández, Jose M. F.; Desmond, Mary E.; Gato, Angel

2017-01-01

Neurogenesis is a very intensive process during early embryonic brain development, becoming dramatically restricted in the adult brain in terms of extension and intensity. We have previously demonstrated the key role of embryonic cerebrospinal fluid (CSF) in developing brain neurogenic activity. We also showed that cultured adult brain neural stem cells (NSCs) remain competent when responding to the neurogenic influence of embryonic CSF. However, adult CSF loses its neurogenic inductive properties. Here, by means of an organotypic culture of adult mouse brain sections, we show that local administration of embryonic CSF in the subventricular zone (SVZ) niche is able to trigger a neurogenic program in NSCs. This leads to a significant increase in the number of non-differentiated NSCs, and also in the number of new neurons which show normal migration, differentiation and maturation. These new data reveal that embryonic CSF activates adult brain NSCs, supporting the previous idea that it contains key instructive components which could be useful in adult brain neuroregenerative strategies. PMID:29311854

Quantitative analysis of serotonin secreted by human embryonic stem cells-derived serotonergic neurons via pH-mediated online stacking-CE-ESI-MRM.

PubMed

Zhong, Xuefei; Hao, Ling; Lu, Jianfeng; Ye, Hui; Zhang, Su-Chun; Li, Lingjun

2016-04-01

A CE-ESI-MRM-based assay was developed for targeted analysis of serotonin released by human embryonic stem cells-derived serotonergic neurons in a chemically defined environment. A discontinuous electrolyte system was optimized for pH-mediated online stacking of serotonin. Combining with a liquid-liquid extraction procedure, LOD of serotonin in the Krebs'-Ringer's solution by CE-ESI-MS/MS on a 3D ion trap MS was0.15 ng/mL. The quantitative results confirmed the serotonergic identity of the in vitro developed neurons and the capacity of these neurons to release serotonin in response to stimulus. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Nuclear receptor TLX regulates cell cycle progression in neural stem cells of the developing brain.

PubMed

Li, Wenwu; Sun, Guoqiang; Yang, Su; Qu, Qiuhao; Nakashima, Kinichi; Shi, Yanhong

2008-01-01

TLX is an orphan nuclear receptor that is expressed exclusively in vertebrate forebrains. Although TLX is known to be expressed in embryonic brains, the mechanism by which it influences neural development remains largely unknown. We show here that TLX is expressed specifically in periventricular neural stem cells in embryonic brains. Significant thinning of neocortex was observed in embryonic d 14.5 TLX-null brains with reduced nestin labeling and decreased cell proliferation in the germinal zone. Cell cycle analysis revealed both prolonged cell cycles and increased cell cycle exit in TLX-null embryonic brains. Increased expression of a cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin D1 provide a molecular basis for the deficiency of cell cycle progression in embryonic brains of TLX-null mice. Furthermore, transient knockdown of TLX by in utero electroporation led to precocious cell cycle exit and differentiation of neural stem cells followed by outward migration. Together these results indicate that TLX plays an important role in neural development by regulating cell cycle progression and exit of neural stem cells in the developing brain.

Nuclear Receptor TLX Regulates Cell Cycle Progression in Neural Stem Cells of the Developing Brain

PubMed Central

Li, Wenwu; Sun, Guoqiang; Yang, Su; Qu, Qiuhao; Nakashima, Kinichi; Shi, Yanhong

2008-01-01

TLX is an orphan nuclear receptor that is expressed exclusively in vertebrate forebrains. Although TLX is known to be expressed in embryonic brains, the mechanism by which it influences neural development remains largely unknown. We show here that TLX is expressed specifically in periventricular neural stem cells in embryonic brains. Significant thinning of neocortex was observed in embryonic d 14.5 TLX-null brains with reduced nestin labeling and decreased cell proliferation in the germinal zone. Cell cycle analysis revealed both prolonged cell cycles and increased cell cycle exit in TLX-null embryonic brains. Increased expression of a cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin D1 provide a molecular basis for the deficiency of cell cycle progression in embryonic brains of TLX-null mice. Furthermore, transient knockdown of TLX by in utero electroporation led to precocious cell cycle exit and differentiation of neural stem cells followed by outward migration. Together these results indicate that TLX plays an important role in neural development by regulating cell cycle progression and exit of neural stem cells in the developing brain. PMID:17901127

Type 1 and 3 inositol trisphosphate receptors are required for extra-embryonic vascular development.

PubMed

Uchida, Keiko; Nakazawa, Maki; Yamagishi, Chihiro; Mikoshiba, Katsuhiko; Yamagishi, Hiroyuki

2016-10-01

The embryonic-maternal interface of the placental labyrinth, allantois, and yolk sac are vital during embryogenesis; however, the precise mechanism underlying the vascularization of these structures remains unknown. Herein we focus on the role of inositol 1,4,5-trisphosphate (IP3) receptors (IP3R), which are intracellular Ca(2+) release channels, in placentation. Double knockout (DKO) of type 1 and 3 IP3Rs (IP3R1 and IP3R3, respectively) in mice resulted in embryonic lethality around embryonic day (E) 11.5. Because IP3R1 and IP3R3 were co-expressed in endothelial cells in the labyrinth, allantois, and yolk sac, we investigated extra-embryonic vascular development in IP3R1- and IP3R3-DKO mice. The formation of chorionic plates and yolk sac vessels seemed dysregulated around the timing of the chorio-allantoic attachment, immediately followed by the disorganization of allantoic vessels, the decreased expression of the spongiotrophoblast cell marker Tpbpa and the growth retardation of the embryos in DKO mice. Fluorescent immunohistochemistry demonstrated downregulation of a vascular endothelial marker, CD31, in labyrinth embryonic vessels and poor elongation of extra-embryonic mesoderm into the labyrinth layer in DKO placenta, whereas the branching of the DKO chorionic trophoblast was initiated. In addition, allantoic and yolk sac vessels in extra-embryonic tissues were less remodeled in DKO mice. In vitro endothelial cord formation and migration activities of cultured vascular endothelial cells derived from human umbilical vein were downregulated under the inhibition of IP3R. Our results suggest that IP3R1 and IP3R3 are required for extra-embryonic vascularization in the placenta, allantois, and yolk sac. This is the first demonstration of the essential role of IP3/IP3Rs signaling in the development of the vasculature at the embryonic-maternal interface. Copyright © 2016 Elsevier Inc. All rights reserved.

Embryonic integument and "molts" in Manduca sexta (Insecta, Lepidoptera).

PubMed

Ziese, Stefanie; Dorn, August

2003-02-01

In Manduca sexta the germ band is formed 12 h post-oviposition (p.o.) (=10% development completed) and is located above the yolk at the egg surface. The cells show a polar organization. They are engaged in the uptake and degradation of yolk globules, pinched off from the yolk cells. This process can be observed in the integumental cells during the first growth phase of the embryo that lasts until "katatrepsis," an embryonic movement that takes place at 40% development completed. At 37% development completed, the ectoderm deposits a thin membrane at its apical surface, the first embryonic membrane, which detaches immediately before katatrepsis. The second period of embryonic growth--from katatrepsis to 84 h p.o. (70% development completed)--starts with the deposition of a second embryonic membrane that is somewhat thicker than the first one and shows a trilaminar, cuticulin-like structure. Whereas the apical cell surface is largely smooth during the deposition of the first embryonic membrane, it forms microvilli during deposition of the second one. At the same time, uptake of formed yolk material ceases and the epidermal cells now contain clusters of mitochondria below the apical surface. Rough endoplasmic reticulum (RER) increases in the perinuclear region. The second embryonic membrane detaches about 63 h p.o. At 69 h p.o., a new generation of microvilli forms and islands of a typical cuticulin layer indicate the onset of the deposition of the larval cuticle. The third growth phase is characterized by a steady increase in the embryo length, the deposition of the larval procuticle, and by cuticular tanning at about 100 h p.o. Beginning at that stage, electron-lucent vesicles aggregate below the epidermal surface and are apparently released below the larval cuticle. Manduca sexta is the first holometabolous insect in which the deposition of embryonic membranes and cuticles has been examined by electron microscopy. In correspondence with hemimetabolous insects, the embryo of M. sexta secretes three covers at approximately the same developmental stage. A marked difference: the second embryonic cover, which in Hemimetabola clearly exhibits a cuticular organization, has instead a membranous, cuticulin-like structure. We see the difference as the result of an evolutionary reductional process promoted by the redundancy of embryonic covers in the egg shell. Embryonic "molts" also occur in noninsect arthropods; their phylogenetical aspects are discussed. Copyright 2002 Wiley-Liss, Inc.

In utero mouse embryonic imaging with OCT for ophthalmologic research

NASA Astrophysics Data System (ADS)

Syed, Saba H.; Larina, Irina V.; Dickinson, Mary E.; Larin, Kirill V.

2011-03-01

Live imaging of an eye during embryonic development in mammalian model is important for understanding dynamic aspects of normal and abnormal eye morphogenesis. In this study, we used Swept Source Optical Coherence Tomography (SS-OCT) for live structural imaging of mouse embryonic eye through the uterine wall. The eye structure was reconstructed in mouse embryos at 13.5 to 17.5 days post coitus (dpc). Despite the limited imaging depth of OCT in turbid tissues, we were able to visualize the whole eye globe at these stages. These results suggest that live in utero OCT imaging is a useful tool to study embryonic eye development in the mouse model.

Experimental evaluation of reproductive response to climate warming in an oviparous skink.

PubMed

Lu, Hongliang; Wang, Yong; Tang, Wenqi; DU, Weiguo

2013-06-01

The impact of climate warming on organisms is increasingly being recognized. The experimental evaluation of phenotypically plastic responses to warming is a critical step in understanding the biological effects and adaptive capacity of organisms to future climate warming. Oviparous Scincella modesta live in deeply-shaded habitats and they require low optimal temperatures during embryonic development, which makes them suitable subjects for testing the effects of warming on reproduction. We raised adult females and incubated their eggs under different thermal conditions that mimicked potential climate warming. Female reproduction, embryonic development and hatchling traits were monitored to evaluate the reproductive response to warming. Experimental warming induced females to lay eggs earlier, but it did not affect the developmental stage of embryos at oviposition or the reproductive output. The high temperatures experienced by gravid females during warming treatments reduced the incubation period and increased embryonic mortality. The locomotor performance of hatchlings was not affected by the maternal thermal environment, but it was affected by the warming treatment during embryonic development. Our results suggest that climate warming might have a profound effect on fitness-relevant traits both at embryonic and post-embryonic stages in oviparous lizards. © 2012 Wiley Publishing Asia Pty Ltd, ISZS and IOZ/CAS.

Comparative hazard analysis and toxicological modeling of diverse nanomaterials using the embryonic zebrafish (EZ) metric of toxicity

NASA Astrophysics Data System (ADS)

Harper, Bryan; Thomas, Dennis; Chikkagoudar, Satish; Baker, Nathan; Tang, Kaizhi; Heredia-Langner, Alejandro; Lins, Roberto; Harper, Stacey

2015-06-01

The integration of rapid assays, large datasets, informatics, and modeling can overcome current barriers in understanding nanomaterial structure-toxicity relationships by providing a weight-of-the-evidence mechanism to generate hazard rankings for nanomaterials. Here, we present the use of a rapid, low-cost assay to perform screening-level toxicity evaluations of nanomaterials in vivo. Calculated EZ Metric scores, a combined measure of morbidity and mortality in developing embryonic zebrafish, were established at realistic exposure levels and used to develop a hazard ranking of diverse nanomaterial toxicity. Hazard ranking and clustering analysis of 68 diverse nanomaterials revealed distinct patterns of toxicity related to both the core composition and outermost surface chemistry of nanomaterials. The resulting clusters guided the development of a surface chemistry-based model of gold nanoparticle toxicity. Our findings suggest that risk assessments based on the size and core composition of nanomaterials alone may be wholly inappropriate, especially when considering complex engineered nanomaterials. Research should continue to focus on methodologies for determining nanomaterial hazard based on multiple sub-lethal responses following realistic, low-dose exposures, thus increasing the availability of quantitative measures of nanomaterial hazard to support the development of nanoparticle structure-activity relationships.

Case Study: Organotypic human in vitro models of embryonic morphogenetic fusion

EPA Science Inventory

Morphogenetic fusion of tissues is a common event in embryonic development and disruption of fusion is associated with birth defects of the eye, heart, neural tube, phallus, palate, and other organ systems. Embryonic tissue fusion requires precise regulation of cell-cell and cell...

Assessment in Work-Based Learning: Investigating a Pedagogical Approach to Enhance Student Learning

ERIC Educational Resources Information Center

Brodie, Pandy; Irving, Kate

2007-01-01

Work-based learning (WBL) is undertaken in a wide variety of higher education contexts and is increasingly viewed as a valuable, and increasingly essential, component of both the undergraduate and postgraduate student learning experience. However, the development of rigorous pedagogies to underpin WBL and its assessment is still embryonic. This…

Effects of different feeder layers on culture of bovine embryonic stem cell-like cells in vitro.

PubMed

Cong, Shan; Cao, Guifang; Liu, Dongjun

2014-12-01

To find a suitable feeder layer is important for successful culture conditions of bovine embryonic stem cell-like cells. In this study, expression of pluripotency-related genes OCT4, SOX2 and NANOG in bovine embryonic stem cell-like cells on mouse embryonic fibroblast feeder layers at 1-5 passages were monitored in order to identify the possible reason that bovine embryonic stem cell-like cells could not continue growth and passage. Here, we developed two novel feeder layers, mixed embryonic fibroblast feeder layers of mouse and bovine embryonic fibroblast at different ratios and sources including mouse fibroblast cell lines. The bovine embryonic stem cell-like cells generated in our study displayed typical stem cell morphology and expressed specific markers such as OCT4, stage-specific embryonic antigen 1 and 4, alkaline phosphatase, SOX2, and NANOG mRNA levels. When feeder layers and cell growth factors were removed, the bovine embryonic stem cell-like cells formed embryoid bodies in a suspension culture. Furthermore, we compared the expression of the pluripotent markers during bovine embryonic stem cell-like cell in culture on mixed embryonic fibroblast feeder layers, including mouse fibroblast cell lines feeder layers and mouse embryonic fibroblast feeder layers by real-time quantitative polymerase chain reaction. Results suggested that mixed embryonic fibroblast and sources including mouse fibroblast cell lines feeder layers were more suitable for long-term culture and growth of bovine embryonic stem cell-like cells than mouse embryonic fibroblast feeder layers. The findings may provide useful experimental data for the establishment of an appropriate culture system for bovine embryonic stem cell lines.

Large-scale production of embryonic red blood cells from human embryonic stem cells.

PubMed

Olivier, Emmanuel N; Qiu, Caihong; Velho, Michelle; Hirsch, Rhoda Elison; Bouhassira, Eric E

2006-12-01

To develop a method to produce in culture large number of erythroid cells from human embryonic stem cells. Human H1 embryonic stem cells were differentiated into hematopoietic cells by coculture with a human fetal liver cell line, and the resulting CD34-positive cells were expanded in vitro in liquid culture using a three-step method. The erythroid cells produced were then analyzed by light microscopy and flow cytometry. Globin expression was characterized by quantitative reverse-transcriptase polymerase chain reaction and by high-performance liquid chromatography. CD34-positive cells produced from human embryonic stem cells could be efficiently differentiated into erythroid cells in liquid culture leading to a more than 5000-fold increase in cell number. The erythroid cells produced are similar to primitive erythroid cells present in the yolk sac of early human embryos and did not enucleate. They are fully hemoglobinized and express a mixture of embryonic and fetal globins but no beta-globin. We have developed an experimental protocol to produce large numbers of primitive erythroid cells starting from undifferentiated human embryonic stem cells. As the earliest human erythroid cells, the nucleated primitive erythroblasts, are not very well characterized because experimental material at this stage of development is very difficult to obtain, this system should prove useful to answer a number of experimental questions regarding the biology of these cells. In addition, production of mature red blood cells from human embryonic stem cells is of great potential practical importance because it could eventually become an alternate source of cell for transfusion.

Maternal dietary manganese protects chick embryos against maternal heat stress via epigenetic-activated antioxidant and anti-apoptotic abilities.

PubMed

Zhu, Yongwen; Lu, Lin; Liao, Xiudong; Li, Wenxiang; Zhang, Liyang; Ji, Cheng; Lin, Xi; Liu, Hsiao-Ching; Odle, Jack; Luo, Xugang

2017-10-27

Maternal heat stress induced the aberrant epigenetic patterns resulting in the abnormal development of offspring embryos. It is unclear whether maternal dietary manganese supplementation as an epigenetic modifier could protect the chick embryonic development against maternal heat stress via epigenetic mechanisms. To test this hypothesis using an avian model, a completely randomized design with a 2 (maternal normal and high environmental temperatures of 21 and 32°C, respectively) × 3 (maternal dietary manganese sources, the control diet without manganese supplementation and the control diet + 120 mg/kg as either inorganic or organic manganese) factorial arrangement was adopted. Maternal environmental hyperthermia increased mRNA expressions of heat shock proteins 90 and 70, cyclin-dependent kinase 6 and B-cell CLL/lymphoma 2-associated X protein displaying oxidative damage and apoptosis in the embryonic heart. Maternal environmental hyperthermia impaired the embryonic development associated with the alteration of epigenetic status, as evidenced by global DNA hypomethylation and histone 3 lysine 9 hypoacetylation in the embryonic heart. Maternal dietary manganese supplementation increased the heart anti-apoptotic gene B-cell CLL/lymphoma 2 expressions under maternal environmental hyperthermia and manganese superoxide dismutase enzyme activity in the embryonic heart. Maternal dietary organic Mn supplementation effectively eliminated the impairment of maternal environmental hyperthermia on the embryonic development. Maternal dietary manganese supplementation up-regulated manganese superoxide dismutase mRNA expression by reducing DNA methylation and increasing histone 3 lysine 9 acetylation of its promoter. It is suggested that maternal dietary manganese addition could protect the chick embryonic development against maternal heat stress via enhancing epigenetic-activated antioxidant and anti-apoptotic abilities.

Arabidopsis LEAFY COTYLEDON1 controls cell fate determination during post-embryonic development

PubMed Central

Huang, Mingkun; Hu, Yilong; Liu, Xu; Li, Yuge; Hou, Xingliang

2015-01-01

Arabidopsis LEAFY COTYLEDON1 (LEC1) transcription factor is a master regulator that shapes plant embryo development and post-embryonic seedling establishment. Loss-of-function of LEC1 alters the cotyledon identity, causing the formation of ectopic trichomes, which does not occur in wild-type seedlings, implying that LEC1 might regulate embryonic cell fate determination during post-embryonic development. To test this hypothesis, we compared the expression of trichome development-related genes between the wild-type and the lec1 mutant. We observed that transcripts of GLABROUS1 (GL1), GL2, and GL3, genes encoding the positive regulators in trichome development, were significantly upregulated, while the TRICHOMELESS1 (TCL2), ENHANCER OF TRY AND CPC1 (ETC1), and ETC2 genes, encoding the negative regulators in trichome development, were downregulated in the lec1 mutant. Furthermore, overexpression of LEC1 activated the expressions of TCL2, CAPPICE (CPC), and ETC1, resulting in production of cotyledonary leaves with no or fewer trichomes during vegetative development. In addition, we demonstrated that LEC1 interacts with TCL2 in yeast and in vitro. A genetic experiment showed that loss-of-function of GL2 rescued the ectopic trichome formation in the lec1 mutant. These findings strongly support that LEC1 regulates trichome development, providing direct evidence for the role of LEC1 in cell fate determination during post-embryonic development. PMID:26579186

HIGH INFORMATION CONTENT TOXICITY SCREENING USING MOUSE AND HUMAN STEM CELL MODELS OF ENDOCRINE DEVELOPMENT AND FUNCTION

EPA Science Inventory

The project will result in the rapid assessment of chemicals for adverse effects on the development of gametes, adipocytes, and islet B-cells; and on the adipocyte and B-cell endocrine signaling function in human and murine embryonic stem cells. Based on the data, hierarchical...

ModuleMiner - improved computational detection of cis-regulatory modules: are there different modes of gene regulation in embryonic development and adult tissues?

PubMed Central

Van Loo, Peter; Aerts, Stein; Thienpont, Bernard; De Moor, Bart; Moreau, Yves; Marynen, Peter

2008-01-01

We present ModuleMiner, a novel algorithm for computationally detecting cis-regulatory modules (CRMs) in a set of co-expressed genes. ModuleMiner outperforms other methods for CRM detection on benchmark data, and successfully detects CRMs in tissue-specific microarray clusters and in embryonic development gene sets. Interestingly, CRM predictions for differentiated tissues exhibit strong enrichment close to the transcription start site, whereas CRM predictions for embryonic development gene sets are depleted in this region. PMID:18394174

Embryonic development of the cricket Gryllus bimaculatus.

PubMed

Donoughe, Seth; Extavour, Cassandra G

2016-03-01

Extensive research into Drosophila melanogaster embryogenesis has improved our understanding of insect developmental mechanisms. However, Drosophila development is thought to be highly divergent from that of the ancestral insect and arthropod in many respects. We therefore need alternative models for arthopod development that are likely to be more representative of basally-branching clades. The cricket Gryllus bimaculatus is such a model, and currently has the most sophisticated functional genetic toolkit of any hemimetabolous insect. The existing cricket embryonic staging system is fragmentary, and it is based on morphological landmarks that are not easily visible on a live, undissected egg. To address this problem, here we present a complementary set of "egg stages" that serve as a guide for identifying the developmental progress of a cricket embryo from fertilization to hatching, based solely on the external appearance of the egg. These stages were characterized using a combination of brightfield timelapse microscopy, timed brightfield micrographs, confocal microscopy, and measurements of egg dimensions. These egg stages are particularly useful in experiments that involve egg injection (including RNA interference, targeted genome modification, and transgenesis), as injection can alter the speed of development, even in control treatments. We also use 3D reconstructions of fixed embryo preparations to provide a comprehensive description of the morphogenesis and anatomy of the cricket embryo during embryonic rudiment assembly, germ band formation, elongation, segmentation, and appendage formation. Finally, we aggregate and schematize a variety of published developmental gene expression patterns. This work will facilitate further studies on G. bimaculatus development, and serve as a useful point of reference for other studies of wild type and experimentally manipulated insect development in fields from evo-devo to disease vector and pest management. Copyright © 2015 Elsevier Inc. All rights reserved.

Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay

EPA Science Inventory

The Embryonic Stem Cell Test (EST) is an assay which evaluates xenobiotic-induced effects using three endpoints: mouse embryonic stem cell (mESC) differentiation, mESC viability, and 3T3-cell viability. Our research goal was to develop an improved high-throughput assay by establi...

The embryonic development of the cnidarian Hydractinia echinata.

PubMed

Kraus, Yulia; Flici, Hakima; Hensel, Katrin; Plickert, Günter; Leitz, Thomas; Frank, Uri

2014-01-01

With the rapid increase of the quantity of molecular data, many animals joined the ranks of the so-called 'emerging models' of Evo-Devo. One of the necessary steps in converting an emerging model into an established one is gaining comprehensive knowledge of its normal embryonic development. The marine colonial hydrozoan Hydractinia echinata - an excellent model for research on stem cells, metamorphosis, and allorecognition - has been studied for decades. Yet knowledge of its embryonic development remains fragmentary and incomplete. Here we provide a detailed account of H. echinata embryonic development using in vivo observations, histology, immunohistochemistry, and electron microscopy. Furthermore, we propose a model describing the cellular basis of the morphogenetic movements occurring during development and also reveal a functional link between canonical Wnt signaling and regional differences in the morphology of the embryo. Hydractinia embryogenesis is an example of the diversity and plasticity of hydrozoan development where multiple routes lead to the same result - the formation of a normal planula larva. © 2014 Wiley Periodicals, Inc.

The physiological basis of geographic variation in rates of embryonic development within a widespread lizard species.

PubMed

Du, Wei-Guo; Warner, Daniel A; Langkilde, Tracy; Robbins, Travis; Shine, Richard

2010-10-01

The duration of embryonic development (e.g., egg incubation period) is a critical life-history variable because it affects both the amount of time that an embryo is exposed to conditions within the nest and the seasonal timing of hatching. Variation in incubation periods among oviparous reptiles might result from variation in either the amount of embryogenesis completed before laying or the subsequent developmental rates of embryos. Selection on incubation duration could change either of those traits. We examined embryonic development of fence lizards (Sceloporus undulatus) from three populations (Indiana, Mississippi, and Florida) that occur at different latitudes and therefore experience different temperatures and season lengths. These data reveal countergradient variation: at identical temperatures in the laboratory, incubation periods were shorter for lizards from cooler areas. This variation was not related to stage at oviposition; eggs of all populations were laid at similar developmental stages. Instead, embryonic development proceeded more rapidly in cooler-climate populations, compensating for the delayed development caused by lower incubation temperatures in the field. The accelerated development appears to occur via an increase in heart mass (and, thus, stroke volume) in one population and an increase in heart rate in the other. Hence, superficially similar adaptations of embryonic developmental rate to local conditions may be generated by dissimilar proximate mechanisms.

Structure and function of gap junction proteins: role of gap junction proteins in embryonic heart development.

PubMed

Ahir, Bhavesh K; Pratten, Margaret K

2014-01-01

Intercellular (cell-to-cell) communication is a crucial and complex mechanism during embryonic heart development. In the cardiovascular system, the beating of the heart is a dynamic and key regulatory process, which is functionally regulated by the coordinated spread of electrical activity through heart muscle cells. Heart tissues are composed of individual cells, each bearing specialized cell surface membrane structures called gap junctions that permit the intercellular exchange of ions and low molecular weight molecules. Gap junction channels are essential in normal heart function and they assist in the mediated spread of electrical impulses that stimulate synchronized contraction (via an electrical syncytium) of cardiac tissues. This present review describes the current knowledge of gap junction biology. In the first part, we summarise some relevant biochemical and physiological properties of gap junction proteins, including their structure and function. In the second part, we review the current evidence demonstrating the role of gap junction proteins in embryonic development with particular reference to those involved in embryonic heart development. Genetics and transgenic animal studies of gap junction protein function in embryonic heart development are considered and the alteration/disruption of gap junction intercellular communication which may lead to abnormal heart development is also discussed.

Periods of cardiovascular susceptibility to hypoxia in embryonic american alligators (Alligator mississippiensis)

PubMed Central

Tate, Kevin B.; Rhen, Turk; Eme, John; Kohl, Zachary F.; Crossley, Janna; Elsey, Ruth M.

2016-01-01

During embryonic development, environmental perturbations can affect organisms' developing phenotype, a process known as developmental plasticity. Resulting phenotypic changes can occur during discrete, critical windows of development. Critical windows are periods when developing embryos are most susceptible to these perturbations. We have previously documented that hypoxia reduces embryo size and increases relative heart mass in American alligator, and this study identified critical windows when hypoxia altered morphological, cardiovascular function and cardiac gene expression of alligator embryos. We hypothesized that incubation in hypoxia (10% O2) would increase relative cardiac size due to cardiac enlargement rather than suppression of somatic growth. We exposed alligator embryos to hypoxia during discrete incubation periods to target windows where the embryonic phenotype is altered. Hypoxia affected heart growth between 20 and 40% of embryonic incubation, whereas somatic growth was affected between 70 and 90% of incubation. Arterial pressure was depressed by hypoxic exposure during 50–70% of incubation, whereas heart rate was depressed in embryos exposed to hypoxia during a period spanning 70–90% of incubation. Expression of Vegf and PdgfB was increased in certain hypoxia-exposed embryo treatment groups, and hypoxia toward the end of incubation altered β-adrenergic tone for arterial pressure and heart rate. It is well known that hypoxia exposure can alter embryonic development, and in the present study, we have identified brief, discrete windows that alter the morphology, cardiovascular physiology, and gene expression in embryonic American alligator. PMID:27101296

Early intrauterine embryonic development in Khawia sinensis Hsü, 1935 (Cestoda, Caryophyllidea, Lytocestidae), an invasive tapeworm of carp (Cyprinus carpio): an ultrastructural study.

PubMed

Bruňanská, Magdaléna; Mackiewicz, John S; Młocicki, Daniel; Swiderski, Zdzisław; Nebesářová, Jana

2012-02-01

Intrauterine embryonic development in the caryophyllidean tapeworm Khawia sinensis has been investigated using transmission electron microscopy and cytochemical staining with periodic acid-thiosemicarbazide-silver proteinate for glycogen. Contrary to previous light microscopy findings that reported the release of non-embryonated eggs of K. sinenesis to the external environment, the present study documents various stages of embryonation (ovoviviparity) within the intrauterine eggs of this cestode. At the initial stage of embryonic development, each fertilised oocyte is accompanied by several vitellocytes that become enclosed within the operculate, electrondense shell. Cleavage divisions result in formation of blastomeres (up to about 24 cells) of various sizes. Mitotic divisions and apparent rosette arrangment of the blastomeres, the latter atypical within the Eucestoda, are observed for the first time in the intrauterine eggs of K. sinenesis. The early embryo enclosed within the electrondense shell is surrounded by a thin membraneous layer which in some enlarged regions shows presence of nuclei. Simultaneously to multiplication and differentiation, some of the blastomeres undergo deterioration. A progressive degeneration of the vitellocytes within eggs provides nutritive reserves, including lipids, for the developing embryo. The possible significance of this atypical timing of the intrauterine embryonic development to (1) the ecology of K. sinensis and that of a recent introduction of another invasive tapeworm, the caryophyllidean Atractolytocestus huronensis Anthony, 1958 to Europe; and (2) the affiliation of caryophyllideans with other lower cestodes, are discussed.

The influence of serum substituents on serum-free Vero cell conditioned culture media manufactured from Dulbecco's modified Eagle medium in mouse embryo culture.

PubMed

Lee, Jong-Seon; Kim, Ju-Hwan; Seo, Young-Seok; Yang, Jung-Bo; Kim, Yong-Il; Kim, Hye-Jin; Lee, Ki-Hwan

2013-09-01

This study was conducted to examine the influences of supplementation of the serum substituents and available period of serum-free Vero cell conditioned media (SF-VCM) manufactured from Dulbecco's modified Eagle medium cultured with Vero cells for in vitro development of mouse preimplantation embryos. A total of 1,099 two-cell embryos collected from imprinting control region mice were cultured in SF-VCM with 10% and 20% human follicular fluid (hFF), serum substitute supplement (SSS), and serum protein substitute (SPS). Development of embryos was observed every 24 hours. Results between different groups were analyzed by chi-square test, and considered statistically significant when P-value was less than 0.05. The rates of embryonic development cultured in SF-VCM supplemented with serum substituents were significantly higher compare with serum-free group (P < 0.05). The rates of embryonic development after 48 hours (morula≤) and 96 hours (blastocyst≤) were significantly higher in 20% SSS and 10% SPS than in 20% hFF supplementation (P < 0.05). And the rates of embryonic development after 96 hours (hatching blastocyst≤) were significantly higher in 10% SPS (94.5%) than in 20% SSS (82.6%) and 20% hFF supplementation (68.5%). The rates of embryonic development according to storage period of the SF-VCM supplemented with 10% SPS showed no significant difference between control, 2 weeks and 4 weeks group. However developmental rate in 6 weeks storage group was significantly lower than other groups. The rate of embryonic development after 96 hours (hatching blastocyst≤) was significantly higher in SF-VCM supplemented with 10% SPS. And storage period of media up to 4 weeks did not affect on embryonic development.

Normal embryonic and germ cell development in mice lacking alpha 1,3-fucosyltransferase IX (Fut9) which show disappearance of stage-specific embryonic antigen 1.

PubMed

Kudo, Takashi; Kaneko, Mika; Iwasaki, Hiroko; Togayachi, Akira; Nishihara, Shoko; Abe, Kuniya; Narimatsu, Hisashi

2004-05-01

Stage-specific embryonic antigen 1 (SSEA-1), an antigenic epitope defined as a Lewis x carbohydrate structure, is expressed during the 8-cell to blastocyst stages in mouse embryos and in primordial germ cells, undifferentiated embryonic stem cells, and embryonic carcinoma cells. For many years, SSEA-1 has been implicated in the development of mouse embryos as a functional carbohydrate epitope in cell-to-cell interaction during morula compaction. In a previous study, alpha 1,3-fucosyltransferase IX (Fut9) exhibited very strong activity for the synthesis of Lewis x compared to other alpha 1,3-fucosyltransferases in an in vitro substrate specificity assay. Fut4 and Fut9 transcripts were expressed in mouse embryos. The Fut9 transcript was detected in embryonic-day-13.5 gonads containing primordial germ cells, but the Fut4 transcript was not. In order to identify the role of SSEA-1 and determine the key enzyme for SSEA-1 synthesis in vivo, we have generated Fut9-deficient (Fut9(-/-)) mice. Fut9(-/-) mice develop normally, with no gross phenotypic abnormalities, and are fertile. Immunohistochemical analysis revealed an absence of SSEA-1 expression in early embryos and primordial germ cells of Fut9(-/-) mice. Therefore, we conclude that expression of the SSEA-1 epitope in the developing mouse embryo is not essential for embryogenesis in vivo.

Human Pluripotent Stem Cell Based Developmental Toxicity Assays for Chemical Safety Screening and Systems Biology Data Generation.

PubMed

Shinde, Vaibhav; Klima, Stefanie; Sureshkumar, Perumal Srinivasan; Meganathan, Kesavan; Jagtap, Smita; Rempel, Eugen; Rahnenführer, Jörg; Hengstler, Jan Georg; Waldmann, Tanja; Hescheler, Jürgen; Leist, Marcel; Sachinidis, Agapios

2015-06-17

Efficient protocols to differentiate human pluripotent stem cells to various tissues in combination with -omics technologies opened up new horizons for in vitro toxicity testing of potential drugs. To provide a solid scientific basis for such assays, it will be important to gain quantitative information on the time course of development and on the underlying regulatory mechanisms by systems biology approaches. Two assays have therefore been tuned here for these requirements. In the UKK test system, human embryonic stem cells (hESC) (or other pluripotent cells) are left to spontaneously differentiate for 14 days in embryoid bodies, to allow generation of cells of all three germ layers. This system recapitulates key steps of early human embryonic development, and it can predict human-specific early embryonic toxicity/teratogenicity, if cells are exposed to chemicals during differentiation. The UKN1 test system is based on hESC differentiating to a population of neuroectodermal progenitor (NEP) cells for 6 days. This system recapitulates early neural development and predicts early developmental neurotoxicity and epigenetic changes triggered by chemicals. Both systems, in combination with transcriptome microarray studies, are suitable for identifying toxicity biomarkers. Moreover, they may be used in combination to generate input data for systems biology analysis. These test systems have advantages over the traditional toxicological studies requiring large amounts of animals. The test systems may contribute to a reduction of the costs for drug development and chemical safety evaluation. Their combination sheds light especially on compounds that may influence neurodevelopment specifically.

Neofunctionalization of embryonic head patterning genes facilitates the positioning of novel traits on the dorsal head of adult beetles.

PubMed

Zattara, Eduardo E; Busey, Hannah A; Linz, David M; Tomoyasu, Yoshinori; Moczek, Armin P

2016-07-13

The origin and integration of novel traits are fundamental processes during the developmental evolution of complex organisms. Yet how novel traits integrate into pre-existing contexts remains poorly understood. Beetle horns represent a spectacular evolutionary novelty integrated within the context of the adult dorsal head, a highly conserved trait complex present since the origin of insects. We investigated whether otd1/2 and six3, members of a highly conserved gene network that instructs the formation of the anterior end of most bilaterians, also play roles in patterning more recently evolved traits. Using ablation-based fate-mapping, comparative larval RNA interference (RNAi) and transcript sequencing, we found that otd1/2, but not six3, play a fundamental role in the post-embryonic formation of the adult dorsal head and head horns of Onthophagus beetles. By contrast, neither gene appears to pattern the adult head of Tribolium flour beetles even though all are expressed in the dorsal head epidermis of both Onthophagus and Tribolium We propose that, at least in beetles, the roles of otd genes during post-embryonic development are decoupled from their embryonic functions, and that potentially non-functional post-embryonic expression in the dorsal head facilitated their co-option into a novel horn-patterning network during Onthophagus evolution. © 2016 The Author(s).

Neofunctionalization of embryonic head patterning genes facilitates the positioning of novel traits on the dorsal head of adult beetles

PubMed Central

Busey, Hannah A.; Linz, David M.; Tomoyasu, Yoshinori; Moczek, Armin P.

2016-01-01

The origin and integration of novel traits are fundamental processes during the developmental evolution of complex organisms. Yet how novel traits integrate into pre-existing contexts remains poorly understood. Beetle horns represent a spectacular evolutionary novelty integrated within the context of the adult dorsal head, a highly conserved trait complex present since the origin of insects. We investigated whether otd1/2 and six3, members of a highly conserved gene network that instructs the formation of the anterior end of most bilaterians, also play roles in patterning more recently evolved traits. Using ablation-based fate-mapping, comparative larval RNA interference (RNAi) and transcript sequencing, we found that otd1/2, but not six3, play a fundamental role in the post-embryonic formation of the adult dorsal head and head horns of Onthophagus beetles. By contrast, neither gene appears to pattern the adult head of Tribolium flour beetles even though all are expressed in the dorsal head epidermis of both Onthophagus and Tribolium. We propose that, at least in beetles, the roles of otd genes during post-embryonic development are decoupled from their embryonic functions, and that potentially non-functional post-embryonic expression in the dorsal head facilitated their co-option into a novel horn-patterning network during Onthophagus evolution. PMID:27412276

Embryonic stem cells and prospects for their use in regenerative medicine approaches to motor neurone disease.

PubMed

Christou, Y A; Moore, H D; Shaw, P J; Monk, P N

2007-10-01

Human embryonic stem cells are pluripotent cells with the potential to differentiate into any cell type in the presence of appropriate stimulatory factors and environmental cues. Their broad developmental potential has led to valuable insights into the principles of developmental and cell biology and to the proposed use of human embryonic stem cells or their differentiated progeny in regenerative medicine. This review focuses on the prospects for the use of embryonic stem cells in cell-based therapy for motor neurone disease or amyotrophic lateral sclerosis, a progressive neurodegenerative disease that specifically affects upper and lower motor neurones and leads ultimately to death from respiratory failure. Stem cell-derived motor neurones could conceivably be used to replace the degenerated cells, to provide authentic substrates for drug development and screening and for furthering our understanding of disease mechanisms. However, to reliably and accurately culture motor neurones, the complex pathways by which differentiation occurs in vivo must be understood and reiterated in vitro by embryonic stem cells. Here we discuss the need for new therapeutic strategies in the treatment of motor neurone disease, the developmental processes that result in motor neurone formation in vivo, a number of experimental approaches to motor neurone production in vitro and recent progress in the application of stem cells to the treatment and understanding of motor neurone disease.

DNA methylation, an epigenetic mechanism connecting folate to healthy embryonic development and aging

USDA-ARS?s Scientific Manuscript database

Experimental studies demonstrated that maternal environmental factors including diet during early embryonic development can influence the phenotype of offspring as well as the risk of disease development at the later life. DNA methylation, an epigenetic phenomenon, has been suggested as a mechanism ...

A toolbox to explore the mechanics of living embryonic tissues

PubMed Central

Campàs, Otger

2016-01-01

The sculpting of embryonic tissues and organs into their functional morphologies involves the spatial and temporal regulation of mechanics at cell and tissue scales. Decades of in vitro work, complemented by some in vivo studies, have shown the relevance of mechanical cues in the control of cell behaviors that are central to developmental processes, but the lack of methodologies enabling precise, quantitative measurements of mechanical cues in vivo have hindered our understanding of the role of mechanics in embryonic development. Several methodologies are starting to enable quantitative studies of mechanics in vivo and in situ, opening new avenues to explore how mechanics contributes to shaping embryonic tissues and how it affects cell behavior within developing embryos. Here we review the present methodologies to study the role of mechanics in living embryonic tissues, considering their strengths and drawbacks as well as the conditions in which they are most suitable. PMID:27061360

A toolbox to explore the mechanics of living embryonic tissues.

PubMed

Campàs, Otger

2016-07-01

The sculpting of embryonic tissues and organs into their functional morphologies involves the spatial and temporal regulation of mechanics at cell and tissue scales. Decades of in vitro work, complemented by some in vivo studies, have shown the relevance of mechanical cues in the control of cell behaviors that are central to developmental processes, but the lack of methodologies enabling precise, quantitative measurements of mechanical cues in vivo have hindered our understanding of the role of mechanics in embryonic development. Several methodologies are starting to enable quantitative studies of mechanics in vivo and in situ, opening new avenues to explore how mechanics contributes to shaping embryonic tissues and how it affects cell behavior within developing embryos. Here we review the present methodologies to study the role of mechanics in living embryonic tissues, considering their strengths and drawbacks as well as the conditions in which they are most suitable. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fibroblast growth factor receptors in in vitro and in vivo chondrogenesis: relating tissue engineering using adult mesenchymal stem cells to embryonic development.

PubMed

Hellingman, Catharine A; Koevoet, Wendy; Kops, Nicole; Farrell, Eric; Jahr, Holger; Liu, Wei; Baatenburg de Jong, Robert J; Frenz, Dorothy A; van Osch, Gerjo J V M

2010-02-01

Adult mesenchymal stem cells (MSCs) are considered promising candidate cells for therapeutic cartilage and bone regeneration. Because tissue regeneration and embryonic development may involve similar pathways, understanding common pathways may lead to advances in regenerative medicine. In embryonic limb development, fibroblast growth factor receptors (FGFRs) play a role in chondrogenic differentiation. The aim of this study was to investigate and compare FGFR expression in in vivo embryonic limb development and in vitro chondrogenesis of MSCs. Our study showed that in in vitro chondrogenesis of MSCs three sequential stages can be found, as in embryonic limb development. A mesenchymal condensation (indicated by N-cadherin) is followed by chondrogenic differentiation (indicated by collagen II), and hypertrophy (indicated by collagen X). FGFR1-3 are expressed in a stage-dependent pattern during in vitro differentiation and in vivo embryonic limb development. In both models FGFR2 is clearly expressed by cells in the condensation phase. No FGFR expression was observed in differentiating and mature hyaline chondrocytes, whereas hypertrophic chondrocytes stained strongly for all FGFRs. To evaluate whether stage-specific modulation of chondrogenic differentiation in MSCs is possible with different subtypes of FGF, FGF2 and FGF9 were added to the chondrogenic medium during different stages in the culture process (early or late). FGF2 and FGF9 differentially affected the amount of cartilage formed by MSCs depending on the stage in which they were added. These results will help us understand the role of FGF signaling in chondrogenesis and find new tools to monitor and control chondrogenic differentiation.

Effects of two strobilurins (azoxystrobin and picoxystrobin) on embryonic development and enzyme activities in juveniles and adult fish livers of zebrafish (Danio rerio).

PubMed

Jia, Wei; Mao, Liangang; Zhang, Lan; Zhang, Yanning; Jiang, Hongyun

2018-09-01

Azoxystrobin and picoxystrobin are two primary strobilurin fungicides used worldwide. This study was conducted to test their effects on embryonic development and the activity of several enzyme in the zebrafish (Danio rerio). After fish eggs were separately exposed to azoxystrobin and picoxystrobin from 24 to 144 h post fertilization (hpf), the mortality, hatching, and teratogenetic rates were measured. Additionally, effects of azoxystrobin and picoxystrobin on activities of three important antioxidant enzymes [catalase (CAT), superoxide dismutase (SOD) and peroxidase (POD)] and two primary detoxification enzymes [carboxylesterase (CarE) and glutathione S-transferase (GST)] and malondialdehyde (MDA) content in zebrafish larvae (96 h) and livers of adult zebrafish of both sexes were also assessed for potential toxicity mechanisms. Based on the embryonic development test results, the mortality, hatching, and teratogenetic rates of eggs treated with azoxystrobin and picoxystrobin all showed significant dose- and time-dependent effects, and the 144-h LC 50 values of azoxystrobin and picoxystrobin were 1174.9 and 213.8 μg L -1 , respectively. In the larval zebrafish (96 h) test, activities of CAT, POD, CarE, and GST and MDA content in azoxystrobin and picoxystrobin-treated zebrafish larvae increased significantly with concentrations of the pesticides compared with those in the control. We further revealed that azoxystrobin and picoxystrobin exposure both caused significant oxidative stress in adult fish livers and the changes differed between the sexes. Our results indicated that picoxystrobin led to higher embryonic development toxicity and oxidative stress than azoxystrobin in zebrafish and the male zebrafish liver had stronger ability to detoxify than that of the females. Copyright © 2018 Elsevier Ltd. All rights reserved.

Student Learning of Early Embryonic Development via the Utilization of Research Resources from the Nematode "Caenorhabditis elegans"

ERIC Educational Resources Information Center

Lu, Fong-Mei; Eliceiri, Kevin W.; Squirrell, Jayne M.; White, John G.; Stewart, James

2008-01-01

This study was undertaken to gain insights into undergraduate students' understanding of early embryonic development, specifically, how well they comprehend the concepts of volume constancy, cell lineages, body plan axes, and temporal and spatial dimensionality in development. To study student learning, a curriculum was developed incorporating…

Embryonic development of a whirligig beetle, Dineutus mellyi, with special reference to external morphology (insecta: Coleoptera, Gyrinidae).

PubMed

Komatsu, Shintaro; Kobayashi, Yukimasa

2012-05-01

The egg morphology and successive changes of developing embryos of the whirligig beetle, Dineutus mellyi (Adephaga: Gyrinidae) are described from observations based on light and scanning electron microscopy. The egg surface is characterized by minute conical projections covering the entire egg surface, a stalk-like micropylar projection at the anterior pole of the egg, and a longitudinal split line along which the chorion is cleaved during the middle embryonic stages. The germ band or embryo is formed on the ventral egg surface, and develops on the surface throughout the egg period; thus, the egg is a superficial type, as is the case in most coleopteran species. A pair of lateral tracheal gills (LTGs) of the first abdominal segment originates from appendage-like projections arising at the lateral side of pleuropodia, and the LTGs of the second to ninth abdominal segments are arranged in a row with that of the first segment. Therefore, LTGs are structures with serial homology. The paired dorsal tracheal gills (DTGs) of the ninth abdominal segment are formed on the regions just latero-dorsal to the LTGs of this segment. Regarding the pleuropodia as the structures being homologous with thoracic legs, neither the LTGs nor DTGs are homologous with thoracic legs, but originate in the more lateral region corresponding to the future pleura of the thoracic segments. The last (10th) abdominal segment in the larva is formed by the fusion of the embryonic 10th and 11th abdominal segments. Four terminal hooks at the end of the last abdominal segment originate from two pairs of swellings on the posterior end of the embryonic 11th abdominal segment. It is proposed that the terminal hooks possibly correspond to the claws of medially fused cerci of the embryonic 11th abdominal segment. Copyright © 2011 Wiley Periodicals, Inc.

COMPARATIVE EMBRYONIC AND LARVAL DEVELOPMENTAL RESPONSES OF AN ESTUARINE SHRIMP (PALAEMONETES PUGIO) TO THE JUVENILE HORMONE AGONIST, FENOXYCARB.

EPA Science Inventory

Grass shrimp (Palaemonetes pugio) were reared separately through both embryonic and total larval development during exposure to fenoxycarb at measured concentrations of <2.2 to 888 ug L-1. A fenoxycarb concentration of 888 ug L-1significantly (p<0.05) inhibited embryonic developm...

Developmental plasticity of mitochondrial function in American alligators, Alligator mississippiensis

PubMed Central

Crossley, Janna; Elsey, Ruth M.; Dzialowski, Edward M.; Shiels, Holly A.; Crossley, Dane A.

2016-01-01

The effect of hypoxia on cellular metabolism is well documented in adult vertebrates, but information is entirely lacking for embryonic organisms. The effect of hypoxia on embryonic physiology is particularly interesting, as metabolic responses during development may have life-long consequences, due to developmental plasticity. To this end, we investigated the effects of chronic developmental hypoxia on cardiac mitochondrial function in embryonic and juvenile American alligators (Alligator mississippiensis). Alligator eggs were incubated in 21% or 10% oxygen from 20 to 90% of embryonic development. Embryos were either harvested at 90% development or allowed to hatch and then reared in 21% oxygen for 3 yr. Ventricular mitochondria were isolated from embryonic/juvenile alligator hearts. Mitochondrial respiration and enzymatic activities of electron transport chain complexes were measured with a microrespirometer and spectrophotometer, respectively. Developmental hypoxia induced growth restriction and increased relative heart mass, and this phenotype persisted into juvenile life. Embryonic mitochondrial function was not affected by developmental hypoxia, but at the juvenile life stage, animals from hypoxic incubations had lower levels of Leak respiration and higher respiratory control ratios, which is indicative of enhanced mitochondrial efficiency. Our results suggest developmental hypoxia can have life-long consequences for alligator morphology and metabolic function. Further investigations are necessary to reveal the adaptive significance of the enhanced mitochondrial efficiency in the hypoxic phenotype. PMID:27707718

Environmental and epigenetic effects upon preimplantation embryo metabolism and development

PubMed Central

Chason, Rebecca J; Csokmay, John; Segars, James H.; DeCherney, Alan H.; Armant, D. Randall

2011-01-01

In vitro fertilization has provided a unique window into the metabolic processes that drive embryonic growth and development from a fertilized ovum to a competent blastocyst. Post-fertilization development is dependent upon a dramatic reshuffling of the parental genomes during meiosis, as well as epigenetic changes that provide a new and autonomous set of instructions to guide cellular differentiation both in the embryo and beyond. While early literature focused simply on the substrates and culture conditions required for progress through embryonic development, more recent insights lead us to suggest that the surrounding environment can alter the epigenome, which can, in turn, impact embryonic metabolism and developmental competence. PMID:21741268

Identification of Estrogen Target Genes during Zebrafish Embryonic Development through Transcriptomic Analysis

EPA Science Inventory

Estrogen signaling is important for vertebrate embryonic development. Here we have used zebrafish (Danio rerio) as a vertebrate model to analyze estrogen signaling during development. Zebrafish embryos were exposed to 1 μM 17β-estradiol (E2) or vehicle from 3 hours to 4 days post...

Observations on germ band development in the cellar spider Pholcus phalangioides.

PubMed

Turetzek, Natascha; Prpic, Nikola-Michael

2016-11-01

Most recent studies of spider embryonic development have focused on representatives of the species-rich group of entelegyne spiders (over 80 % of all extant species). Embryogenesis in the smaller spider groups, however, is less well studied. Here, we describe the development of the germ band in the spider species Pholcus phalangioides, a representative of the haplogyne spiders that are phylogenetically the sister group of the entelegyne spiders. We show that the transition from radially symmetric embryonic anlage to the bilaterally symmetric germ band involves the accumulation of cells in the centre of the embryonic anlage (primary thickening). These cells then disperse all across the embryonic anlage. A secondary thickening of cells then appears in the centre of the embryonic anlage, and this thickening expands and forms the segment addition zone. We also confirm that the major part of the opisthosoma initially develops as a tube shaped structure, and its segments are then sequentially folded down on the yolk during inversion. This special mode of opisthosoma formation has not been reported for entelegyne spiders, but a more comprehensive sampling of this diverse group is necessary to decide whether this peculiarity is indeed lacking in the entelegyne spiders.

The Phosphatase PTP-PEST/PTPN12 Regulates Endothelial Cell Migration and Adhesion, but Not Permeability, and Controls Vascular Development and Embryonic Viability*

PubMed Central

Souza, Cleiton Martins; Davidson, Dominique; Rhee, Inmoo; Gratton, Jean-Philippe; Davis, Elaine C.; Veillette, André

2012-01-01

Protein-tyrosine phosphatase (PTP)-PEST (PTPN12) is ubiquitously expressed. It is essential for normal embryonic development and embryonic viability in mice. Herein we addressed the involvement of PTP-PEST in endothelial cell functions using a combination of genetic and biochemical approaches. By generating primary endothelial cells from an inducible PTP-PEST-deficient mouse, we found that PTP-PEST is not needed for endothelial cell differentiation and proliferation or for the control of endothelial cell permeability. Nevertheless, it is required for integrin-mediated adhesion and migration of endothelial cells. PTP-PEST-deficient endothelial cells displayed increased tyrosine phosphorylation of Cas, paxillin, and Pyk2, which were previously also implicated in integrin functions. By eliminating PTP-PEST in endothelial cells in vivo, we obtained evidence that expression of PTP-PEST in endothelial cells is required for normal vascular development and embryonic viability. Therefore, PTP-PEST is a key regulator of integrin-mediated functions in endothelial cells seemingly through its capacity to control Cas, paxillin, and Pyk2. This function explains at least in part the essential role of PTP-PEST in embryonic development and viability. PMID:23105101

Generation of embryos directly from embryonic stem cells by tetraploid embryo complementation reveals a role for GATA factors in organogenesis.

PubMed

Duncan, S A

2005-12-01

Gene targeting in ES (embryonic stem) cells has been used extensively to study the role of proteins during embryonic development. In the traditional procedure, this requires the generation of chimaeric mice by introducing ES cells into blastocysts and allowing them to develop to term. Once chimaeric mice are produced, they are bred into a recipient mouse strain to establish germline transmission of the allele of interest. Although this approach has been used very successfully, the breeding cycles involved are time consuming. In addition, genes that are essential for organogenesis often have roles in the formation of extra-embryonic tissues that are essential for early stages of post-implantation development. For example, mice lacking the GATA transcription factors, GATA4 or GATA6, arrest during gastrulation due to an essential role for these factors in differentiation of extra-embryonic endoderm. This lethality has frustrated the study of these factors during the development of organs such as the liver and heart. Extraembryonic defects can, however, be circumvented by generating clonal mouse embryos directly from ES cells by tetraploid complementation. Here, we describe the usefulness and efficacy of this approach using GATA factors as an example.

Serial block face-scanning electron microscopy: a tool for studying embryonic development at the cell-matrix interface.

PubMed

Starborg, Tobias; Kadler, Karl E

2015-03-01

Studies of gene regulation, signaling pathways, and stem cell biology are contributing greatly to our understanding of early embryonic vertebrate development. However, much less is known about the events during the latter half of embryonic development, when tissues comprising mostly extracellular matrix (ECM) are formed. The matrix extends far beyond the boundaries of individual cells and is refractory to study by conventional biochemical and molecular techniques; thus major gaps exist in our knowledge of the formation and three-dimensional (3D) organization of the dense tissues that form the bulk of adult vertebrates. Serial block face-scanning electron microscopy (SBF-SEM) has the ability to image volumes of tissue containing numerous cells at a resolution sufficient to study the organization of the ECM. Furthermore, whereas light microscopy was once relatively straightforward and electron microscopy was performed in specialist laboratories, the tables are turned; SBF-SEM is relatively straightforward and is becoming routine in high-end resolution studies of embryonic structures in vivo. In this review, we discuss the emergence of SBF-SEM as a tool for studying embryonic vertebrate development. © 2015 Wiley Periodicals, Inc.

Observation of human embryonic behavior in vitro by high-resolution time-lapse cinematography.

PubMed

Iwata, Kyoko; Mio, Yasuyuki

2016-07-01

Assisted reproductive technology (ART) has yielded vast amounts of information and knowledge on human embryonic development in vitro; however, still images provide limited data on dynamic changes in the developing embryos. Using our high-resolution time-lapse cinematography (hR-TLC) system, we were able to describe normal human embryonic development continuously from the fertilization process to the hatched blastocyst stage in detail. Our hR-TLC observation also showed the embryonic abnormality of a third polar body (PB)-like substance likely containing a small pronucleus being extruded and resulting in single-pronucleus (1PN) formation, while our molecular biological investigations suggested the possibility that some 1PN embryos could be diploid, carrying both maternal and paternal genomes. Furthermore, in some embryos the extruded third PB-like substance was eventually re-absorbed into the ooplasm resulting in the formation of an uneven-sized, two-PN zygote. In addition, other hR-TLC observations showed that cytokinetic failure was correlated with equal-sized, multi-nucleated blastomeres that were also observed in the embryo showing early initiation of compaction. Assessment combining our hR-TLC with molecular biological techniques enables a better understanding of embryonic development and potential improvements in ART outcomes.

Generation of genetically modified mice using CRISPR/Cas9 and haploid embryonic stem cell systems

PubMed Central

JIN, Li-Fang; LI, Jin-Song

2016-01-01

With the development of high-throughput sequencing technology in the post-genomic era, researchers have concentrated their efforts on elucidating the relationships between genes and their corresponding functions. Recently, important progress has been achieved in the generation of genetically modified mice based on CRISPR/Cas9 and haploid embryonic stem cell (haESC) approaches, which provide new platforms for gene function analysis, human disease modeling, and gene therapy. Here, we review the CRISPR/Cas9 and haESC technology for the generation of genetically modified mice and discuss the key challenges in the application of these approaches. PMID:27469251

Prolactin modulates luteal activity in the short-nosed fruit bat, Cynopterus sphinx during delayed embryonic development.

PubMed

Anuradha; Krishna, Amitabh

2017-07-01

The aim of this study was to evaluate the role of prolactin as a modulator of luteal steroidogenesis during the period of delayed embryonic development in Cynopterus sphinx. A marked decline in circulating prolactin levels was noted during the months of November through December coinciding with the period of decreased serum progesterone and delayed embryonic development. The seasonal changes in serum prolactin levels correlated positively with circulating progesterone (P) level, but inversely with circulating melatonin level during first pregnancy showing delayed development in Cynopterus sphinx. The results also showed decreased expression of prolactin receptor-short form (PRL-RS) both in the corpus luteum and in the utero-embryonic unit during the period of delayed embryonic development. Bats treated in vivo with prolactin during the period of delayed development showed significant increase in serum progesterone and estradiol levels together with significant increase in the expression of PRL-RS, luteinizing hormone receptor (LH-R), steroidogenic acute receptor protein (STAR) and 3β-hydroxysteroid dehydrogenase (3β-HSD) in the ovary. Prolactin stimulated ovarian angiogenesis (vascular endothelial growth factor) and cell survival (B-cell lymphoma 2) in vivo. Significant increases in ovarian progesterone production and the expression of prolactin-receptor, LH-R, STAR and 3β-HSD proteins were noted following the exposure of LH or prolactin in vitro during the delayed period. In conclusion, short-day associated increased melatonin level may be responsible for decreased prolactin release during November-December. The decline in prolactin level might play a role in suppressing P and estradiol-17β (E2) estradiol levels thereby causing delayed embryonic development in C. sphinx. Copyright © 2017 Elsevier Inc. All rights reserved.

Past primary sex-ratio estimates of 4 populations of Loggerhead sea turtle based on TSP durations.

NASA Astrophysics Data System (ADS)

Monsinjon, Jonathan; Kaska, Yakup; Tucker, Tony; LeBlanc, Anne Marie; Williams, Kristina; Rostal, David; Girondot, Marc

2016-04-01

Ectothermic species are supposed to be strongly affected by climate change and particularly those that exhibit temperature-dependent sex-determination (TSD). Actually, predicting the embryonic response of such organism to incubation-temperature variations in natural conditions remains challenging. In order to assess the vulnerability of sea turtles, primary sex-ratio estimates should be produced at pertinent ecological time and spatial scales. Although information on this important demographic parameter is one of the priorities for conservation purpose, accurate methodology to produce such an estimate is still lacking. The most commonly used method invocates incubation duration as a proxy for sex-ratio. This method is inappropriate because temperature influences incubation duration during all development whereas sex is influenced by temperature during only part of development. The thermosensitive period of development for sex determination (TSP) lies in the middle third of development. A model of embryonic growth must be used to define precisely the position of the TSP at non-constant incubation temperatures. The thermal reaction norm for embryonic growth rate have been estimated for 4 distinct populations of the globally distributed and threatened marine turtle Caretta caretta. A thermal reaction norm describes the pattern of phenotypic expression of a single genotype across a range of temperatures. Moreover, incubation temperatures have been reconstructed for the last 35 years using a multi-correlative model with climate temperature. After development of embryos have been modelled, we estimated the primary sex-ratio based on the duration of the TSP. Our results suggests that Loggerhead sea turtles nesting phenology is linked with the period within which both sexes can be produced in variable proportions. Several hypotheses will be discussed to explain why Caretta caretta could be more resilient to climate change than generally thought for sex determination.

Novel Method To Differentiate Human Embryonic Stem Cells Into Dopaminergic Nerve Cells | NCI Technology Transfer Center | TTC

Cancer.gov

The National Institute on Drug Abuse's Development and Plasticity Section is seeking statements of capability or interest from parties interested in licensing opportunities to further develop, evaluate, or commercialize novel methods to differentiate human embryonic stem cells into dopaminergic nerve cells. The invention described here is a novel method of differentiating human embryonic stem cells (hESCs) into dopaminergic nerve cells, which is preferable to the currently available dopaminergic differentiation techniques.

Monosaccharide uptake by erythrocytes of the embryonic and adult chicken.

PubMed

Ingermann, R L; Stock, M K; Metcalfe, J; Bissonnette, J M

1985-01-01

Rates of monosaccharide uptake by adult and 10-18 day old embryonic chicken erythrocytes were quantitated. The rate of carrier-mediated, stereospecific transport decreased 28% from day 10 to day 14 of incubation and was unchanged thereafter. At no time, however, did the rate of carrier-mediated transport by embryonic erythrocytes differ significantly from that of the adult cells. The rate of transfer by simple diffusion was 3-5 fold faster in embryonic than in adult erythrocytes. Uptake by simple diffusion decreased slightly as the embryo developed. Chronic hyperoxic incubation (70% O2) had little influence on total monosaccharide uptake by embryonic erythrocytes.

Effects of heavy ion radiation on the brain vascular system and embryonic development

NASA Technical Reports Server (NTRS)

Yang, T. C.; Tobias, C. A.

1984-01-01

The present investigation is concerned with the effects of heavy-ion radiation on the vascular system and the embryonic development, taking into account the results of experiments with neonatal rats and mouse embryos. It is found that heavy ions can be highly effective in producing brain hemorrhages and in causing body deformities. Attention is given to aspects of methodology, the induction of brain hemorrhages by X-rays and heavy ions, and the effect of iron particles on embryonic development. Reported results suggest that high linear energy transfer (LET) heavy ions can be very effective in producing developmental abnormalities.

Platelets regulate lymphatic vascular development through CLEC-2-SLP-76 signaling.

PubMed

Bertozzi, Cara C; Schmaier, Alec A; Mericko, Patricia; Hess, Paul R; Zou, Zhiying; Chen, Mei; Chen, Chiu-Yu; Xu, Bin; Lu, Min-min; Zhou, Diane; Sebzda, Eric; Santore, Matthew T; Merianos, Demetri J; Stadtfeld, Matthias; Flake, Alan W; Graf, Thomas; Skoda, Radek; Maltzman, Jonathan S; Koretzky, Gary A; Kahn, Mark L

2010-07-29

Although platelets appear by embryonic day 10.5 in the developing mouse, an embryonic role for these cells has not been identified. The SYK-SLP-76 signaling pathway is required in blood cells to regulate embryonic blood-lymphatic vascular separation, but the cell type and molecular mechanism underlying this regulatory pathway are not known. In the present study we demonstrate that platelets regulate lymphatic vascular development by directly interacting with lymphatic endothelial cells through C-type lectin-like receptor 2 (CLEC-2) receptors. PODOPLANIN (PDPN), a transmembrane protein expressed on the surface of lymphatic endothelial cells, is required in nonhematopoietic cells for blood-lymphatic separation. Genetic loss of the PDPN receptor CLEC-2 ablates PDPN binding by platelets and confers embryonic lymphatic vascular defects like those seen in animals lacking PDPN or SLP-76. Platelet factor 4-Cre-mediated deletion of Slp-76 is sufficient to confer lymphatic vascular defects, identifying platelets as the cell type in which SLP-76 signaling is required to regulate lymphatic vascular development. Consistent with these genetic findings, we observe SLP-76-dependent platelet aggregate formation on the surface of lymphatic endothelial cells in vivo and ex vivo. These studies identify a nonhemostatic pathway in which platelet CLEC-2 receptors bind lymphatic endothelial PDPN and activate SLP-76 signaling to regulate embryonic vascular development.

Correlation of Versican Expression, Accumulation, and Degradation during Embryonic Development by Quantitative Immunohistochemistry

PubMed Central

Snyder, Jessica M.; Washington, Ida M.; Birkland, Timothy; Chang, Mary Y.; Frevert, Charles W.

2015-01-01

Versican, a chondroitin sulfate proteoglycan, is important in embryonic development, and disruption of the versican gene is embryonically lethal in the mouse. Although several studies show that versican is increased in various organs during development, a focused quantitative study on versican expression and distribution during lung and central nervous system development in the mouse has not previously been performed. We tracked changes in versican (Vcan) gene expression and in the accumulation and degradation of versican. Vcan expression and quantitative immunohistochemistry performed from embryonic day (E) 11.5 to E15.5 showed peak Vcan expression at E13.5 in the lungs and brain. Quantitative mRNA analysis and versican immunohistochemistry showed differences in the expression of the versican isoforms in the embryonic lung and head. The expression of Vcan mRNA and accumulation of versican in tissues was complementary. Immunohistochemistry demonstrated co-localization of versican accumulation and degradation, suggesting distinct roles of versican deposition and degradation in embryogenesis. Very little versican mRNA or protein was found in the lungs of 12- to 16-week-old mice but versican accumulation was significantly increased in mice with Pseudomonas aeruginosa lung infection. These data suggest that versican plays an important role in fundamental, overlapping cellular processes in lung development and infection. PMID:26385570

For the Children's Sake: How To Develop a Tribal Teacher Education Program.

ERIC Educational Resources Information Center

Barden, Jack; Davis, Carol

1999-01-01

Describes the development of a teacher preparation program at Turtle Mountain Community College, which tries to balance pedagogy, subject matter, and culture in a holistic way. States that the program seeks to assess students in the program based on the outcomes they have achieved. Asserts that the hallmark of this embryonic program is…

Pío del Río-Hortega: A Visionary in the Pathology of Central Nervous System Tumors

PubMed Central

Ramon y Cajal Agüeras, Santiago

2016-01-01

The last 140 years have seen considerable advances in knowledge of central nervous system tumors. However, the main tumor types had already been described during the early years of the twentieth century. The studies of Dr. Pío del Río Hortega have been ones of the most exhaustive histology and cytology-based studies of nervous system tumors. Río Hortega's work was performed using silver staining methods, which require a high level of practical skill and were therefore difficult to standardize. His technical aptitude and interest in nervous system tumors played a key role in the establishment of his classification, which was based on cell lineage and embryonic development. Río Hortega's approach was controversial when he proposed it. Current classifications are not only based on cell type and embryonic lineage, as well as on clinical characteristics, anatomical site, and age. PMID:26973470

Acellular Mouse Kidney ECM can be Used as a Three-Dimensional Substrate to Test the Differentiation Potential of Embryonic Stem Cell Derived Renal Progenitors.

PubMed

Sambi, Manpreet; Chow, Theresa; Whiteley, Jennifer; Li, Mira; Chua, Shawn; Raileanu, Vanessa; Rogers, Ian M

2017-08-01

The development of strategies for tissue regeneration and bio-artificial organ development is based on our understanding of embryogenesis. Differentiation protocols attempt to recapitulate the signaling modalities of gastrulation and organogenesis, coupled with cell selection regimens to isolate the cells of choice. This strategy is impeded by the lack of optimal in vitro culture systems since traditional culture systems do not allow for the three-dimensional interaction between cells and the extracellular matrix. While artificial three-dimensional scaffolds are available, using the natural extracellular matrix scaffold is advantageous because it has a distinct architecture that is difficult to replicate. The adult extracellular matrix is predicted to mediate signaling related to tissue repair not embryogenesis but existing similarities between the two argues that the extracellular matrix will influence the differentiation of stem and progenitor cells. Previous studies using undifferentiated embryonic stem cells grown directly on acellular kidney ECM demonstrated that the acellular kidney supported cell growth but limited differentiation occurred. Using mouse kidney extracellular matrix and mouse embryonic stem cells we report that the extracellular matrix can support the development of kidney structures if the stem cells are first differentiated to kidney progenitor cells before being applied to the acellular organ.

Surgical inflammatory stress: the embryo takes hold of the reins again

PubMed Central

2013-01-01

The surgical inflammatory response can be a type of high-grade acute stress response associated with an increasingly complex trophic functional system for using oxygen. This systemic neuro-immune-endocrine response seems to induce the re-expression of 2 extraembryonic-like functional axes, i.e. coelomic-amniotic and trophoblastic-yolk-sac-related, within injured tissues and organs, thus favoring their re-development. Accordingly, through the up-regulation of two systemic inflammatory phenotypes, i.e. neurogenic and immune-related, a gestational-like response using embryonic functions would be induced in the patient’s injured tissues and organs, which would therefore result in their repair. Here we establish a comparison between the pathophysiological mechanisms that are produced during the inflammatory response and the physiological mechanisms that are expressed during early embryonic development. In this way, surgical inflammation could be a high-grade stress response whose pathophysiological mechanisms would be based on the recapitulation of ontogenic and phylogenetic-related functions. Thus, the ultimate objective of surgical inflammation, as a gestational process, is creating new tissues/organs for repairing the injured ones. Since surgical inflammation and early embryonic development share common production mechanisms, the factors that hamper the wound healing reaction in surgical patients could be similar to those that impair the gestational process. PMID:23374964

In utero imaging of mouse embryonic development with optical coherence tomography

NASA Astrophysics Data System (ADS)

Syed, Saba H.; Dickinson, Mary E.; Larin, Kirill V.; Larina, Irina V.

2011-03-01

Studying progression of congenital diseases in animal models can greatly benefit from live embryonic imaging Mouse have long served as a model of mammalian embryonic developmental processes, however, due to intra-uterine nature of mammalian development live imaging is challenging. In this report we present results on live mouse embryonic imaging in utero with Optical Coherence Tomography. Embryos from 12.5 through 17.5 days post-coitus (dpc) were studied through the uterine wall. In longitudinal studies, same embryos were imaged at developmental stages 13.5, 15.5 and 17.5 dpc. This study suggests that OCT can serve as a powerful tool for live mouse embryo imaging. Potentially this technique can contribute to our understanding developmental abnormalities associated with mutations, toxic drugs.

Birth characteristics and the risk of childhood rhabdomyosarcoma based on histological subtype.

PubMed

Ognjanovic, S; Carozza, S E; Chow, E J; Fox, E E; Horel, S; McLaughlin, C C; Mueller, B A; Puumala, S; Reynolds, P; Von Behren, J; Spector, L

2010-01-05

Little is known about risk factors for childhood rhabdomyosarcoma (RMS) and the histology-specific details are rare. Case-control studies formed by linking cancer and birth registries of California, Minnesota, New York, Texas and Washington, which included 583 RMS cases (363 embryonal and 85 alveolar RMS) and 57 966 randomly selected control subjects, were analysed using logistic regression. The associations of RMS (overall, and based on embryonal or alveolar histology) with birth weight across five 500 g categories (from 2000 to 4500 g) were examined using normal birth weight (2500-3999 g) as a reference. Large (>90th percentile) and small (<10th percentile) size for gestational age were calculated based on birth weight distributions in controls and were similarly examined. High birth weight increased the risk of embryonal RMS and RMS overall. Each 500 g increase in birth weight increased the risk of embryonal RMS (odds ratio (OR)=1.27, 95% confidence interval (CI)=1.14-1.42) and RMS overall (OR=1.18, 95% CI=1.09-1.29). Large size for gestational age also significantly increased the risk of embryonal RMS (OR=1.42, 95% CI=1.03-1.96). These data suggest a positive association between accelerated in utero growth and embryonal RMS, but not alveolar RMS. These results warrant cautious interpretation owing to the small number of alveolar RMS cases.

New Developmental Evidence Clarifies the Evolution of Wrist Bones in the Dinosaur–Bird Transition

PubMed Central

Botelho, João Francisco; Ossa-Fuentes, Luis; Soto-Acuña, Sergio; Smith-Paredes, Daniel; Nuñez-León, Daniel; Salinas-Saavedra, Miguel; Ruiz-Flores, Macarena; Vargas, Alexander O.

2014-01-01

From early dinosaurs with as many as nine wrist bones, modern birds evolved to develop only four ossifications. Their identity is uncertain, with different labels used in palaeontology and developmental biology. We examined embryos of several species and studied chicken embryos in detail through a new technique allowing whole-mount immunofluorescence of the embryonic cartilaginous skeleton. Beyond previous controversy, we establish that the proximal–anterior ossification develops from a composite radiale+intermedium cartilage, consistent with fusion of radiale and intermedium observed in some theropod dinosaurs. Despite previous claims that the development of the distal–anterior ossification does not support the dinosaur–bird link, we found its embryonic precursor shows two distinct regions of both collagen type II and collagen type IX expression, resembling the composite semilunate bone of bird-like dinosaurs (distal carpal 1+distal carpal 2). The distal–posterior ossification develops from a cartilage referred to as “element x,” but its position corresponds to distal carpal 3. The proximal–posterior ossification is perhaps most controversial: It is labelled as the ulnare in palaeontology, but we confirm the embryonic ulnare is lost during development. Re-examination of the fossil evidence reveals the ulnare was actually absent in bird-like dinosaurs. We confirm the proximal–posterior bone is a pisiform in terms of embryonic position and its development as a sesamoid associated to a tendon. However, the pisiform is absent in bird-like dinosaurs, which are known from several articulated specimens. The combined data provide compelling evidence of a remarkable evolutionary reversal: A large, ossified pisiform re-evolved in the lineage leading to birds, after a period in which it was either absent, nonossified, or very small, consistently escaping fossil preservation. The bird wrist provides a modern example of how developmental and paleontological data illuminate each other. Based on all available data, we introduce a new nomenclature for bird wrist ossifications. PMID:25268520

New developmental evidence clarifies the evolution of wrist bones in the dinosaur-bird transition.

PubMed

Botelho, João Francisco; Ossa-Fuentes, Luis; Soto-Acuña, Sergio; Smith-Paredes, Daniel; Nuñez-León, Daniel; Salinas-Saavedra, Miguel; Ruiz-Flores, Macarena; Vargas, Alexander O

2014-09-01

From early dinosaurs with as many as nine wrist bones, modern birds evolved to develop only four ossifications. Their identity is uncertain, with different labels used in palaeontology and developmental biology. We examined embryos of several species and studied chicken embryos in detail through a new technique allowing whole-mount immunofluorescence of the embryonic cartilaginous skeleton. Beyond previous controversy, we establish that the proximal-anterior ossification develops from a composite radiale+intermedium cartilage, consistent with fusion of radiale and intermedium observed in some theropod dinosaurs. Despite previous claims that the development of the distal-anterior ossification does not support the dinosaur-bird link, we found its embryonic precursor shows two distinct regions of both collagen type II and collagen type IX expression, resembling the composite semilunate bone of bird-like dinosaurs (distal carpal 1+distal carpal 2). The distal-posterior ossification develops from a cartilage referred to as "element x," but its position corresponds to distal carpal 3. The proximal-posterior ossification is perhaps most controversial: It is labelled as the ulnare in palaeontology, but we confirm the embryonic ulnare is lost during development. Re-examination of the fossil evidence reveals the ulnare was actually absent in bird-like dinosaurs. We confirm the proximal-posterior bone is a pisiform in terms of embryonic position and its development as a sesamoid associated to a tendon. However, the pisiform is absent in bird-like dinosaurs, which are known from several articulated specimens. The combined data provide compelling evidence of a remarkable evolutionary reversal: A large, ossified pisiform re-evolved in the lineage leading to birds, after a period in which it was either absent, nonossified, or very small, consistently escaping fossil preservation. The bird wrist provides a modern example of how developmental and paleontological data illuminate each other. Based on all available data, we introduce a new nomenclature for bird wrist ossifications.

Nitric Oxide Synthase-3 Promotes Embryonic Development of Atrioventricular Valves

PubMed Central

Liu, Yin; Lu, Xiangru; Xiang, Fu-Li; Lu, Man; Feng, Qingping

2013-01-01

Nitric oxide synthase-3 (NOS3) has recently been shown to promote endothelial-to-mesenchymal transition (EndMT) in the developing atrioventricular (AV) canal. The present study was aimed to investigate the role of NOS3 in embryonic development of AV valves. We hypothesized that NOS3 promotes embryonic development of AV valves via EndMT. To test this hypothesis, morphological and functional analysis of AV valves were performed in wild-type (WT) and NOS3−/− mice at postnatal day 0. Our data show that the overall size and length of mitral and tricuspid valves were decreased in NOS3−/− compared with WT mice. Echocardiographic assessment showed significant regurgitation of mitral and tricuspid valves during systole in NOS3−/− mice. These phenotypes were all rescued by cardiac specific NOS3 overexpression. To assess EndMT, immunostaining of Snail1 was performed in the embryonic heart. Both total mesenchymal and Snail1+ cells in the AV cushion were decreased in NOS3−/− compared with WT mice at E10.5 and E12.5, which was completely restored by cardiac specific NOS3 overexpression. In cultured embryonic hearts, NOS3 promoted transforming growth factor (TGFβ), bone morphogenetic protein (BMP2) and Snail1expression through cGMP. Furthermore, mesenchymal cell formation and migration from cultured AV cushion explants were decreased in the NOS3−/− compared with WT mice. We conclude that NOS3 promotes AV valve formation during embryonic heart development and deficiency in NOS3 results in AV valve insufficiency. PMID:24204893

Engineering human cell spheroids to model embryonic tissue fusion in vitro.

EPA Science Inventory

Epithelial-mesenchymal interactions drive embryonic fusion events during development and upon perturbation can result in birth defects. Cleft palate and neural tube defects can result from genetic defects or environmental exposures during development, yet very little is known abo...

Towards Building an AOP-based Prenatal Developmental Toxicity Ontology (CEFIC LRI Workshop - Brussels)

EPA Science Inventory

Ontologies are a way to formalize domain-specific scientific knowledge. A developmental ontology would help researchers describe the pathways and processes critical to embryonic development and provide a way to link their chemical disruption to adverse outcomes. Designing one for...

Dynamic Imaging of Mouse Embryos and Cardiodynamics in Static Culture.

PubMed

Lopez, Andrew L; Larina, Irina V

2018-01-01

The heart is a dynamic organ that quickly undergoes morphological and mechanical changes through early embryonic development. Characterizing these early moments is important for our understanding of proper embryonic development and the treatment of heart disease. Traditionally, tomographic imaging modalities and fluorescence-based microscopy are excellent approaches to visualize structural features and gene expression patterns, respectively, and connect aberrant gene programs to pathological phenotypes. However, these approaches usually require static samples or fluorescent markers, which can limit how much information we can derive from the dynamic and mechanical changes that regulate heart development. Optical coherence tomography (OCT) is unique in this circumstance because it allows for the acquisition of three-dimensional structural and four-dimensional (3D + time) functional images of living mouse embryos without fixation or contrast reagents. In this chapter, we focus on how OCT can visualize heart morphology at different stages of development and provide cardiodynamic information to reveal mechanical properties of the developing heart.

Intrinsic vs. extrinsic influences on life history expression: metabolism and parentally induced temperature influences on embryo development rate.

PubMed

Martin, Thomas E; Ton, Riccardo; Niklison, Alina

2013-06-01

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. © 2013 John Wiley & Sons Ltd/CNRS.

Intrinsic vs. extrinsic influences on life history expression: metabolism and parentally induced temperature influences on embryo development rate

USGS Publications Warehouse

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.

Single nucleotide polymorphisms in candidate genes associated with fertilizing ability of sperm and subsequent embryonic development in cattle

USDA-ARS?s Scientific Manuscript database

Fertilization and development of the preimplantation embryo is under genetic control. The goal of the current study was to test 434 single nucleotide polymorphisms (SNPs) for association with genetic variation in fertilization and early embryonic development. The approach was to produce embryos from...

Maternal transfer of methimazole and effects on thyroid hormone availability in embryonic tissues.

PubMed

Van Herck, Stijn L J; Geysens, Stijn; Bald, Edward; Chwatko, Grazyna; Delezie, Evelyne; Dianati, Elham; Ahmed, R G; Darras, Veerle M

2013-07-01

Methimazole (MMI) is an anti-thyroid drug used in the treatment of chronic hyperthyroidism. There is, however, some debate about its use during pregnancy as MMI is known to cross the mammalian placenta and reach the developing foetus. A similar problem occurs in birds, where MMI is deposited in the egg and taken up by the developing embryo. To investigate whether maternally derived MMI can have detrimental effects on embryonic development, we treated laying hens with MMI (0.03% in drinking water) and measured total and reduced MMI contents in the tissues of hens and embryos at different stages of development. In hens, MMI was selectively increased in the thyroid gland, while its levels in the liver and especially brain remained relatively low. Long-term MMI treatment induced a pronounced goitre with a decrease in thyroxine (T₄) content but an increase in thyroidal 3,5,3'-triiodothyronine (T₃) content. This resulted in normal T₃ levels in tissues except in the brain. In chicken embryos, MMI levels were similar in the liver and brain. They gradually decreased during development but always remained above those in the corresponding maternal tissues. Contrary to the situation in hens, T₄ availability was only moderately affected in embryos. Peripheral T₃ levels were reduced in 14-day-old embryos but normal in 18-day-old embryos, while brain T₃ content was decreased at all embryonic stages tested. We conclude that all embryonic tissues are exposed to relatively high doses of MMI and its oxidised metabolites. The effect of maternal MMI treatment on embryonic thyroid hormone availability is most pronounced for brain T₃ content, which is reduced throughout the embryonic development period.

Neural Differentiation of Embryonic Stem Cells In Vitro: A Road Map to Neurogenesis in the Embryo

PubMed Central

Abranches, Elsa; Silva, Margarida; Pradier, Laurent; Schulz, Herbert; Hummel, Oliver; Henrique, Domingos; Bekman, Evguenia

2009-01-01

Background The in vitro generation of neurons from embryonic stem (ES) cells is a promising approach to produce cells suitable for neural tissue repair and cell-based replacement therapies of the nervous system. Available methods to promote ES cell differentiation towards neural lineages attempt to replicate, in different ways, the multistep process of embryonic neural development. However, to achieve this aim in an efficient and reproducible way, a better knowledge of the cellular and molecular events that are involved in the process, from the initial specification of neuroepithelial progenitors to their terminal differentiation into neurons and glial cells, is required. Methodology/Principal Findings In this work, we characterize the main stages and transitions that occur when ES cells are driven into a neural fate, using an adherent monolayer culture system. We established improved conditions to routinely produce highly homogeneous cultures of neuroepithelial progenitors, which organize into neural tube-like rosettes when they acquire competence for neuronal production. Within rosettes, neuroepithelial progenitors display morphological and functional characteristics of their embryonic counterparts, namely, apico-basal polarity, active Notch signalling, and proper timing of production of neurons and glia. In order to characterize the global gene activity correlated with each particular stage of neural development, the full transcriptome of different cell populations that arise during the in vitro differentiation protocol was determined by microarray analysis. By using embryo-oriented criteria to cluster the differentially expressed genes, we define five gene expression signatures that correlate with successive stages in the path from ES cells to neurons. These include a gene signature for a primitive ectoderm-like stage that appears after ES cells enter differentiation, and three gene signatures for subsequent stages of neural progenitor development, from an early stage that follows neural induction to a final stage preceding terminal differentiation. Conclusions/Significance Overall, our work confirms and extends the cellular and molecular parallels between monolayer ES cell neural differentiation and embryonic neural development, revealing in addition novel aspects of the genetic network underlying the multistep process that leads from uncommitted cells to differentiated neurons. PMID:19621087

DNA methylation analysis of the gene CDKN2B in Gallus gallus (chicken).

PubMed

Gryzińska, Magdalena; Andraszek, Katarzyna; Jocek, Grzegorz

2013-01-01

Methylation is an epigenetic modification of DNA affecting gene expression without changing the structure of nucleotides. It plays a crucial role in the embryonic and post-embryonic development of living organisms. Methylation level is tissue and species-specific and changes with age. The study was aimed at identifying the methylation of the CDKN2B gene situated at locus bar in Polbar chickens on the 6th and 18th day of embryonic development using the MSP (methylation-specific PCR) method. Methylation was not detected in the promoter region of gene CDKN2B on the 6th and 18th day of embryonic development. As one of the five genes responsible for melanine activity in melanocytes and highly active, it can contribute to the production of this pigment. The present research broadens the current knowledge of the chicken epigenome and the mechanism of autosexing in birds.

Alterations to embryonic serotonin change aggression and fearfulness

USDA-ARS?s Scientific Manuscript database

Prenatal environment, including maternal hormones, affects the development of the serotonin (5-HT) system, with long-lasting effects on mood and behavioral exhibition in children and adults. The chicken provides a unique animal model to study the effects of embryonic development on childhood and ado...

In silico Testing of Environmental Impact on Embryonic Vascular Development

EPA Science Inventory

Understanding risks to embryonic development from exposure to environmental chemicals is a significant challenge given the diverse chemical landscape and paucity of data for most of these compounds. EPA’s Virtual Embryo project is building in silico models of morphogenesis to tes...

Developing an Experimental Model of Vascular Toxicity in Embryonic Zebrafish

EPA Science Inventory

Developing an Experimental Model of Vascular Toxicity in Embryonic Zebrafish Tamara Tal, Integrated Systems Toxicology Division, U.S. EPA Background: There are tens of thousands of chemicals that have yet to be fully evaluated for their toxicity by validated in vivo testing ...

Lipid content and fatty acid profile during lake whitefish embryonic development at different incubation temperatures.

PubMed

Mueller, Casey A; Doyle, Liam; Eme, John; Manzon, Richard G; Somers, Christopher M; Boreham, Douglas R; Wilson, Joanna Y

2017-01-01

Lipids serve as energy sources, structural components, and signaling molecules during fish embryonic development, and utilization of lipids may vary with temperature. Embryonic energy utilization under different temperatures is an important area of research in light of the changing global climate. Therefore, we examined percent lipid content and fatty acid profiles of lake whitefish (Coregonus clupeaformis) throughout embryonic development at three incubation temperatures. We sampled fertilized eggs and embryos at gastrulation, eyed and fin flutter stages following chronic incubation at temperatures of 1.8, 4.9 and 8.0°C. Hatchlings were also sampled following incubation at temperatures of 3.3, 4.9 and 8.0°C. Fertilized eggs had an initial high percentage of dry mass composed of lipid (percent lipid content; ~29%) consisting of ~20% saturated fatty acids (SFA), ~32% monounsaturated fatty acids (MUFA), ~44% polyunsaturated fatty acids (PUFA), and 4% unidentified. The most abundant fatty acids were 16:0, 16:1, 18:1(n-9c), 20:4(n-6), 20:5(n-3) and 22:6(n-3). This lipid profile matches that of other cold-water fish species. Percent lipid content increased during embryonic development, suggesting protein or other yolk components were preferentially used for energy. Total percentage of MUFA decreased during development, which indicated MUFA were the primary lipid catabolized for energy during embryonic development. Total percentage of PUFA increased during development, driven largely by an increase in 22:6(n-3). Temperature did not influence percent lipid content or percent MUFA at any development stage, and had inconsistent effects on percent SFA and percent PUFA during development. Thus, lake whitefish embryos appear to be highly adapted to low temperatures, and do not alter lipids in response to temperature within their natural incubation conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Impacts of maternal dietary protein intake on fetal survival, growth, and development.

PubMed

Herring, Cassandra M; Bazer, Fuller W; Johnson, Gregory A; Wu, Guoyao

2018-03-01

Maternal nutrition during gestation, especially dietary protein intake, is a key determinant in embryonic survival, growth, and development. Low maternal dietary protein intake can cause embryonic losses, intra-uterine growth restriction, and reduced postnatal growth due to a deficiency in specific amino acids that are important for cell metabolism and function. Of note, high maternal dietary protein intake can also result in intra-uterine growth restriction and embryonic death, due to amino acid excesses, as well as the toxicity of ammonia, homocysteine, and H 2 S that are generated from amino acid catabolism. Maternal protein nutrition has a pronounced impact on fetal programming and alters the expression of genes in the fetal genome. As a precursor to the synthesis of molecules (e.g. nitric oxide, polyamines, and creatine) with cell signaling and metabolic functions, L-arginine (Arg) is essential during pregnancy for growth and development of the conceptus. With inadequate maternal dietary protein intake, Arg and other important amino acids are deficient in mother and fetus. Dietary supplementation of Arg during gestation has been effective in improving embryonic survival and development of the conceptus in many species, including humans, pigs, sheep, mice, and rats. Both the balance among amino acids and their quantity are critical for healthy pregnancies and offspring. Impact statement This review aims at: highlighting adverse effects of elevated levels of ammonia in mother or fetus on embryonic/fetal survival, growth, and development; helping nutritionists and practitioners to understand the mechanisms whereby elevated levels of ammonia in mother or fetus results in embryonic/fetal death, growth restriction, and developmental abnormalities; and bringing, into the attention of nutritionists and practitioners, the problems of excess or inadequate dietary intake of protein or amino acids on pregnancy outcomes in animals and humans. The article provides new, effective means to improve embryonic/fetal survival and growth in mammals.

Effects of temperature on embryonic and early larval growth and development in the rough-skinned newt (Taricha granulosa).

PubMed

Smith, Geoffrey D; Hopkins, Gareth R; Mohammadi, Shabnam; M Skinner, Heather; Hansen, Tyler; Brodie, Edmund D; French, Susannah S

2015-07-01

We investigated the effects of temperature on the growth and development of embryonic and early larval stages of a western North American amphibian, the rough-skinned newt (Taricha granulosa). We assigned newt eggs to different temperatures (7, 14, or 21°C); after hatching, we re-assigned the newt larvae into the three different temperatures. Over the course of three to four weeks, we measured total length and developmental stage of the larvae. Our results indicated a strong positive relationship over time between temperature and both length and developmental stage. Importantly, individuals assigned to cooler embryonic temperatures did not achieve the larval sizes of individuals from the warmer embryonic treatments, regardless of larval temperature. Our investigation of growth and development at different temperatures demonstrates carry-over effects and provides a more comprehensive understanding of how organisms respond to temperature changes during early development. Copyright © 2015 Elsevier Ltd. All rights reserved.

A 3D magnetic tissue stretcher for remote mechanical control of embryonic stem cell differentiation.

PubMed

Du, Vicard; Luciani, Nathalie; Richard, Sophie; Mary, Gaëtan; Gay, Cyprien; Mazuel, François; Reffay, Myriam; Menasché, Philippe; Agbulut, Onnik; Wilhelm, Claire

2017-09-12

The ability to create a 3D tissue structure from individual cells and then to stimulate it at will is a major goal for both the biophysics and regenerative medicine communities. Here we show an integrated set of magnetic techniques that meet this challenge using embryonic stem cells (ESCs). We assessed the impact of magnetic nanoparticles internalization on ESCs viability, proliferation, pluripotency and differentiation profiles. We developed magnetic attractors capable of aggregating the cells remotely into a 3D embryoid body. This magnetic approach to embryoid body formation has no discernible impact on ESC differentiation pathways, as compared to the hanging drop method. It is also the base of the final magnetic device, composed of opposing magnetic attractors in order to form embryoid bodies in situ, then stretch them, and mechanically stimulate them at will. These stretched and cyclic purely mechanical stimulations were sufficient to drive ESCs differentiation towards the mesodermal cardiac pathway.The development of embryoid bodies that are responsive to external stimuli is of great interest in tissue engineering. Here, the authors culture embryonic stem cells with magnetic nanoparticles and show that the presence of magnetic fields could affect their aggregation and differentiation.

Visualisation of chicken macrophages using transgenic reporter genes: insights into the development of the avian macrophage lineage.

PubMed

Balic, Adam; Garcia-Morales, Carla; Vervelde, Lonneke; Gilhooley, Hazel; Sherman, Adrian; Garceau, Valerie; Gutowska, Maria W; Burt, David W; Kaiser, Pete; Hume, David A; Sang, Helen M

2014-08-01

We have generated the first transgenic chickens in which reporter genes are expressed in a specific immune cell lineage, based upon control elements of the colony stimulating factor 1 receptor (CSF1R) locus. The Fms intronic regulatory element (FIRE) within CSF1R is shown to be highly conserved in amniotes and absolutely required for myeloid-restricted expression of fluorescent reporter genes. As in mammals, CSF1R-reporter genes were specifically expressed at high levels in cells of the macrophage lineage and at a much lower level in granulocytes. The cell lineage specificity of reporter gene expression was confirmed by demonstration of coincident expression with the endogenous CSF1R protein. In transgenic birds, expression of the reporter gene provided a defined marker for macrophage-lineage cells, identifying the earliest stages in the yolk sac, throughout embryonic development and in all adult tissues. The reporter genes permit detailed and dynamic visualisation of embryonic chicken macrophages. Chicken embryonic macrophages are not recruited to incisional wounds, but are able to recognise and phagocytose microbial antigens. © 2014. Published by The Company of Biologists Ltd.

De novo formation of nucleoli in developing mouse embryos originating from enucleolated zygotes.

PubMed

Kyogoku, Hirohisa; Fulka, Josef; Wakayama, Teruhiko; Miyano, Takashi

2014-06-01

The large, compact oocyte nucleoli, sometimes referred to as nucleolus precursor bodies (NPBs), are essential for embryonic development in mammals; in their absence, the oocytes complete maturation and can be fertilized, but no nucleoli are formed in the zygote or embryo, leading to developmental failure. It has been convincingly documented that zygotes inherit the oocyte nucleolar material and form NPBs again in pronuclei. It is commonly accepted that during early embryonic development, the original compact zygote NPBs gradually transform into reticulated nucleoli of somatic cells. Here, we show that zygote NPBs are not required for embryonic and full-term development in the mouse. When NPBs were removed from late-stage zygotes by micromanipulation, the enucleolated zygotes developed to the blastocyst stage and, after transfer to recipients, live pups were obtained. We also describe de novo formation of nucleoli in developing embryos. After removal of NPBs from zygotes, they formed new nucleoli after several divisions. These results indicate that the zygote NPBs are not used in embryonic development and that the nucleoli in developing embryos originate from de novo synthesized materials. © 2014. Published by The Company of Biologists Ltd.

Impaired cardiac energy metabolism in embryos lacking adrenergic stimulation

PubMed Central

Baker, Candice N.; Gidus, Sarah A.; Price, George F.; Peoples, Jessica N. R.

2014-01-01

As development proceeds from the embryonic to fetal stages, cardiac energy demands increase substantially, and oxidative phosphorylation of ADP to ATP in mitochondria becomes vital. Relatively little, however, is known about the signaling mechanisms regulating the transition from anaerobic to aerobic metabolism that occurs during the embryonic period. The main objective of this study was to test the hypothesis that adrenergic hormones provide critical stimulation of energy metabolism during embryonic/fetal development. We examined ATP and ADP concentrations in mouse embryos lacking adrenergic hormones due to targeted disruption of the essential dopamine β-hydroxylase (Dbh) gene. Embryonic ATP concentrations decreased dramatically, whereas ADP concentrations rose such that the ATP/ADP ratio in the adrenergic-deficient group was nearly 50-fold less than that found in littermate controls by embryonic day 11.5. We also found that cardiac extracellular acidification and oxygen consumption rates were significantly decreased, and mitochondria were significantly larger and more branched in adrenergic-deficient hearts. Notably, however, the mitochondria were intact with well-formed cristae, and there was no significant difference observed in mitochondrial membrane potential. Maternal administration of the adrenergic receptor agonists isoproterenol or l-phenylephrine significantly ameliorated the decreases in ATP observed in Dbh−/− embryos, suggesting that α- and β-adrenergic receptors were effective modulators of ATP concentrations in mouse embryos in vivo. These data demonstrate that adrenergic hormones stimulate cardiac energy metabolism during a critical period of embryonic development. PMID:25516547

Clonal analysis of Notch1-expressing cells reveals the existence of unipotent stem cells that retain long-term plasticity in the embryonic mammary gland.

PubMed

Lilja, Anna M; Rodilla, Veronica; Huyghe, Mathilde; Hannezo, Edouard; Landragin, Camille; Renaud, Olivier; Leroy, Olivier; Rulands, Steffen; Simons, Benjamin D; Fre, Silvia

2018-06-01

Recent lineage tracing studies have revealed that mammary gland homeostasis relies on unipotent stem cells. However, whether and when lineage restriction occurs during embryonic mammary development, and which signals orchestrate cell fate specification, remain unknown. Using a combination of in vivo clonal analysis with whole mount immunofluorescence and mathematical modelling of clonal dynamics, we found that embryonic multipotent mammary cells become lineage-restricted surprisingly early in development, with evidence for unipotency as early as E12.5 and no statistically discernable bipotency after E15.5. To gain insights into the mechanisms governing the switch from multipotency to unipotency, we used gain-of-function Notch1 mice and demonstrated that Notch activation cell autonomously dictates luminal cell fate specification to both embryonic and basally committed mammary cells. These functional studies have important implications for understanding the signals underlying cell plasticity and serve to clarify how reactivation of embryonic programs in adult cells can lead to cancer.

Transcriptional Profiling of Ectoderm Specification to Keratinocyte Fate in Human Embryonic Stem Cells

PubMed Central

Tadeu, Ana Mafalda Baptista; Lin, Samantha; Hou, Lin; Chung, Lisa; Zhong, Mei; Zhao, Hongyu; Horsley, Valerie

2015-01-01

In recent years, several studies have shed light into the processes that regulate epidermal specification and homeostasis. We previously showed that a broad-spectrum γ–secretase inhibitor DAPT promoted early keratinocyte specification in human embryonic stem cells triggered to undergo ectoderm specification. Here, we show that DAPT accelerates human embryonic stem cell differentiation and induces expression of the ectoderm protein AP2. Furthermore, we utilize RNA sequencing to identify several candidate regulators of ectoderm specification including those involved in epithelial and epidermal development in human embryonic stem cells. Genes associated with transcriptional regulation and growth factor activity are significantly enriched upon DAPT treatment during specification of human embryonic stem cells to the ectoderm lineage. The human ectoderm cell signature identified in this study contains several genes expressed in ectodermal and epithelial tissues. Importantly, these genes are also associated with skin disorders and ectodermal defects, providing a platform for understanding the biology of human epidermal keratinocyte development under diseased and homeostatic conditions. PMID:25849374

[Acceleration of Embryonic Development of Pinus sibirica Trees with a One-Year Reproductive Cycle].

PubMed

Tret'yakova, I N; Lukina, N V

2016-01-01

The study of the formation of embryonic structures in Pinus sibirica forms with a one-year reproductive cycle showed that the acceleration of the embryonic process manifested itself as a reduction of the coenocytic stage of the female gametophyte development (1.5 months instead of 1 year). The egg was not fertilized because of the asynchronous maturation of male and female gametophytes. Seeds without embryos were formed. We assumed that the acceleration of the reproductive process in Pinus sibirica was caused by a mutation in the female generative organs.

Generation of Functional Thyroid Tissue Using 3D-Based Culture of Embryonic Stem Cells.

PubMed

Antonica, Francesco; Kasprzyk, Dominika Figini; Schiavo, Andrea Alex; Romitti, Mírian; Costagliola, Sabine

2017-01-01

During the last decade three-dimensional (3D) cultures of pluripotent stem cells have been intensively used to understand morphogenesis and molecular signaling important for the embryonic development of many tissues. In addition, pluripotent stem cells have been shown to be a valid tool for the in vitro modeling of several congenital or chronic human diseases, opening new possibilities to study their physiopathology without using animal models. Even more interestingly, 3D culture has proved to be a powerful and versatile tool to successfully generate functional tissues ex vivo. Using similar approaches, we here describe a protocol for the generation of functional thyroid tissue using mouse embryonic stem cells and give all the details and references for its characterization and analysis both in vitro and in vivo. This model is a valid approach to study the expression and the function of genes involved in the correct morphogenesis of thyroid gland, to elucidate the mechanisms of production and secretion of thyroid hormones and to test anti-thyroid drugs.

The transcriptomes of novel marmoset monkey embryonic stem cell lines reflect distinct genomic features

PubMed Central

Debowski, Katharina; Drummer, Charis; Lentes, Jana; Cors, Maren; Dressel, Ralf; Lingner, Thomas; Salinas-Riester, Gabriela; Fuchs, Sigrid; Sasaki, Erika; Behr, Rüdiger

2016-01-01

Embryonic stem cells (ESCs) are useful for the study of embryonic development. However, since research on naturally conceived human embryos is limited, non-human primate (NHP) embryos and NHP ESCs represent an excellent alternative to the corresponding human entities. Though, ESC lines derived from naturally conceived NHP embryos are still very rare. Here, we report the generation and characterization of four novel ESC lines derived from natural preimplantation embryos of the common marmoset monkey (Callithrix jacchus). For the first time we document derivation of NHP ESCs derived from morula stages. We show that quantitative chromosome-wise transcriptome analyses precisely reflect trisomies present in both morula-derived ESC lines. We also demonstrate that the female ESC lines exhibit different states of X-inactivation which is impressively reflected by the abundance of the lncRNA X inactive-specific transcript (XIST). The novel marmoset ESC lines will promote basic primate embryo and ESC studies as well as preclinical testing of ESC-based regenerative approaches in NHP. PMID:27385131

The transcriptomes of novel marmoset monkey embryonic stem cell lines reflect distinct genomic features.

PubMed

Debowski, Katharina; Drummer, Charis; Lentes, Jana; Cors, Maren; Dressel, Ralf; Lingner, Thomas; Salinas-Riester, Gabriela; Fuchs, Sigrid; Sasaki, Erika; Behr, Rüdiger

2016-07-07

Embryonic stem cells (ESCs) are useful for the study of embryonic development. However, since research on naturally conceived human embryos is limited, non-human primate (NHP) embryos and NHP ESCs represent an excellent alternative to the corresponding human entities. Though, ESC lines derived from naturally conceived NHP embryos are still very rare. Here, we report the generation and characterization of four novel ESC lines derived from natural preimplantation embryos of the common marmoset monkey (Callithrix jacchus). For the first time we document derivation of NHP ESCs derived from morula stages. We show that quantitative chromosome-wise transcriptome analyses precisely reflect trisomies present in both morula-derived ESC lines. We also demonstrate that the female ESC lines exhibit different states of X-inactivation which is impressively reflected by the abundance of the lncRNA X inactive-specific transcript (XIST). The novel marmoset ESC lines will promote basic primate embryo and ESC studies as well as preclinical testing of ESC-based regenerative approaches in NHP.

Effect of micro-vibration culture system on embryo development.

PubMed

Hur, Yong Soo; Park, Jeong Hyun; Ryu, Eun Kyung; Park, Sung Jin; Lee, Jun Ho; Lee, Soo Hee; Yoon, Jung; Yoon, San Hyun; Hur, Chang Young; Lee, Won Don; Lim, Jin Ho

2013-06-01

Micro-vibration culture system was examined to determine the effects on mouse and human embryo development and possible improvement of clinical outcomes in poor responders. The embryonic development rates and cell numbers of blastocysts were compared between a static culture group (n = 178) and a micro-vibration culture group (n = 181) in mice. The embryonic development rates and clinical results were compared between a static culture group (n = 159 cycles) and a micro-vibration culture group (n = 166 cycles) in poor responders. A micro-vibrator was set at a frequency of 42 Hz, 5 s/60 min duration for mouse and human embryo development. The embryonic development rate was significantly improved in the micro-vibration culture group in mice (p < 0.05). The cell numbers of mouse blastocysts were significantly higher in the micro-vibration group than in the static culture group (p < 0.05). In the poor responders, the rate of high grade embryos was not significantly improved in the micro-vibration culture group on day 3. However, the optimal embryonic development rate on day 5 was improved in the micro-vibration group, and the total pregnancy rate and implantation rate were significantly higher in the micro-vibration group than in the static culture group (p < 0.05). Micro-vibration culture methods have a beneficial effect on embryonic development in mouse embryos. In poor responders, the embryo development rate was improved to a limited extent under the micro-vibration culture conditions, but the clinical results were significantly improved.

Cardiotoxicity evaluation using human embryonic stem cells and induced pluripotent stem cell-derived cardiomyocytes.

PubMed

Zhao, Qi; Wang, Xijie; Wang, Shuyan; Song, Zheng; Wang, Jiaxian; Ma, Jing

2017-03-09

Cardiotoxicity remains an important concern in drug discovery. Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have become an attractive platform to evaluate cardiotoxicity. However, the consistency between human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in prediction of cardiotoxicity has yet to be elucidated. Here we screened the toxicities of four representative drugs (E-4031, isoprenaline, quinidine, and haloperidol) using both hESC-CMs and hiPSC-CMs, combined with an impedance-based bioanalytical method. It showed that both hESC-CMs and hiPSC-CMs can recapitulate cardiotoxicity and identify the effects of well-characterized compounds. The combined platform of hPSC-CMs and an impedance-based bioanalytical method could improve preclinical cardiotoxicity screening, holding great potential for increasing drug development accuracy.

The laboratory curse: variation in temperature stimulates embryonic development and shortens diapause

USDA-ARS?s Scientific Manuscript database

An ongoing biological debate is the difference in trait expression in continuous versus cycling temperature regimes, but are even daily cycling temperatures sufficient to generate natural expression of traits? We compared embryonic development and the duration of diapause for Mormon cricket eggs in...

[Embryonic stem cells and therapeutic cloning].

PubMed

Sunde, A; Eftedal, I

2001-08-30

Increased interest in the therapeutic use of human stem cells has emerged following significant progress in ongoing research. The cloning of a sheep, the isolation of human embryonic stem cells, and the discovery that adult stem cells may be reprogrammed taken together give substance to hopes that novel principles of treatment may be developed for a variety of serious conditions. Embryonic stem cells are derived from pre-embryos at the blastocyst stage and may give rise to all bodily tissues and cells. Animal models have demonstrated that embryonic stem cells when transplanted into adult hosts may differentiate and develop into cells and tissues applicable for treatment of a variety of conditions, including Parkinson's disease, multiple sclerosis, spinal injuries, cardiac stroke and cancer. Transplanted embryonic stem cells are exposed to immune reactions similar to those acting on organ transplants, hence immunosuppression of the recipient is generally required. It is, however, possible to obtain embryonic stem cells that are genetically identical to the patient's own cells by means of therapeutic cloning techniques. The nucleus from a somatic cell is transferred into an egg after removal of the egg's own genetic material. Under specific condition the egg will use genetic information from the somatic cell in organising the formation of a blastocyst which in turn generates embryonic stem cells. These cells have a genetic composition identical to that of the patient and are suitable for stem cell therapy.

Early embryonic sensitivity to cyclophosphamide in cardiac differentiation from human embryonic stem cells.

PubMed

Zhu, Ming-Xia; Zhao, Jin-Yuan; Chen, Gui-An; Guan, Li

2011-09-01

hESCs (human embryonic stem cells) can differentiate into tissue derivatives of all three germ layers in vitro and mimic the development of the embryo in vivo. In this study, we have investigated the potential of an hESC-based assay for the detection of toxicity to cardiac differentiation in embryonic development. First of all, we developed the protocol of cardiac induction from hESCs according to our previous work and distinguished cardiac precursor cells and late mature cardiomyocytes from differentiated cells, demonstrated by the Q-PCR (quantitative real-time PCR), immunocytochemistry and flow cytometry analysis. In order to test whether CPA (cyclophosphamide) induces developmental and cellular toxicity in the human embryo, we exposed the differentiating cells from hESCs to CPA (a well-known proteratogen) at different stages. We have found that a high concentration of CPA could inhibit cardiac differentiation of hESCs. Two separate exposure intervals were used to determine the effects of CPA on cardiac precursor cells and late mature cardiomyocytes respectively. The cardiac precursor cells were sensitive to CPA in non-cytotoxic concentrations for the expression of the cardiac-specific mRNA markers Nkx2.5 (NK2 transcription factor related, locus 5), GATA-4 (GATA binding protein 4 transcription factor) and TNNT2 (troponin T type 2). Non-cytotoxic CPA concentrations did not affect the mRNA markers' expression in late mature cardiomyocytes, indicating that cardiac precursors were more sensitive to CPA than late cardiomyocytes in cardiogenesis. We set up the in vitro developmental toxicity test model so as to reduce the number of test animals and expenses without compromising the safety of consumers and patients. Furthermore, such in vitro methods may be possibly suited to test a large number of chemicals than the classical employed in vivo tests.

The effects of incubation temperature and experimental design on heart rates of lizard embryos.

PubMed

Hulbert, Austin C; Mitchell, Timothy S; Hall, Joshua M; Guiffre, Cassia M; Douglas, Danielle C; Warner, Daniel A

2017-08-01

Many studies of phenotypic plasticity alter environmental conditions during embryonic development, yet only measure phenotypes at the neonatal stage (after embryonic development). However, measuring aspects of embryo physiology enhances our understanding of how environmental factors immediately affect embryos, which aids our understanding of developmental plasticity. While current research on reptile developmental plasticity has demonstrated that fluctuating incubation temperatures affect development differently than constant temperatures, most research on embryo physiology is still performed with constant temperature experiments. In this study, we noninvasively measured embryonic heart rates of the brown anole (Anolis sagrei), across ecologically relevant fluctuating temperatures. We incubated eggs under temperatures measured from potential nests in the field and examined how heart rates change through a diel cycle and throughout embryonic development. We also evaluated how experimental design (e.g., repeated vs. single measures designs, constant vs. fluctuating temperatures) and different protocols (e.g., removing eggs from incubators) might influence heart rate. We found that heart rates were correlated with daily temperature and increased through development. Our findings suggest that experimenters have reasonable flexibility in choosing an experimental design to address their questions; however, some aspects of design and protocol can potentially influence estimations of heart rates. Overall, we present the first ecologically relevant measures of anole embryonic heart rates and provide recommendations for experimental designs for future experiments. © 2017 Wiley Periodicals, Inc.

Redundant functions of I-BAR family members, IRSp53 and IRTKS, are essential for embryonic development

PubMed Central

Chou, Ai Mei; Sem, Kai Ping; Lam, Wei Jun; Ahmed, Sohail; Lim, Chin Yan

2017-01-01

The insulin receptor substrate of 53 kDa, IRSp53, is an adaptor protein that works with activated GTPases, Cdc42 and Rac, to modulate actin dynamics and generate membrane protrusions in response to cell signaling. Adult mice that lack IRSp53 fail to regulate synaptic plasticity and exhibit hippocampus-associated learning deficiencies. Here, we show that 60% of IRSp53 null embryos die at mid to late gestation, indicating a vital IRSp53 function in embryonic development. We find that IRSp53 KO embryos displayed pleiotropic phenotypes such as developmental delay, oligodactyly and subcutaneous edema, and died of severely impaired cardiac and placental development. We further show that double knockout of IRSp53 and its closest family member, IRTKS, resulted in exacerbated placental abnormalities, particularly in spongiotrophoblast differentiation and development, giving rise to complete embryonic lethality. Hence, our findings demonstrate a hitherto under-appreciated IRSp53 function in embryonic development, and further establish an essential genetic interaction between IRSp53 and IRTKS in placental formation. PMID:28067313

Maternal thyroid hormones are essential for neural development in zebrafish.

PubMed

Campinho, Marco A; Saraiva, João; Florindo, Claudia; Power, Deborah M

2014-07-01

Teleost eggs contain an abundant store of maternal thyroid hormones (THs), and early in zebrafish embryonic development, all the genes necessary for TH signaling are expressed. Nonetheless the function of THs in embryonic development remains elusive. To test the hypothesis that THs are fundamental for zebrafish embryonic development, an monocarboxilic transporter 8 (Mct8) knockdown strategy was deployed to prevent maternal TH uptake. Absence of maternal THs did not affect early specification of the neural epithelia but profoundly modified later dorsal specification of the brain and spinal cord as well as specific neuron differentiation. Maternal THs acted upstream of pax2a, pax7, and pax8 genes but downstream of shha and fgf8a signaling. The lack of inhibitory spinal cord interneurons and increased motoneurons in the mct8 morphants is consistent with their stiff axial body and impaired mobility. The mct8 mutations are associated with X-linked mental retardation in humans, and the cellular and molecular consequences of MCT8 knockdown during embryonic development in zebrafish provides new insight into the potential role of THs in this condition.

Maternal Thyroid Hormones Are Essential for Neural Development in Zebrafish

PubMed Central

Saraiva, João; Florindo, Claudia; Power, Deborah M.

2014-01-01

Teleost eggs contain an abundant store of maternal thyroid hormones (THs), and early in zebrafish embryonic development, all the genes necessary for TH signaling are expressed. Nonetheless the function of THs in embryonic development remains elusive. To test the hypothesis that THs are fundamental for zebrafish embryonic development, an monocarboxilic transporter 8 (Mct8) knockdown strategy was deployed to prevent maternal TH uptake. Absence of maternal THs did not affect early specification of the neural epithelia but profoundly modified later dorsal specification of the brain and spinal cord as well as specific neuron differentiation. Maternal THs acted upstream of pax2a, pax7, and pax8 genes but downstream of shha and fgf8a signaling. The lack of inhibitory spinal cord interneurons and increased motoneurons in the mct8 morphants is consistent with their stiff axial body and impaired mobility. The mct8 mutations are associated with X-linked mental retardation in humans, and the cellular and molecular consequences of MCT8 knockdown during embryonic development in zebrafish provides new insight into the potential role of THs in this condition. PMID:24877564

The role of platelets during reproduction.

PubMed

Isermann, Berend; Nawroth, Peter P

2006-01-01

The availability of mice with defined defects within the hemostatic system enabled researchers to identify a role the coagulation system for embryonic and placental development. However, the role of platelets during development has only recently been experimentally addressed, giving some insight into potential functions of platelets during development. Thus, a quantitative embryonic platelet defect (severe thrombopenia secondary to NF-E2 deficiency) is associated with an embryonic growth retardation and reduced vascularisation of the placenta. Maternal platelet deficiency is associated with placental hemorrhage, which, however, does not impair embryonic or maternal survival. In vitro studies established that platelets or platelet conditioned medium regulate the invasive properties of human extravillous trophoblast cells and induce a phenotypical switch of trophoblast cells. These data imply that platelets are of relevance during placentation. Conversely, platelets and the formation of platelet-fibrin aggregates are dispensable for the development of the embryo proper, establishing that the lethal phenotypes observed in some embryo slacking coagulation regulators does not result from an inability to form platelet-fibrin aggregates, but likely reflects altered protease dependent signaling during vascular development.

The role of flow in the morphodynamics of embryonic heart

NASA Astrophysics Data System (ADS)

Gharib, Morteza

2017-11-01

Nature has shown us that some hearts do not require valves to achieve unidirectional flow. In its earliest stages, the vertebrate heart consists of a primitive tube that drives blood through a simple vascular network nourishing tissues and other developing organ systems. We have shown that in the case of the embryonic zebrafish heart, an elastic wave resonance mechanism based on impedance mismatches at the boundaries of the heart tube is the likely mechanism responsible for the valveless pumping behavior. When functioning normally, mature heart valves prevent intracardiac retrograde blood flow; before valves develop there is considerable regurgitation, resulting in oscillatory flow between the atrium and ventricle. We show that reversing flows are particularly strong stimuli to endothelial cells and that heart valves form as a developmental response to oscillatory blood flow through the maturing heart.

The essential role of endogenous ghrelin in growth hormone expression during zebrafish adenohypophysis development.

PubMed

Li, Xi; He, Jiangyan; Hu, Wei; Yin, Zhan

2009-06-01

Ghrelin, a multifunctional hormone, including potent GH stimulation activity, has been suggested to be important during embryonic development. Expression of ghrelin has been confirmed in the zebrafish pancreas during embryonic stages. Interfering with ghrelin function using two specific antisense morpholino oligonucleotides causes defects during zebrafish embryonic development. In ghrelin morphants the expression of GH was abolished in zebrafish somatotropes, whereas the expression patterns of the other key molecules involved in hypothalamic-pituitary development and distinct pituitary hormones genes remain largely intact at the appropriate time during zebrafish adenohypophysis development. Effective rescue of the ghrelin morphants with exogenous ghrelin mRNA showed that the correct gene had been targeted. Moreover, by analyzing the efficiencies of the ghrelin morphants rescue experiments with various forms of exogenous mutant ghrelin mRNAs, we also demonstrated the essentiality of the form acyl-ghrelin on GH stimulation during zebrafish adenohypophysis development. Our in vivo experiments, for the first time, also provided evidence of the existence of functional obestatin in the C-terminal part of zebrafish proghrelin peptides. Our research here has demonstrated that zebrafish is a unique model for functional studies of endogenous ghrelin, especially during embryonic development.

Imaging of murine embryonic cardiovascular development using optical coherence tomography (Conference Presentation)

NASA Astrophysics Data System (ADS)

Huang, Yongyang; Degenhardt, Karl R.; Astrof, Sophie; Zhou, Chao

2016-03-01

We have demonstrated the capability of spectral domain optical coherence tomography (SDOCT) system to image full development of mouse embryonic cardiovascular system. Monitoring morphological changes of mouse embryonic heart occurred in different embryonic stages helps identify structural or functional cardiac anomalies and understand how these anomalies lead to congenital heart diseases (CHD) present at birth. In this study, mouse embryo hearts ranging from E9.5 to E15.5 were prepared and imaged in vitro. A customized spectral domain OCT system was used for imaging, with a central wavelength of 1310nm, spectral bandwidth of ~100nm and imaging speed of 47kHz A-scans/s. Axial resolution of this system was 8.3µm in air, and transverse resolution was 6.2 µm with 5X objective. Key features of mouse embryonic cardiovascular development such as vasculature remodeling into circulatory system, separation of atria and ventricles and emergence of valves could be clearly seen in three-dimensional OCT images. Optical clearing was applied to overcome the penetration limit of OCT system. With high resolution, fast imaging speed, 3D imaging capability, OCT proves to be a promising biomedical imaging modality for developmental biology studies, rivaling histology and micro-CT.

Dihydroartemisinin promotes angiogenesis during the early embryonic development of zebrafish

PubMed Central

Ba, Qian; Duan, Juan; Tian, Jia-qiang; Wang, Zi-liang; Chen, Tao; Li, Xiao-guang; Chen, Pei-zhan; Wu, Song-jie; Xiang, Li; Li, Jing-quan; Chu, Rui-ai; Wang, Hui

2013-01-01

Aim: To investigate the embryotoxicity of dihydroartemisinin (DHA), the main active metabolite of artemisinin, in zebrafish, and explore the corresponding mechanisms. Methods: The embryos of wild type and TG (flk1:GFP) transgenic zebrafish were exposed to DHA. Developmental phenotypes of the embryos were observed. Development of blood vessels was directly observed in living embryos of TG (flk1:GFP) transgenic zebrafish under fluorescence microscope. The expression of angiogenesis marker genes vegfa, flk1, and flt1 in the embryos was detected using real-time PCR and RNA in situ hybridization assays. Results: Exposure to DHA (1–10 mg/L) dose-dependently caused abnormal zebrafish embryonic phenotypes in the early developmental stage. Furthermore, exposure to DHA (10 mg/L) resulted in more pronounced embryonic angiogenesis in TG (flk1:GFP) zebrafish line. Exposure to DHA (10 mg/L) significantly increased the mRNA expression of vegfa, flk1, and flt1 in the embryos. Knockdown of the flk1 protein partially blocked the effects of DHA on embryogenesis. Conclusion: DHA causes abnormal embryonic phenotypes and promotes angiogenesis in zebrafish early embryonic development, demonstrating the potential embryotoxicity of DHA. PMID:23708556

Incubation times of dinosaur eggs via embryonic metabolism

NASA Astrophysics Data System (ADS)

Lee, Scott A.

2016-08-01

The incubation times for the eggs of 21 dinosaurs are determined from an estimate of their embyronic metabolic rate and the mass of the hatchlings via a mass growth model based on conservation of energy. Embryos in extant birds and crocodiles are studied in order to determine the best model for embryonic metabolism and growth. These results are used to develop a theoretical model that predicts the incubation times of an egg. This model is applied to dinosaur eggs and provides a unique window into dinosaur reproduction. The dinosaurs studied come from both Saurischia and Ornithischia. The incubation times vary from about 28 days for Archaeopteryx lithographica to about 76 days for Alamosaurus sanjuanensis.

Incubation times of dinosaur eggs via embryonic metabolism.

PubMed

Lee, Scott A

2016-08-01

The incubation times for the eggs of 21 dinosaurs are determined from an estimate of their embyronic metabolic rate and the mass of the hatchlings via a mass growth model based on conservation of energy. Embryos in extant birds and crocodiles are studied in order to determine the best model for embryonic metabolism and growth. These results are used to develop a theoretical model that predicts the incubation times of an egg. This model is applied to dinosaur eggs and provides a unique window into dinosaur reproduction. The dinosaurs studied come from both Saurischia and Ornithischia. The incubation times vary from about 28 days for Archaeopteryx lithographica to about 76 days for Alamosaurus sanjuanensis.

A Single-Cell Roadmap of Lineage Bifurcation in Human ESC Models of Embryonic Brain Development.

PubMed

Yao, Zizhen; Mich, John K; Ku, Sherman; Menon, Vilas; Krostag, Anne-Rachel; Martinez, Refugio A; Furchtgott, Leon; Mulholland, Heather; Bort, Susan; Fuqua, Margaret A; Gregor, Ben W; Hodge, Rebecca D; Jayabalu, Anu; May, Ryan C; Melton, Samuel; Nelson, Angelique M; Ngo, N Kiet; Shapovalova, Nadiya V; Shehata, Soraya I; Smith, Michael W; Tait, Leah J; Thompson, Carol L; Thomsen, Elliot R; Ye, Chaoyang; Glass, Ian A; Kaykas, Ajamete; Yao, Shuyuan; Phillips, John W; Grimley, Joshua S; Levi, Boaz P; Wang, Yanling; Ramanathan, Sharad

2017-01-05

During human brain development, multiple signaling pathways generate diverse cell types with varied regional identities. Here, we integrate single-cell RNA sequencing and clonal analyses to reveal lineage trees and molecular signals underlying early forebrain and mid/hindbrain cell differentiation from human embryonic stem cells (hESCs). Clustering single-cell transcriptomic data identified 41 distinct populations of progenitor, neuronal, and non-neural cells across our differentiation time course. Comparisons with primary mouse and human gene expression data demonstrated rostral and caudal progenitor and neuronal identities from early brain development. Bayesian analyses inferred a unified cell-type lineage tree that bifurcates between cortical and mid/hindbrain cell types. Two methods of clonal analyses confirmed these findings and further revealed the importance of Wnt/β-catenin signaling in controlling this lineage decision. Together, these findings provide a rich transcriptome-based lineage map for studying human brain development and modeling developmental disorders. Copyright © 2017 Elsevier Inc. All rights reserved.

Embryonic common snapping turtles (Chelydra serpentina) preferentially regulate intracellular tissue pH during acid-base challenges.

PubMed

Shartau, Ryan B; Crossley, Dane A; Kohl, Zachary F; Brauner, Colin J

2016-07-01

The nests of embryonic turtles naturally experience elevated CO2 (hypercarbia), which leads to increased blood PCO2  and a respiratory acidosis, resulting in reduced blood pH [extracellular pH (pHe)]. Some fishes preferentially regulate tissue pH [intracellular pH (pHi)] against changes in pHe; this has been proposed to be associated with exceptional CO2 tolerance and has never been identified in amniotes. As embryonic turtles may be CO2 tolerant based on nesting strategy, we hypothesized that they preferentially regulate pHi, conferring tolerance to severe acute acid-base challenges. This hypothesis was tested by investigating pH regulation in common snapping turtles (Chelydra serpentina) reared in normoxia then exposed to hypercarbia (13 kPa PCO2 ) for 1 h at three developmental ages: 70% and 90% of incubation, and yearlings. Hypercarbia reduced pHe but not pHi, at all developmental ages. At 70% of incubation, pHe was depressed by 0.324 pH units while pHi of brain, white muscle and lung increased; heart, liver and kidney pHi remained unchanged. At 90% of incubation, pHe was depressed by 0.352 pH units but heart pHi increased with no change in pHi of other tissues. Yearlings exhibited a pHe reduction of 0.235 pH units but had no changes in pHi of any tissues. The results indicate common snapping turtles preferentially regulate pHi during development, but the degree of response is reduced throughout development. This is the first time preferential pHi regulation has been identified in an amniote. These findings may provide insight into the evolution of acid-base homeostasis during development of amniotes, and vertebrates in general. © 2016. Published by The Company of Biologists Ltd.

Development of a 3D co-culture model using human stem cells for studying embryonic palatal fusion.

EPA Science Inventory

Morphogenetic tissue fusion is a critical and complex event in embryonic development and failure of this event leads to birth defects, such as cleft palate. Palatal fusion requires adhesion and subsequent dissolution of the medial epithelial layer of the mesenchymal palatal shelv...

Developing predictions of in vivo developmental toxicity of ToxCast chemicals using mouse embryonic stem cells.

EPA Science Inventory

Developing predictions of in vivo developmental toxicity of ToxCast chemicals using mouse embryonic stem cells S. Hunter, M. Rosen, M. Hoopes, H. Nichols, S. Jeffay, K. Chandler1, Integrated Systems Toxicology Division, National Health and Environmental Effects Research Labor...

Periconceptional maternal one-carbon biomarkers are associated with embryonic development according to the Carnegie stages.

PubMed

Parisi, F; Rousian, M; Koning, A H J; Willemsen, S P; Cetin, I; Steegers-Theunissen, R P M

2017-03-01

Is periconceptional maternal one-carbon (I-C) metabolism associated with embryonic morphological development in non-malformed ongoing pregnancies? Serum vitamin B12, red blood cell (RBC) folate and plasma total homocysteine (tHcy) are associated with embryonic development according to the Carnegie stages. Derangements in maternal I-C metabolism affect reproductive and pregnancy outcomes, as well as future health of the offspring. Between 2010 and 2014, women with singleton ongoing pregnancies were enrolled in a prospective periconceptional cohort study. A total of 234 pregnancies, including 138 spontaneous or IUI pregnancies with strict pregnancy dating and 96 pregnancies derived from IVF, ICSI or cryopreserved embryo transfer (IVF/ICSI pregnancies), underwent longitudinal transvaginal three-dimensional ultrasound (3D US) scans from 6+0 up to 10+2 weeks of gestation. Carnegie stages were defined using internal and external morphologic criteria in a virtual reality system. Maternal venous blood samples were collected at enrollment for serum vitamin B12, RBC folate and plasma tHcy assessment. Associations between biomarker concentrations and longitudinal Carnegie stages were investigated using linear mixed models. We performed a median of three 3D US scans per pregnancy (range 1-5) resulting in 600 good quality data sets for the Carnegie stage annotation (80.5%). Vitamin B12 was positively associated with embryonic development in the total study population (β = 0.001 (95% CI: 0.000; 0.002), P < 0.05) and in the subgroup of strictly dated spontaneous pregnancies (β = 0.002 (95% CI: 0.001; 0.003), P < 0.05). Low vitamin B12 concentrations (-2SD, 73.4 pmol/l) were associated with delayed embryonic development by 1.4 days (95% CI: 1.3-1.4) compared with high concentrations (+2SD, 563.1 pmol/l). RBC folate was positively associated with Carnegie stages only in IVF/ICSI pregnancies (β = 0.001 (95% CI: 0.0005; 0.0015), P < 0.05). In this group, low RBC folate concentrations (-2SD, 875.4 nmol/l) were associated with a 1.8-day delay (95% CI: 1.7-1.8) in development compared with high concentrations (+2SD, 2119.9 nmol/l). tHcy was negatively associated with embryonic development in the total study population (β = -0.08 (95% CI: -0.14; -0.02), P < 0.01), as well as in the IVF/ICSI subgroup (β = -0.08 (95% CI: -0.15; -0.01), P < 0.05). High tHcy concentrations (+2SD, 10.4 µmol/l) were associated with a delay of 1.6 days (95% CI: 1.5-1.7) in embryonic development compared with low concentrations (-2SD, 3.0 µmol/l). The study was performed in a tertiary care center, resulting in high rates of folic acid supplement use and comorbidity that may reduce the external validity of our findings. In periconceptional care, maternal I-C biomarkers should be taken into account as predictors of embryonic morphological development. Combining embryonic size measurements with morphological assessment could better define normal embryonic development. The work was funded by the Department of Obstetrics and Gynaecology, Erasmus MC, University Medical Centre, Rotterdam, The Netherlands. RPMST is CSO of the startup company Slimmere Zorg and CEO of eHealth Care Solutions. The authors declare no conflicts of interest. Not applicable. © The Author 2017. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

Viviparity in high-altitude Phrynocephalus lizards is adaptive because embryos cannot fully develop without maternal thermoregulation.

PubMed

Wang, Zheng; Lu, Hong-Liang; Ma, Li; Ji, Xiang

2014-03-01

Viviparous Phrynocephalus lizards (Agamidae) are mainly restricted to the Qinghai-Tibet Plateau of China. In this study, we used Phrynocephalus vlangalii females kept under seven thermal regimes for the whole gestation period to test the hypothesis that viviparity in high-altitude Phrynocephalus lizards is adaptive because embryos cannot fully develop without maternal thermoregulation. All females at 24 °C and 93% of the females at 28 °C failed to give birth or produced stillborns, and proportionally fewer females gave birth at 29 or 35 °C than at 32 °C. Though the daily temperatures encountered were unsuitable for embryonic development, 95% of the females in nature and 89% of the females thermoregulating in the laboratory gave birth. There was no shift in the thermal preferences of females when they were pregnant. Although thermal conditions inside natural burrows were unsuitable for embryonic development, mass and sprint speed were both greater in neonates produced in nature. Our data show that (1) long-term exposure of P. vlangalii embryos to temperatures outside the range of 29-35 °C may result in the failure of development, but daily or short-term exposure may not necessarily increase embryonic mortality; (2) low gestation temperatures slow but do not arrest embryonic development, and females produce high-quality offspring in the shortest possible time by maintaining gestation temperatures close to the upper thermal limit for embryonic development; and (3) viviparity is currently adaptive at high elevations because embryos in nature cannot fully develop without relying on maternal thermoregulation. Our data validate the hypothesis tested.

Diverging functions of Scr between embryonic and post-embryonic development in a hemimetabolous insect, Oncopeltus fasciatus.

PubMed

Chesebro, John; Hrycaj, Steven; Mahfooz, Najmus; Popadić, Aleksandar

2009-05-01

Hemimetabolous insects undergo an ancestral mode of development in which embryos hatch into first nymphs that resemble miniature adults. While recent studies have shown that homeotic (hox) genes establish segmental identity of first nymphs during embryogenesis, no information exists on the function of these genes during post-embryogenesis. To determine whether and to what degree hox genes influence the formation of adult morphologies, we performed a functional analysis of Sex combs reduced (Scr) during post-embryonic development in Oncopeltus fasciatus. The main effect was observed in prothorax of Scr-RNAi adults, and ranged from significant alterations in its size and shape to a near complete transformation of its posterior half toward a T2-like identity. Furthermore, while the consecutive application of Scr-RNAi at both of the final two post-embryonic stages (fourth and fifth) did result in formation of ectopic wings on T1, the individual applications at each of these stages did not. These experiments provide two new insights into evolution of wings. First, the role of Scr in wing repression appears to be conserved in both holo- and hemimetabolous insects. Second, the prolonged Scr-depletion (spanning at least two nymphal stages) is both necessary and sufficient to restart wing program. At the same time, other structures that were previously established during embryogenesis are either unaffected (T1 legs) or display only minor changes (labium) in adults. These observations reveal a temporal and spatial divergence of Scr roles during embryonic (main effect in labium) and post-embryonic (main effect in prothorax) development.

Stage specific requirement of platelet-derived growth factor receptor-α in embryonic development.

PubMed

Qian, Chen; Wong, Carol Wing Yan; Wu, Zhongluan; He, Qiuming; Xia, Huimin; Tam, Paul Kwong Hang; Wong, Kenneth Kak Yuen; Lui, Vincent Chi Hang

2017-01-01

Platelet-derived growth factor receptor alpha (PDGFRα) is a cell-surface receptor tyrosine kinase for platelet-derived growth factors. Correct timing and level of Pdgfra expression is crucial for embryo development, and deletion of Pdgfra caused developmental defects of multiple endoderm and mesoderm derived structures, resulting in a complex phenotypes including orofacial cleft, spina bifida, rib deformities, and omphalocele in mice. However, it is not clear if deletion of Pdgfra at different embryonic stages differentially affects these structures. To address the temporal requirement of Pdgfra in embryonic development. We have deleted the Pdgfra in Pdgfra-expressing tissues at different embryonic stages in mice, examined and quantified the developmental anomalies. Current study showed that (i) conditional deletion of Pdgfra at different embryonic days (between E7.5 and E10.5) resulted in orofacial cleft, spina bifida, rib cage deformities, and omphalocele, and (ii) the day of Pdgfra deletion influenced the combinations, incidence and severities of these anomalies. Deletion of Pdgfra caused apoptosis of Pdgfra-expressing tissues, and developmental defects of their derivatives. Orofacial cleft, spina bifida and omphalocele are among the commonest skeletal and abdominal wall defects of newborns, but their genetic etiologies are largely unknown. The remarkable resemblance of our conditional Pdgfra knockout embryos to theses human congenital anomalies, suggesting that dysregulated PDGFRA expression could cause these anomalies in human. Future work should aim at defining (a) the regulatory elements for the expression of the human PDGFRA during embryonic development, and (b) if mutations / sequence variations of these regulatory elements cause these anomalies.

Diverging functions of Scr between embryonic and post-embryonic development in a hemimetabolous insect, Oncopeltus fasciatus

PubMed Central

Chesebro, John; Hrycaj, Steven; Mahfooz, Najmus; Popadić, Aleksandar

2009-01-01

Hemimetabolous insects undergo an ancestral mode of development in which embryos hatch into first nymphs that resemble miniature adults. While recent studies have shown that homeotic (hox) genes establish segmental identity of first nymphs during embryogenesis, no information exists on the function of these genes during post-embryogenesis. To determine whether and to what degree hox genes influence the formation of adult morphologies, we performed a functional analysis of Sex combs reduced (Scr) during post-embryonic development in Oncopeltus fasciatus. The main effect was observed in prothorax of Scr-RNAi adults, and ranged from significant alterations in its size and shape to a near complete transformation of its posterior half toward a T2-like identity. Furthermore, while the consecutive application of Scr-RNAi at both of the final two post-embryonic stages (fourth and fifth) did result in formation of ectopic wings on T1, the individual applications at each of these stages did not. These experiments provide two new insights into evolution of wings. First, the role of Scr in wing repression appears to be conserved in both holo- and hemimetabolous insects. Second, the prolonged Scr-depletion (spanning at least two nymphal stages) is both necessary and sufficient to restart wing program. At the same time, other structures that were previously established during embryogenesis are either unaffected (T1 legs) or display only minor changes (labium) in adults. These observations reveal a temporal and spatial divergence of Scr roles during embryonic (main effect in labium) and post-embryonic (main effect in prothorax) development. PMID:19382295

A new method of estimating thermal performance of embryonic development rate yields accurate prediction of embryonic age in wild reptile nests.

PubMed

Rollinson, Njal; Holt, Sarah M; Massey, Melanie D; Holt, Richard C; Nancekivell, E Graham; Brooks, Ronald J

2018-05-01

Temperature has a strong effect on ectotherm development rate. It is therefore possible to construct predictive models of development that rely solely on temperature, which have applications in a range of biological fields. Here, we leverage a reference series of development stages for embryos of the turtle Chelydra serpentina, which was described at a constant temperature of 20 °C. The reference series acts to map each distinct developmental stage onto embryonic age (in days) at 20 °C. By extension, an embryo taken from any given incubation environment, once staged, can be assigned an equivalent age at 20 °C. We call this concept "Equivalent Development", as it maps the development stage of an embryo incubated at a given temperature to its equivalent age at a reference temperature. In the laboratory, we used the concept of Equivalent Development to estimate development rate of embryos of C. serpentina across a series of constant temperatures. Using these estimates of development rate, we created a thermal performance curve measured in units of Equivalent Development (TPC ED ). We then used the TPC ED to predict developmental stage of embryos in several natural turtle nests across six years. We found that 85% of the variation of development stage in natural nests could be explained. Further, we compared the predictive accuracy of the model based on the TPC ED to the predictive accuracy of a degree-day model, where development is assumed to be linearly related to temperature and the amount of accumulated heat is summed over time. Information theory suggested that the model based on the TPC ED better describes variation in developmental stage in wild nests than the degree-day model. We suggest the concept of Equivalent Development has several strengths and can be broadly applied. In particular, studies on temperature-dependent sex determination may be facilitated by the concept of Equivalent Development, as development age maps directly onto the developmental series of the organism, allowing critical periods of sex determination to be delineated without invasive sampling, even under fluctuating temperature. Copyright © 2018 Elsevier Ltd. All rights reserved.

Establishment of mouse embryonic stem cells from isolated blastomeres and whole embryos using three derivation methods

PubMed Central

González, Sheyla; Ibáñez, Elena

2010-01-01

Purpose The aim of the present study is to compare three previously described mouse embryonic stem cell derivation methods to evaluate the influence of culture conditions, number of isolated blastomeres and embryonic stage in the derivation process. Methods Three embryonic stem cell derivation methods: standard, pre-adhesion and defined culture medium method, were compared in the derivation from isolated blastomeres and whole embryos at 4- and 8-cell stages. Results A total of 200 embryonic stem cell lines were obtained with an efficiency ranging from 1.9% to 72%. Conclusions Using either isolated blastomeres or whole embryos, the highest rates of mouse embryonic stem cell establishment were achieved with the defined culture medium method and efficiencies increased as development progressed. Using isolated blastomeres, efficiencies increased in parallel to the proportion of the embryo volume used to start the derivation process. PMID:20862536

The primary role of zebrafish nanog is in extra-embryonic tissue.

PubMed

Gagnon, James A; Obbad, Kamal; Schier, Alexander F

2018-01-09

The role of the zebrafish transcription factor Nanog has been controversial. It has been suggested that Nanog is primarily required for the proper formation of the extra-embryonic yolk syncytial layer (YSL) and only indirectly regulates gene expression in embryonic cells. In an alternative scenario, Nanog has been proposed to directly regulate transcription in embryonic cells during zygotic genome activation. To clarify the roles of Nanog, we performed a detailed analysis of zebrafish nanog mutants. Whereas zygotic nanog mutants survive to adulthood, maternal-zygotic (MZ nanog ) and maternal mutants exhibit developmental arrest at the blastula stage. In the absence of Nanog, YSL formation and epiboly are abnormal, embryonic tissue detaches from the yolk, and the expression of dozens of YSL and embryonic genes is reduced. Epiboly defects can be rescued by generating chimeric embryos of MZ nanog embryonic tissue with wild-type vegetal tissue that includes the YSL and yolk cell. Notably, cells lacking Nanog readily respond to Nodal signals and when transplanted into wild-type hosts proliferate and contribute to embryonic tissues and adult organs from all germ layers. These results indicate that zebrafish Nanog is necessary for proper YSL development but is not directly required for embryonic cell differentiation. © 2018. Published by The Company of Biologists Ltd.

The Most Prevalent Freeman-Sheldon Syndrome Mutations in the Embryonic Myosin Motor Share Functional Defects.

PubMed

Walklate, Jonathan; Vera, Carlos; Bloemink, Marieke J; Geeves, Michael A; Leinwand, Leslie

2016-05-06

The embryonic myosin isoform is expressed during fetal development and rapidly down-regulated after birth. Freeman-Sheldon syndrome (FSS) is a disease associated with missense mutations in the motor domain of this myosin. It is the most severe form of distal arthrogryposis, leading to overcontraction of the hands, feet, and orofacial muscles and other joints of the body. Availability of human embryonic muscle tissue has been a limiting factor in investigating the properties of this isoform and its mutations. Using a recombinant expression system, we have studied homogeneous samples of human motors for the WT and three of the most common FSS mutants: R672H, R672C, and T178I. Our data suggest that the WT embryonic myosin motor is similar in contractile speed to the slow type I/β cardiac based on the rate constant for ADP release and ADP affinity for actin-myosin. All three FSS mutations show dramatic changes in kinetic properties, most notably the slowing of the apparent ATP hydrolysis step (reduced 5-9-fold), leading to a longer lived detached state and a slowed Vmax of the ATPase (2-35-fold), indicating a slower cycling time. These mutations therefore seriously disrupt myosin function. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Conserved developmental alternative splicing of muscleblind-like (MBNL) transcripts regulates MBNL localization and activity.

PubMed

Terenzi, Fulvia; Ladd, Andrea N

2010-01-01

Muscleblind-like (MBNL) proteins have been shown to regulate pre-mRNA alternative splicing, and MBNL1 has been implicated in regulating fetal-to-adult transitions in alternative splicing in the heart. MBNL1 is highly conserved, exhibiting more than 95% identity at the amino acid level between birds and mammals. To investigate MBNL1 expression during embryonic heart development, we examined MBNL1 transcript and protein expression in the embryonic chicken heart from the formation of the primitive heart tube through cardiac morphogenesis (embryonic days 1.5 through 8). MBNL1 transcript levels remained steady throughout these stages, whereas MBNL1 protein levels increased and exhibited a shift in isoforms. MBNL1 has several alternatively spliced exons. Using RT-PCR, we determined that the inclusion of one of these, exon 5, decreases dramatically during cardiac morphogenesis. This developmental transition is conserved in mice. Functional analyses of MBNL1 isoforms containing or lacking exon 5-encoded sequences revealed that exon 5 is important for the regulation of the subcellular localization, RNA binding affinity, and alternative splicing activity of MBNL1 proteins. A second MBNL protein, MBNL2, is also expressed in the embryonic heart. We found that MBNL2 exon 5, which is paralogous to MBNL1 exon 5, is similarly regulated during embryonic heart development. Analysis of MBNL1 and MBNL2 transcripts in several embryonic tissues in chicken and mouse indicate that exon 5 alternative splicing is highly conserved and tissue-specific. Thus, we propose that conserved developmental stage- and tissue-specific alternative splicing of MBNL transcripts is an important mechanism by which MBNL activity is regulated during embryonic development.

Impaired Embryonic Development in Mice Overexpressing the RNA-Binding Protein TIAR

PubMed Central

Kharraz, Yacine; Salmand, Pierre-Adrien; Camus, Anne; Auriol, Jacques; Gueydan, Cyril; Kruys, Véronique; Morello, Dominique

2010-01-01

Background TIA-1-related (TIAR) protein is a shuttling RNA-binding protein involved in several steps of RNA metabolism. While in the nucleus TIAR participates to alternative splicing events, in the cytoplasm TIAR acts as a translational repressor on specific transcripts such as those containing AU-Rich Elements (AREs). Due to its ability to assemble abortive pre-initiation complexes coalescing into cytoplasmic granules called stress granules, TIAR is also involved in the general translational arrest observed in cells exposed to environmental stress. However, the in vivo role of this protein has not been studied so far mainly due to severe embryonic lethality upon tiar invalidation. Methodology/Principal Findings To examine potential TIAR tissue-specificity in various cellular contexts, either embryonic or adult, we constructed a TIAR transgenic allele (loxPGFPloxPTIAR) allowing the conditional expression of TIAR protein upon Cre recombinase activity. Here, we report the role of TIAR during mouse embryogenesis. We observed that early TIAR overexpression led to low transgene transmission associated with embryonic lethality starting at early post-implantation stages. Interestingly, while pre-implantation steps evolved correctly in utero, in vitro cultured embryos were very sensitive to culture medium. Control and transgenic embryos developed equally well in the G2 medium, whereas culture in M16 medium led to the phosphorylation of eIF2α that accumulated in cytoplasmic granules precluding transgenic blastocyst hatching. Our results thus reveal a differential TIAR-mediated embryonic response following artificial or natural growth environment. Conclusions/Significance This study reports the importance of the tightly balanced expression of the RNA-binding protein TIAR for normal embryonic development, thereby emphasizing the role of post-transcriptional regulations in early embryonic programming. PMID:20596534

Effect of recombinant-LH and hCG in the absence of FSH on in vitro maturation (IVM) fertilization and early embryonic development of mouse germinal vesicle (GV)-stage oocytes.

PubMed

Dinopoulou, Vasiliki; Drakakis, Peter; Kefala, Stella; Kiapekou, Erasmia; Bletsa, Ritsa; Anagnostou, Elli; Kallianidis, Konstantinos; Loutradis, Dimitrios

2016-06-01

During in vitro maturation (IVM), intrinsic and extrinsic factors must co-operate properly in order to ensure cytoplasmic and nuclear maturation. We examined the possible effect of LH/hCG in the process of oocyte maturation in mice with the addition of recombinant LH (r-LH) and hCG in our IVM cultures of mouse germinal vesicle (GV)-stage oocytes. Moreover, the effects of these hormones on fertilization, early embryonic development and the expression of LH/hCG receptor were examined. Nuclear maturation of GV-stage oocytes was evaluated after culture in the presence of r-LH or hCG. Fertilization rates and embryonic development were assessed after 24h. Total RNA was isolated from oocytes of different stages of maturation and from zygotes and embryos of different stages of development in order to examine the expression of LH/hCG receptor, using RT-PCR. The in vitro nuclear maturation rate of GV-stage oocytes that received hCG was significantly higher compared to the control group. Early embryonic development was increased in the hCG and LH cultures of GV oocytes when LH was further added. The LH/hCG receptor was expressed in all stages of in vitro matured mouse oocytes and in every stage of early embryonic development. Addition of hCG in IVM cultures of mouse GV oocytes increased maturation rates significantly. LH, however, was more beneficial to early embryonic development than hCG. This suggests a promising new technique in basic science research or in clinical reproductive medicine. Copyright © 2016 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

The energy cost of embryonic development in fishes and amphibians, with emphasis on new data from the Australian lungfish, Neoceratodus forsteri.

PubMed

Mueller, Casey A; Joss, Jean M P; Seymour, Roger S

2011-01-01

The rate of oxygen consumption throughout embryonic development is used to indirectly determine the 'cost' of development, which includes both differentiation and growth. This cost is affected by temperature and the duration of incubation in anamniote fish and amphibian embryos. The influences of temperature on embryonic development rate, respiration rate and energetics were investigated in the Australian lungfish, Neoceratodus forsteri, and compared with published data. Developmental stage and oxygen consumption rate were measured until hatching, upon which wet and dry gut-free masses were determined. A measure of the cost of development, the total oxygen required to produce 1 mg of embryonic dry tissue, increased as temperature decreased. The relationship between the oxygen cost of development (C, ml mg(-1)) and dry hatchling mass (M, mg) in fishes and amphibians is described by C = 0.30 M(0.22 0.13 (95% CI)), r (2) = 0.52. The scaling exponent indicates that the cost of embryonic development increases disproportionally with increasing hatchling mass. At 15 and 20°C, N. forsteri cost of development is significantly lower than the regression mean for all species, and at 25°C is lower than the allometrically scaled data set. Unexpectedly, incubation of N. forsteri is long, despite natural development under relatively warm conditions, and may be related to a large genome size. The low cost of development may be associated with construction of a rather sluggish fish with a low capacity for aerobic metabolism. The metabolic rate is lower in N. forsteri hatchlings than in any other fishes or amphibians at the same temperature, which matches the extremely low aerobic metabolic scope of the juveniles.

Fecundity of Quail in Spacelab Microgravity

NASA Technical Reports Server (NTRS)

Wentworth, B. C.; Wentworth, A. L.

1996-01-01

Flight experiments in which fertilized Japanese quail eggs were allowed to develop to various ages in space, and the results of the following laboratory tests are described. Laboratory-based experiments concerned with the embryonic development of Japanese quail in gravity using simulated vibrations and G-force are reported. Effect of turning and ambient temperature at various days of incubation on the development of Japanese quail, and method to feed and water adult and newly hatched Japanese quail in microgravity using a gelatin-based diet as a solid water supply, are also described.

Injurious Effects of Emodin on Maturation of Mouse Oocytes, Fertilization and Fetal Development via Apoptosis

PubMed Central

Chang, Mei-Hui; Chang, Shao-Chung; Chan, Wen-Hsiung

2012-01-01

Emodin (1,3,8-trihydroxy-6-methylanthraquinone), a major constituent of rhubarb, has a wide range of therapeutic applications. Previous studies have established that emodin induces apoptosis in the inner cell mass and trophectoderm of mouse blastocysts and leads to decreased embryonic development and viability, indicating a role as an injury risk factor for normal embryonic development. However, the mechanisms underlying its hazardous effects have yet to be characterized. In the current study, we further investigated the effects of emodin on oocyte maturation and subsequent pre- and post-implantation development, both in vitro and in vivo. Notably, emodin induced a significant reduction in the rates of oocyte maturation, fertilization, and in vitro embryonic development. Treatment of oocytes with emodin during in vitro maturation (IVM) led to increased resorption of postimplantation embryos and decreased fetal weight. Experiments using an in vivo mouse model disclosed that consumption of drinking water containing 20–40 μM emodin led to decreased oocyte maturation and in vitro fertilization, as well as early embryonic developmental injury. Notably, pretreatment with a caspase-3-specific inhibitor effectively prevented emodin-triggered injury effects, suggesting that impairment of embryo development occurs via a caspase-dependent apoptotic process. PMID:23203041

Injurious effects of emodin on maturation of mouse oocytes, fertilization and fetal development via apoptosis.

PubMed

Chang, Mei-Hui; Chang, Shao-Chung; Chan, Wen-Hsiung

2012-10-29

Emodin (1,3,8-trihydroxy-6-methylanthraquinone), a major constituent of rhubarb, has a wide range of therapeutic applications. Previous studies have established that emodin induces apoptosis in the inner cell mass and trophectoderm of mouse blastocysts and leads to decreased embryonic development and viability, indicating a role as an injury risk factor for normal embryonic development. However, the mechanisms underlying its hazardous effects have yet to be characterized. In the current study, we further investigated the effects of emodin on oocyte maturation and subsequent pre- and post-implantation development, both in vitro and in vivo. Notably, emodin induced a significant reduction in the rates of oocyte maturation, fertilization, and in vitro embryonic development. Treatment of oocytes with emodin during in vitro maturation (IVM) led to increased resorption of postimplantation embryos and decreased fetal weight. Experiments using an in vivo mouse model disclosed that consumption of drinking water containing 20-40 μM emodin led to decreased oocyte maturation and in vitro fertilization, as well as early embryonic developmental injury. Notably, pretreatment with a caspase-3-specific inhibitor effectively prevented emodin-triggered injury effects, suggesting that impairment of embryo development occurs via a caspase-dependent apoptotic process.

The case for DNA methylation based molecular profiling to improve diagnostic accuracy for central nervous system embryonal tumors (not otherwise specified) in adults.

PubMed

Halliday, Gail C; Junckerstorff, Reimar C; Bentel, Jacqueline M; Miles, Andrew; Jones, David T W; Hovestadt, Volker; Capper, David; Endersby, Raelene; Cole, Catherine H; van Hagen, Tom; Gottardo, Nicholas G

2018-01-01

Central nervous system primitive neuro-ectodermal tumors (CNS-PNETs), have recently been re-classified in the most recent 2016 WHO Classification into a standby catch all category, "CNS Embryonal Tumor, not otherwise specified" (CNS embryonal tumor, NOS) based on epigenetic, biologic and histopathologic criteria. CNS embryonal tumors (NOS) are a rare, histologically and molecularly heterogeneous group of tumors that predominantly affect children, and occasionally adults. Diagnosis of this entity continues to be challenging and the ramifications of misdiagnosis of this aggressive class of brain tumors are significant. We report the case of a 45-year-old woman who was diagnosed with a central nervous system embryonal tumor (NOS) based on immunohistochemical analysis of the patient's tumor at diagnosis. However, later genome-wide methylation profiling of the diagnostic tumor undertaken to guide treatment, revealed characteristics most consistent with IDH-mutant astrocytoma. DNA sequencing and immunohistochemistry confirmed the presence of IDH1 and ATRX mutations resulting in a revised diagnosis of high-grade small cell astrocytoma, and the implementation of a less aggressive treatment regime tailored more appropriately to the patient's tumor type. This case highlights the inadequacy of histology alone for the diagnosis of brain tumours and the utility of methylation profiling and integrated genomic analysis for the diagnostic verification of adults with suspected CNS embryonal tumor (NOS), and is consistent with the increasing realization in the field that a combined diagnostic approach based on clinical, histopathological and molecular data is required to more accurately distinguish brain tumor subtypes and inform more effective therapy. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Does gravity influence the early stages of the development of the nervous system in an amphibian?

PubMed

Duprat, A M; Husson, D; Gualandris-Parisot, L

1998-11-01

As a result of previous studies using hypergravity (centrifuge) or virtual microgravity (clinostat), it was proposed that gravity was involved in embryonic development, i.e., in the establishment of the embryonic polarities and the body plan pattern which subsequently direct morphogenesis and organogenesis of the central nervous system and of sensory organs. Recent experiments were performed in space using sounding rockets and orbiting space-modules to ascertain whether gravity is indeed required for embryogenesis in Invertebrates and Vertebrates. Eggs fertilised in vivo or in vitro in microgravity showed some abnormalities during embryonic development but were able to regulate and produce nearly normal larvae. Copyright 1998 Elsevier Science B.V.

Kisspeptin regulates ovarian steroidogenesis during delayed embryonic development in the fruit bat, Cynopterus sphinx.

PubMed

Anuradha; Krishna, Amitabh

2017-11-01

Cynopterus sphinx, a fruit bat, undergoes delayed embryonic development during the winter months, a period that corresponds to low levels of progesterone and estradiol synthesis by the ovary. Kisspeptins (KPs) are a group of neuropeptide hormones that act via G-protein coupled receptor 54 (GPR54) to stimulate hypothalamic secretion of Gonadotropin-releasing hormone, thereby regulating ovarian steroidogenesis, folliculogenesis, and ovulation. GPR54 is also expressed in the ovary, suggesting a direct role for KPs in ovarian steroidogenesis. The aim of present study was to determine if a low serum level of KP is responsible for reduced progesterone and estradiol levels during the period of delayed embryonic development in C. sphinx. Indeed, low serum KP abundance corresponded to reduced expression of GPR54 in ovarian luteal cells during the period of delayed development compared to normal development. In vitro and in vivo treatment with KP increased GPR54 abundance, via Extracellular signal regulated kinase and its downstream mediators, leading to increased progesterone synthesis in the ovary during delayed embryonic development. KP treatment also increased cholesterol uptake and elevated expression of Luteinizing hormone receptor and Steroid acute regulatory protein in the ovary, suggesting that elevation in circulating KP during delayed embryonic development may reactivate luteal activity. KPs may also enhance cell survival (BCL-2, reduced Caspase 3 activity) and angiogenesis (Vascular endothelium growth factor) during this period. The findings of this study thus demonstrate a regulatory role for KPs in the maintenance of luteal steroidogenesis during pregnancy in C. sphinx. © 2017 Wiley Periodicals, Inc.

Rotational imaging optical coherence tomography for full-body mouse embryonic imaging

PubMed Central

Wu, Chen; Sudheendran, Narendran; Singh, Manmohan; Larina, Irina V.; Dickinson, Mary E.; Larin, Kirill V.

2016-01-01

Abstract. Optical coherence tomography (OCT) has been widely used to study mammalian embryonic development with the advantages of high spatial and temporal resolutions and without the need for any contrast enhancement probes. However, the limited imaging depth of traditional OCT might prohibit visualization of the full embryonic body. To overcome this limitation, we have developed a new methodology to enhance the imaging range of OCT in embryonic day (E) 9.5 and 10.5 mouse embryos using rotational imaging. Rotational imaging OCT (RI-OCT) enables full-body imaging of mouse embryos by performing multiangle imaging. A series of postprocessing procedures was performed on each cross-section image, resulting in the final composited image. The results demonstrate that RI-OCT is able to improve the visualization of internal mouse embryo structures as compared to conventional OCT. PMID:26848543

Impaired cardiac energy metabolism in embryos lacking adrenergic stimulation.

PubMed

Baker, Candice N; Gidus, Sarah A; Price, George F; Peoples, Jessica N R; Ebert, Steven N

2015-03-01

As development proceeds from the embryonic to fetal stages, cardiac energy demands increase substantially, and oxidative phosphorylation of ADP to ATP in mitochondria becomes vital. Relatively little, however, is known about the signaling mechanisms regulating the transition from anaerobic to aerobic metabolism that occurs during the embryonic period. The main objective of this study was to test the hypothesis that adrenergic hormones provide critical stimulation of energy metabolism during embryonic/fetal development. We examined ATP and ADP concentrations in mouse embryos lacking adrenergic hormones due to targeted disruption of the essential dopamine β-hydroxylase (Dbh) gene. Embryonic ATP concentrations decreased dramatically, whereas ADP concentrations rose such that the ATP/ADP ratio in the adrenergic-deficient group was nearly 50-fold less than that found in littermate controls by embryonic day 11.5. We also found that cardiac extracellular acidification and oxygen consumption rates were significantly decreased, and mitochondria were significantly larger and more branched in adrenergic-deficient hearts. Notably, however, the mitochondria were intact with well-formed cristae, and there was no significant difference observed in mitochondrial membrane potential. Maternal administration of the adrenergic receptor agonists isoproterenol or l-phenylephrine significantly ameliorated the decreases in ATP observed in Dbh-/- embryos, suggesting that α- and β-adrenergic receptors were effective modulators of ATP concentrations in mouse embryos in vivo. These data demonstrate that adrenergic hormones stimulate cardiac energy metabolism during a critical period of embryonic development. Copyright © 2015 the American Physiological Society.

UTX regulates mesoderm differentiation of embryonic stem cells independent of H3K27 demethylase activity.

PubMed

Wang, Chaochen; Lee, Ji-Eun; Cho, Young-Wook; Xiao, Ying; Jin, Qihuang; Liu, Chengyu; Ge, Kai

2012-09-18

To investigate the role of histone H3K27 demethylase UTX in embryonic stem (ES) cell differentiation, we have generated UTX knockout (KO) and enzyme-dead knock-in male ES cells. Deletion of the X-chromosome-encoded UTX gene in male ES cells markedly decreases expression of the paralogous UTY gene encoded by Y chromosome, but has no effect on global H3K27me3 level, Hox gene expression, or ES cell self-renewal. However, UTX KO cells show severe defects in mesoderm differentiation and induction of Brachyury, a transcription factor essential for mesoderm development. Surprisingly, UTX regulates mesoderm differentiation and Brachyury expression independent of its enzymatic activity. UTY, which lacks detectable demethylase activity, compensates for the loss of UTX in regulating Brachyury expression. UTX and UTY bind directly to Brachyury promoter and are required for Wnt/β-catenin signaling-induced Brachyury expression in ES cells. Interestingly, male UTX KO embryos express normal levels of UTY and survive until birth. In contrast, female UTX KO mice, which lack the UTY gene, show embryonic lethality before embryonic day 11.5. Female UTX KO embryos show severe defects in both Brachyury expression and embryonic development of mesoderm-derived posterior notochord, cardiac, and hematopoietic tissues. These results indicate that UTX controls mesoderm differentiation and Brachyury expression independent of H3K27 demethylase activity, and suggest that UTX and UTY are functionally redundant in ES cell differentiation and early embryonic development.

EMG1 is essential for mouse pre-implantation embryo development.

PubMed

Wu, Xiaoli; Sandhu, Sumit; Patel, Nehal; Triggs-Raine, Barbara; Ding, Hao

2010-09-21

Essential for mitotic growth 1 (EMG1) is a highly conserved nucleolar protein identified in yeast to have a critical function in ribosome biogenesis. A mutation in the human EMG1 homolog causes Bowen-Conradi syndrome (BCS), a developmental disorder characterized by severe growth failure and psychomotor retardation leading to death in early childhood. To begin to understand the role of EMG1 in mammalian development, and how its deficiency could lead to Bowen-Conradi syndrome, we have used mouse as a model. The expression of Emg1 during mouse development was examined and mice carrying a null mutation for Emg1 were generated and characterized. Our studies indicated that Emg1 is broadly expressed during early mouse embryonic development. However, in late embryonic stages and during postnatal development, Emg1 exhibited specific expression patterns. To assess a developmental role for EMG1 in vivo, we exploited a mouse gene-targeting approach. Loss of EMG1 function in mice arrested embryonic development prior to the blastocyst stage. The arrested Emg1-/- embryos exhibited defects in early cell lineage-specification as well as in nucleologenesis. Further, loss of p53, which has been shown to rescue some phenotypes resulting from defects in ribosome biogenesis, failed to rescue the Emg1-/- pre-implantation lethality. Our data demonstrate that Emg1 is highly expressed during mouse embryonic development, and essential for mouse pre-implantation development. The absolute requirement for EMG1 in early embryonic development is consistent with its essential role in yeast. Further, our findings also lend support to the previous study that showed Bowen-Conradi syndrome results from a partial EMG1 deficiency. A complete deficiency would not be expected to be compatible with a live birth.

ELF (Extremely Low Frequency) Communications System Ecological Monitoring Program: Summary of 1987 Progress

DTIC Science & Technology

1989-04-01

Development . Prenatal developmental stages are especially sensitive to environmental perturbations. At present, there is conflicting evidence of direct EM...effects on embryonic or fetal development . In addition, possible effects of the ELF system on parental behavior could also have an indirect effect on... development . The purpose of this element is to determine the incidence of abnormalities in embryonic development in tree swallows at treatment and control

In vitro organogenesis of gut-like structures from mouse embryonic stem cells.

PubMed

Kuwahara, M; Ogaeri, T; Matsuura, R; Kogo, H; Fujimoto, T; Torihashi, S

2004-04-01

Embryonic stem (ES) cells have pluripotency and give rise to many cell types and tissues, including representatives of all three germ layers in the embryo. We have reported previously that mouse ES cells formed contracting gut-like organs from embryoid bodies (EBs). These gut-like structures contracted spontaneously, and had large lumens surrounded by three layers, i.e. epithelium, lamina propria and muscularis. Ganglia were scattered along the periphery, and interstitial cells of Cajal (ICC) were distributed among the smooth muscle cells. In the present study, to determine whether they can be a model of gut organogenesis, we investigated the formation process of the gut-like structures in comparison with embryonic gut development. As a result, we found that the fundamental process of formation in vitro was similar to embryonic gut development in vivo. The result indicates that the gut-like structure is a useful tool not only for developmental study to determine the factors that induce gut organogenesis, but also for studies of enteric neurone and ICC development.

Isolation and expression of homeobox genes from the embryonic chicken eye.

PubMed

Dhawan, R R; Schoen, T J; Beebe, D C

1997-06-11

To identify homeobox-containing genes that may play a role in the differentiation of ocular tissues. Total RNA was isolated from microdissected chicken embryo eye tissues at 3.5 days of development (embryonic day 3.5; E3.5). An "anchor-oligo-dT primer" was used for the synthesis of cDNA. Degenerate oligonucleotides designed from highly-conserved sequences in the third helix of the homeobox and the "anchor-primer" were used to amplify cDNAs by polymerase chain reaction (PCR). PCR products were cloned and sequenced. The spatial and temporal expression of selected transcripts was mapped by whole-mount in situ hybridization and northern blot analysis. After sequencing eighteen clones we identified a member of the distal-less family (dlx-3) in cDNA from presumptive neural retina and three chicken homologs of the Xenopus "anterior neural fold" (Xanf-1) in cDNA from anterior eye tissue. Dlx transcripts were mapped by in situ hybridization. Expression began at Hamburger and Hamilton stage 14 (E2.5) and was widely distributed in embryonic mesenchyme on E3 and E4. Expression increased in the retina during early development and persisted until after hatching. The one anf clone selected for further study was not detected by in situ or northern blot analysis. It is feasible to isolate homeobox cDNAs directly from microdissected embryonic tissues. Chicken dlx-3 mRNA has a wider distribution in the embryo than expected, based on the expression of the mouse homolog. Dlx-3 may play a role in establishing or maintaining the differentiation of the retina.

Revising the embryonic origin of thyroid C cells in mice and humans

PubMed Central

Johansson, Ellen; Andersson, Louise; Örnros, Jessica; Carlsson, Therese; Ingeson-Carlsson, Camilla; Liang, Shawn; Dahlberg, Jakob; Jansson, Svante; Parrillo, Luca; Zoppoli, Pietro; Barila, Guillermo O.; Altschuler, Daniel L.; Padula, Daniela; Lickert, Heiko; Fagman, Henrik; Nilsson, Mikael

2015-01-01

Current understanding infers a neural crest origin of thyroid C cells, the major source of calcitonin in mammals and ancestors to neuroendocrine thyroid tumors. The concept is primarily based on investigations in quail–chick chimeras involving fate mapping of neural crest cells to the ultimobranchial glands that regulate Ca2+ homeostasis in birds, reptiles, amphibians and fishes, but whether mammalian C cell development involves a homologous ontogenetic trajectory has not been experimentally verified. With lineage tracing, we now provide direct evidence that Sox17+ anterior endoderm is the only source of differentiated C cells and their progenitors in mice. Like many gut endoderm derivatives, embryonic C cells were found to coexpress pioneer factors forkhead box (Fox) a1 and Foxa2 before neuroendocrine differentiation takes place. In the ultimobranchial body epithelium emerging from pharyngeal pouch endoderm in early organogenesis, differential Foxa1/Foxa2 expression distinguished two spatially separated pools of C cell precursors with different growth properties. A similar expression pattern was recapitulated in medullary thyroid carcinoma cells in vivo, consistent with a growth-promoting role of Foxa1. In contrast to embryonic precursor cells, C cell-derived tumor cells invading the stromal compartment downregulated Foxa2, foregoing epithelial-to-mesenchymal transition designated by loss of E-cadherin; both Foxa2 and E-cadherin were re-expressed at metastatic sites. These findings revise mammalian C cell ontogeny, expand the neuroendocrine repertoire of endoderm and redefine the boundaries of neural crest diversification. The data further underpin distinct functions of Foxa1 and Foxa2 in both embryonic and tumor development. PMID:26395490

G-quadruplexes as novel cis-elements controlling transcription during embryonic development.

PubMed

David, Aldana P; Margarit, Ezequiel; Domizi, Pablo; Banchio, Claudia; Armas, Pablo; Calcaterra, Nora B

2016-05-19

G-quadruplexes are dynamic structures folded in G-rich single-stranded DNA regions. These structures have been recognized as a potential nucleic acid based mechanism for regulating multiple cellular processes such as replication, transcription and genomic maintenance. So far, their transcriptional role in vivo during vertebrate embryonic development has not yet been addressed. Here, we performed an in silico search to find conserved putative G-quadruplex sequences (PQSs) within proximal promoter regions of human, mouse and zebrafish developmental genes. Among the PQSs able to fold in vitro as G-quadruplex, those present in nog3, col2a1 and fzd5 promoters were selected for further studies. In cellulo studies revealed that the selected G-quadruplexes affected the transcription of luciferase controlled by the SV40 nonrelated promoter. G-quadruplex disruption in vivo by microinjection in zebrafish embryos of either small ligands or DNA oligonucleotides complementary to the selected PQSs resulted in lower transcription of the targeted genes. Moreover, zebrafish embryos and larvae phenotypes caused by the presence of complementary oligonucleotides fully resembled those ones reported for nog3, col2a1 and fzd5 morphants. To our knowledge, this is the first work revealing in vivo the role of conserved G-quadruplexes in the embryonic development, one of the most regulated processes of the vertebrates biology. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

G-quadruplexes as novel cis-elements controlling transcription during embryonic development

PubMed Central

David, Aldana P.; Margarit, Ezequiel; Domizi, Pablo; Banchio, Claudia; Armas, Pablo; Calcaterra, Nora B.

2016-01-01

G-quadruplexes are dynamic structures folded in G-rich single-stranded DNA regions. These structures have been recognized as a potential nucleic acid based mechanism for regulating multiple cellular processes such as replication, transcription and genomic maintenance. So far, their transcriptional role in vivo during vertebrate embryonic development has not yet been addressed. Here, we performed an in silico search to find conserved putative G-quadruplex sequences (PQSs) within proximal promoter regions of human, mouse and zebrafish developmental genes. Among the PQSs able to fold in vitro as G-quadruplex, those present in nog3, col2a1 and fzd5 promoters were selected for further studies. In cellulo studies revealed that the selected G-quadruplexes affected the transcription of luciferase controlled by the SV40 nonrelated promoter. G-quadruplex disruption in vivo by microinjection in zebrafish embryos of either small ligands or DNA oligonucleotides complementary to the selected PQSs resulted in lower transcription of the targeted genes. Moreover, zebrafish embryos and larvae phenotypes caused by the presence of complementary oligonucleotides fully resembled those ones reported for nog3, col2a1 and fzd5 morphants. To our knowledge, this is the first work revealing in vivo the role of conserved G-quadruplexes in the embryonic development, one of the most regulated processes of the vertebrates biology. PMID:26773060

Effects of catechins and low temperature on embryonic development and hatching in Heterodera glycines and Meloidogyne incognita

USDA-ARS?s Scientific Manuscript database

Mimics of two natural influences, a chemical similar to one present in cyst nematodes and low temperature exposure of nematode eggs, were evaluated for their effects on quantitative and qualitative features of embryonic development and hatching. The polyphenol epigallocatechin gallate (EGCG), an ana...

Intrauterine air impairs embryonic postimplantation development in mice.

PubMed

Liu, Ruonan; Li, Yimeng; Miao, Yanping; Wei, Yanhui; Guan, Mo; Zhou, Rongyan; Li, Xiangyun

2017-12-01

Although most embryologists load air bubbles into the catheter along with embryos during embryo transfer, the effects of these air bubbles on embryo transfer success rate are not clear. Air bubbles were nonsurgically injected into unilateral uterine horns of mice to demonstrate the negative effects of intrauterine air bubbles on embryonic development. Our data showed that when air bubbles are nonsurgically injected into unilateral uterine horns of pregnant 4days mice the litter size is significantly decreased. Four days after the introduction of air, abnormal decidua and dead conceptuses were detected in the uterine horns receiving the air bubbles. In addition, intrauterine air also significantly impaired murine embryo transfer success rates, and induced an increase in endometrial capillary permeability and decidualization in mice on day 4 of pseudopregnancy. These results strongly indicated that the air bubbles loaded into embryo transfer catheters to bracket the embryo-containing medium may have negative effect on embryonic implantation and development. Intrauterine air impaired murine embryonic postimplantation development, and this provided some clues for improving embryo transfer techniques in human. Copyright © 2017 Elsevier B.V. All rights reserved.

Tension (re)builds: Biophysical mechanisms of embryonic wound repair.

PubMed

Zulueta-Coarasa, Teresa; Fernandez-Gonzalez, Rodrigo

2017-04-01

Embryonic tissues display an outstanding ability to rapidly repair wounds. Epithelia, in particular, serve as protective layers that line internal organs and form the skin. Thus, maintenance of epithelial integrity is of utmost importance for animal survival, particularly at embryonic stages, when an immune system has not yet fully developed. Rapid embryonic repair of epithelial tissues is conserved across species, and involves the collective migration of the cells around the wound. The migratory cell behaviours associated with wound repair require the generation and transmission of mechanical forces, not only for the cells to move, but also to coordinate their movements. Here, we review the forces involved in embryonic wound repair. We discuss how different force-generating structures are assembled at the molecular level, and the mechanisms that maintain the balance between force-generating structures as wounds close. Finally, we describe the mechanisms that cells use to coordinate the generation of mechanical forces around the wound. Collective cell movements and their misregulation have been associated with defective tissue repair, developmental abnormalities and cancer metastasis. Thus, we propose that understanding the role of mechanical forces during embryonic wound closure will be crucial to develop therapeutic interventions that promote or prevent collective cell movements under pathological conditions. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Maternal dietary zinc supplementation enhances the epigenetic-activated antioxidant ability of chick embryos from maternal normal and high temperatures.

PubMed

Zhu, Yongwen; Liao, Xiudong; Lu, Lin; Li, Wenxiang; Zhang, Liyang; Ji, Cheng; Lin, Xi; Liu, Hsiao-Ching; Odle, Jack; Luo, Xugang

2017-03-21

The role of maternal dietary zinc supplementation in protecting the embryos from maternal hyperthermia-induced negative effects via epigenetic mechanisms was examined using an avian model (Gallus gallus). Broiler breeder hens were exposed to two maternal temperatures (21°C and 32°C) × three maternal dietary zinc treatments (zinc-unsupplemented control diet, the control diet + 110 mg zinc/kg inorganic or organic zinc) for 8 weeks. Maternal hyperthermia increased the embryonic mortality and induced oxidative damage evidenced by the elevated mRNA expressions of heat shock protein genes. Maternal dietary zinc deficiency damaged the embryonic development associated with the global DNA hypomethylation and histone 3 lysine 9 hyperacetylation in the embryonic liver. Supplementation of zinc in maternal diets effectively eliminated the embryonic mortality induced by maternal hyperthermia and enhanced antioxidant ability with the increased mRNA and protein expressions of metallothionein IV in the embryonic liver. The increased metallothionein IV mRNA expression was due to the reduced DNA methylation and increased histone 3 lysine 9 acetylation of the metallothionein IV promoter regardless of zinc source. These data demonstrate that maternal dietary zinc addition as an epigenetic modifier could protect the offspring embryonic development against maternal heat stress via enhancing the epigenetic-activated antioxidant ability.

Early zebrafish development: It’s in the maternal genes

PubMed Central

Abrams, Elliott W.; Mullins, Mary C.

2009-01-01

Summary The earliest stages of embryonic development in all animals examined rely on maternal gene products that are generated during oogenesis and supplied to the egg. The period of maternal control of embryonic development varies among animals according to the onset of zygotic transcription and the persistence of maternal gene products. This maternal regulation has been little studied in vertebrates, due to the difficulty in manipulating maternal gene function and lack of basic molecular information. However, recent maternal-effect screens in the zebrafish have generated more than 40 unique mutants that are providing new molecular entry points to the maternal control of early vertebrate development. Here we discuss recent studies of 12 zebrafish mutant genes that illuminate the maternal molecular controls on embryonic development, including advances in the regulation of animal-vegetal polarity, egg activation, cleavage development, body plan formation, tissue morphogenesis, microRNA function and germ cell development. PMID:19608405

Adult mortality probability and nest predation rates explain parental effort in warming eggs with consequences for embryonic development time

USGS Publications Warehouse

Martin, Thomas E.; Oteyza, Juan C.; Boyce, Andy J.; Lloyd, Penn; Ton, Riccardo

2015-01-01

Parental behavior and effort vary extensively among species. Life-history theory suggests that age-specific mortality could cause this interspecific variation, but past tests have focused on fecundity as the measure of parental effort. Fecundity can cause costs of reproduction that confuse whether mortality is the cause or the consequence of parental effort. We focus on a trait, parental allocation of time and effort in warming embryos, that varies widely among species of diverse taxa and is not tied to fecundity. We conducted studies on songbirds of four continents and show that time spent warming eggs varies widely among species and latitudes and is not correlated with clutch size. Adult and offspring (nest) mortality explained most of the interspecific variation in time and effort that parents spend warming eggs, measured by average egg temperatures. Parental effort in warming eggs is important because embryonic temperature can influence embryonic development period and hence exposure time to predation risk. We show through correlative evidence and experimental swapping of embryos between species that parentally induced egg temperatures cause interspecific variation in embryonic development period. The strong association of age-specific mortality with parental effort in warming eggs and the subsequent effects on embryonic development time are unique results that can advance understanding of broad geographic patterns of life-history variation.

Correlation of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced apoptotic cell death in the embryonic vasculature with embryotoxicity

USGS Publications Warehouse

Cantrell, Susannah M.; Joy-Schlezinger, Jennifer; Stegeman, John J.; Tillitt, Donald E.; Hannington, Mark D.

1998-01-01

Vertebrate embryos are particularly sensitive to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Identification of tissues that are susceptible to the adverse effects of TCDD is requisite for understanding the embryo toxic effects of TCDD. The objective of the present study was to quantitate the temporal appearance of and dose dependence of apoptosis in TCDD-exposed medaka embryos (Oryzias latipes). A fluorescent-based DNA end-labeling assay provided a sensitive method for detection of TCDD-induced apoptosis in tissue sections of medaka embryos. Apoptotic cells were readily apparent in the medial yolk vein at all observed embryonic stages in TCDD-exposed embryos. Slope-comparison analysis indicated that TCDD-induced programmed cell death in the embryonic medial yolk vein was mechanistically linked to embryo mortality. These data are consistent with the hypothesis that vascular damage contributes to the acute embryo toxic effects of TCDD. However, as sublethal concentrations of dioxin-like compounds are more typical of environmental exposures, tissue damage was also assessed in medaka fry that were exposed to low doses of TCDD during embryonic development. Cell death was detected in gill and digestive tissues in visibly healthy medaka fry that had been exposed to low doses of TCDD during embryonic development. Increased expression of cytochrome P450 1A is a major biochemical consequence of TCDD exposure and is often used as a biomarker for exposure to dioxin-like compounds. Therefore, we compared the tissue distribution of TCDD-induced P450 1A expression and TCDD-induced programmed cell death. TCDD-induced programmed cell death co-localized with TCDD-induced P450 1A expression in both embryos and in visibly healthy post-hatch fry. Our results suggest that aberrant programmed cell death may be a suitable marker for exposure of feral organisms to dioxin-like compounds.

Hox genes require homothorax and extradenticle for body wall identity specification but not for appendage identity specification during metamorphosis of Tribolium castaneum.

PubMed

Smith, Frank W; Jockusch, Elizabeth L

2014-11-01

The establishment of segment identity is a key developmental process that allows for divergence along the anteroposterior body axis in arthropods. In Drosophila, the identity of a segment is determined by the complement of Hox genes it expresses. In many contexts, Hox transcription factors require the protein products of extradenticle (exd) and homothorax (hth) as cofactors to perform their identity specification functions. In holometabolous insects, segment identity may be specified twice, during embryogenesis and metamorphosis. To glean insight into the relationship between embryonic and metamorphic segmental identity specification, we have compared these processes in the flour beetle Tribolium castaneum, which develops ventral appendages during embryogenesis that later metamorphose into adult appendages with distinct morphologies. At metamorphosis, comparisons of RNAi phenotypes indicate that Hox genes function jointly with Tc-hth and Tc-exd to specify several region-specific aspects of the adult body wall. On the other hand, Hox genes specify appendage identities along the anteroposterior axis independently of Tc-hth/Tc-exd and Tc-hth/Tc-exd specify proximal vs. distal identity within appendages independently of Hox genes during this stage. During embryogenesis, Tc-hth and Tc-exd play a broad role in the segmentation process and are required for specification of body wall identities in the thorax; however, contrasting with results from other species, we did not obtain homeotic transformations of embryonic appendages in response to Tc-hth or Tc-exd RNAi. In general, the homeotic effects of interference with the function of Hox genes and Tc-hth/Tc-exd during metamorphosis did not match predictions based on embryonic roles of these genes. Comparing metamorphic patterning in T. castaneum to embryonic and post-embryonic development in hemimetabolous insects suggests that holometabolous metamorphosis combines patterning processes of both late embryogenesis and metamorphosis of the hemimetabolous life cycle. Copyright © 2014 Elsevier Inc. All rights reserved.

Bypass of lethality with mosaic mice generated by Cre-loxP-mediated recombination.

PubMed

Betz, U A; Vosshenrich, C A; Rajewsky, K; Müller, W

1996-10-01

The analysis of gene function based on the generation of mutant mice by homologous recombination in embryonic stem cells is limited if gene disruption results in embryonic lethality. Mosaic mice, which contain a certain proportion of mutant cells in all organs, allow lethality to be circumvented and the potential of mutant cells to contribute to different cell lineages to be analyzed. To generate mosaic animals, we used the bacteriophage P1-derived Cre-loxP recombination system, which allows gene alteration by Cre-mediated deletion of loxP-flanked gene segments. We generated nestin-cre transgenic mouse lines, which expressed the Cre recombinase under the control of the rat nestin promoter and its second intron enhancer. In crosses to animals carrying a loxP-flanked target gene, partial deletion of the loxP-flanked allele occurred before day 10.5 post coitum and was detectable in all adult organs examined, including germ-line cells. Using this approach, we generated mosaic mice containing cells deficient in the gamma-chain of the interleukin-2 receptor (IL-2R gamma); in these animals, the IL-2R gamma-deficient cells were underrepresented in the thymus and spleen. Because mice deficient in DNA polymerase beta die perinatally, we studied the effects of DNA polymerase beta deficiency in mosaic animals. We found that some of the mosaic polymerase beta-deficient animals were viable, but were often reduced in size and weight. The fraction of DNA polymerase beta-deficient cells in mosaic embryos decreased during embryonic development, presumably because wild-type cells had a competitive advantage. The nestin-cre transgenic mice can be used to generate mosaic animals in which target genes are mutated by Cre-mediated recombination of loxP-flanked target genes. By using mosaic animals, embryonic lethality can be bypassed and cell lineages for whose development a given target gene is critical can be identified. In the case of DNA polymerase beta, deficient cells are already selected against during embryonic development, demonstrating the general importance of this protein in multiple cell types.

EXTRA-EMBRYONIC-SPECIFIC IMPRINTED EXPRESSION IS RESTRICTED TO DEFINED LINEAGES IN THE POST-IMPLANTATION EMBRYO

PubMed Central

Hudson, Quanah J.; Seidl, Christine I.M.; Kulinski, Tomasz M.; Huang, Ru; Warczok, Katarzyna E.; Bittner, Romana; Bartolomei, Marisa S.; Barlow, Denise P.

2011-01-01

A subset of imprinted genes in the mouse have been reported to show imprinted expression that is restricted to the placenta, a short-lived extra-embryonic organ. Notably these so-called 'placental-specific' imprinted genes are expressed from both parental alleles in embryo and adult tissues. The placenta is an embryonic-derived organ that is closely associated with maternal tissue and as a consequence, maternal contamination can be mistaken for maternal-specific imprinted expression. The complexity of the placenta, which arises from multiple embryonic lineages, poses additional problems in accurately assessing allele-specific repressive epigenetic modifications in genes that also show lineage-specific silencing in this organ. These problems require that extra evidence be obtained to support the imprinted status of genes whose imprinted expression is restricted to the placenta. We show here that the extra-embryonic visceral yolk sac (VYS), a nutritive membrane surrounding the developing embryo, shows a similar 'extra-embryonic-lineage-specific' pattern of imprinted expression. We present an improved enzymatic technique for separating the bilaminar VYS and show that this pattern of imprinted expression is restricted to the endoderm layer. Finally, we show that VYS 'extra-embryonic-lineage-specific' imprinted expression is regulated by DNA methylation in a similar manner as shown for genes showing multi-lineage imprinted expression in extra-embryonic, embryonic and adult tissues. These results show that the VYS is an improved model for studying the epigenetic mechanisms regulating extra-embryonic-lineage-specific imprinted expression. PMID:21354127

Expression and regulation of glucocorticoid-induced leucine zipper in the developing anterior pituitary gland.

PubMed

Ellestad, Laura E; Malkiewicz, Stefanie A; Guthrie, H David; Welch, Glenn R; Porter, Tom E

2009-02-01

The expression profile of glucocorticoid-induced leucine zipper (GILZ) in the anterior pituitary during the second half of embryonic development in the chick is consistent with in vivo regulation by circulating corticosteroids. However, nothing else has been reported about the presence of GILZ in the neuroendocrine system. We sought to characterize expression and regulation of GILZ in the chicken embryonic pituitary gland and determine the effect of GILZ overexpression on anterior pituitary hormone levels. Pituitary GILZ mRNA levels increased during embryogenesis to a maximum on the day of hatch, and decreased through the first week after hatch. GILZ expression was rapidly upregulated by corticosterone in embryonic pituitary cells. To determine whether GILZ regulates hormone gene expression in the developing anterior pituitary, we overexpressed GILZ in embryonic pituitary cells and measured mRNA for the major pituitary hormones. Exogenous GILZ increased prolactin mRNA above basal levels, but not as high as that in corticosterone-treated cells, indicating that GILZ may play a small role in lactotroph differentiation. The largest effect we observed was a twofold increase in FSH beta subunit in cells transfected with GILZ but not treated with corticosterone, suggesting that GILZ may positively regulate gonadotroph development in a manner not involving glucocorticoids. In conclusion, this is the first report to characterize avian GILZ and examine its regulation in the developing neuroendocrine system. We have shown that GILZ is upregulated by glucocorticoids in the embryonic pituitary gland and may regulate expression of several pituitary hormones.

4D Subject-Specific Inverse Modeling of the Chick Embryonic Heart Outflow Tract Hemodynamics

PubMed Central

Goenezen, Sevan; Chivukula, Venkat Keshav; Midgett, Madeline; Phan, Ly; Rugonyi, Sandra

2015-01-01

Blood flow plays a critical role in regulating embryonic cardiac growth and development, with altered flow leading to congenital heart disease. Progress in the field, however, is hindered by a lack of quantification of hemodynamic conditions in the developing heart. In this study, we present a methodology to quantify blood flow dynamics in the embryonic heart using subject-specific computational fluid dynamics (CFD) models. While the methodology is general, we focused on a model of the chick embryonic heart outflow tract (OFT), which distally connects the heart to the arterial system, and is the region of origin of many congenital cardiac defects. Using structural and Doppler velocity data collected from optical coherence tomography (OCT), we generated 4D (3D + time) embryo-specific CFD models of the heart OFT. To replicate the blood flow dynamics over time during the cardiac cycle, we developed an iterative inverse-method optimization algorithm, which determines the CFD model boundary conditions such that differences between computed velocities and measured velocities at one point within the OFT lumen are minimized. Results from our developed CFD model agree with previously measured hemodynamics in the OFT. Further, computed velocities and measured velocities differ by less than 15% at locations that were not used in the optimization, validating the model. The presented methodology can be used in quantifications of embryonic cardiac hemodynamics under normal and altered blood flow conditions, enabling an in depth quantitative study of how blood flow influences cardiac development. PMID:26361767

Gas exchange in avian embryos and hatchlings.

PubMed

Mortola, Jacopo P

2009-08-01

The avian egg has been proven to be an excellent model for the study of the physical principles and the physiological characteristics of embryonic gas exchange. In recent years, it has become a model for the studies of the prenatal development of pulmonary ventilation, its chemical control and its interaction with extra-pulmonary gas exchange. Differently from mammals, in birds the initiation of pulmonary ventilation and the transition from diffusive to convective gas exchange are gradual and slow-occurring events amenable to detailed investigations. The absence of the placenta and of the mother permits the study of the mechanisms of embryonic adaptation to prenatal perturbations in a way that would be impossible with mammalian preparations. First, this review summarises the general aspects of the natural history of the avian egg that are pertinent to embryonic metabolism, growth and gas exchange and the characteristics of the structures participating in gas exchange. Then, the review focuses on the embryonic development of pulmonary ventilation, its regulation in relation to the embryo's environment and metabolic state, the effects that acute or sustained changes in embryonic temperature or oxygenation can have on growth, metabolism and ventilatory control.

Somatic Donor Cell Type Correlates with Embryonic, but Not Extra-Embryonic, Gene Expression in Postimplantation Cloned Embryos

PubMed Central

Inoue, Kimiko; Ogura, Atsuo

2013-01-01

The great majority of embryos generated by somatic cell nuclear transfer (SCNT) display defined abnormal phenotypes after implantation, such as an increased likelihood of death and abnormal placentation. To gain better insight into the underlying mechanisms, we analyzed genome-wide gene expression profiles of day 6.5 postimplantation mouse embryos cloned from three different cell types (cumulus cells, neonatal Sertoli cells and fibroblasts). The embryos retrieved from the uteri were separated into embryonic (epiblast) and extraembryonic (extraembryonic ectoderm and ectoplacental cone) tissues and were subjected to gene microarray analysis. Genotype- and sex-matched embryos produced by in vitro fertilization were used as controls. Principal component analysis revealed that whereas the gene expression patterns in the embryonic tissues varied according to the donor cell type, those in extraembryonic tissues were relatively consistent across all groups. Within each group, the embryonic tissues had more differentially expressed genes (DEGs) (>2-fold vs. controls) than did the extraembryonic tissues (P<1.0×10–26). In the embryonic tissues, one of the common abnormalities was upregulation of Dlk1, a paternally imprinted gene. This might be a potential cause of the occasional placenta-only conceptuses seen in SCNT-generated mouse embryos (1–5% per embryos transferred in our laboratory), because dysregulation of the same gene is known to cause developmental failure of embryos derived from induced pluripotent stem cells. There were also some DEGs in the extraembryonic tissues, which might explain the poor development of SCNT-derived placentas at early stages. These findings suggest that SCNT affects the embryonic and extraembryonic development differentially and might cause further deterioration in the embryonic lineage in a donor cell-specific manner. This could explain donor cell-dependent variations in cloning efficiency using SCNT. PMID:24146866

Epidermal differentiation in embryos of the tuatara Sphenodon punctatus (Reptilia, Sphenodontidae) in comparison with the epidermis of other reptiles.

PubMed

Alibardi, L; Gill, B J

2007-07-01

Studying the epidermis in primitive reptiles can provide clues regarding evolution of the epidermis during land adaptation in vertebrates. With this aim, the development of the skin of the relatively primitive reptile Sphenodon punctatus in representative embryonic stages was studied by light and electron microscopy and compared with that of other reptiles previously studied. The dermis organizes into a superficial and deep portion when the epidermis starts to form the first layers. At embryonic stages comparable with those of lizards, only one layer of the inner periderm is formed beneath the outer periderm. This also occurs in lizards and snakes so far studied. The outer and inner periderm form the embryonic epidermis and accumulate thick, coarse filaments (25-30 nm thick) and sparse alpha-keratin filaments as in other reptiles. Beneath the embryonic epidermis an oberhautchen and beta-cells form small horny tips that represent overlapping borders along the margin of beta-cells that overlap other beta-cells (in a tile-like arrangement). The tips resemble those of agamine lizards but at a small scale, forming a lamellate-spinulated pattern as previously described in adult epidermis. The embryonic epidermis matures by the dispersion of coarse filaments among keratin at the end of embryonic development and is shed around hatching. The presence of these matrix organelles in the embryonic epidermis of this primitive reptile further indicates that amniote epidermis acquired interkeratin matrix proteins early for land adaptation. Unlike the condition in lizards and snakes, a shedding complex is not formed in the epidermis of embryonic S. punctatus that is like that of the adult. Therefore, as in chelonians and crocodilians, the epidermis of S. punctatus also represents an initial stage that preceded the evolution of the shedding complex for moulting.

Epidermal differentiation in embryos of the tuatara Sphenodon punctatus (Reptilia, Sphenodontidae) in comparison with the epidermis of other reptiles

PubMed Central

Alibardi, L; Gill, B J

2007-01-01

Studying the epidermis in primitive reptiles can provide clues regarding evolution of the epidermis during land adaptation in vertebrates. With this aim, the development of the skin of the relatively primitive reptile Sphenodon punctatus in representative embryonic stages was studied by light and electron microscopy and compared with that of other reptiles previously studied. The dermis organizes into a superficial and deep portion when the epidermis starts to form the first layers. At embryonic stages comparable with those of lizards, only one layer of the inner periderm is formed beneath the outer periderm. This also occurs in lizards and snakes so far studied. The outer and inner periderm form the embryonic epidermis and accumulate thick, coarse filaments (25–30 nm thick) and sparse alpha-keratin filaments as in other reptiles. Beneath the embryonic epidermis an oberhautchen and beta-cells form small horny tips that represent overlapping borders along the margin of beta-cells that overlap other beta-cells (in a tile-like arrangement). The tips resemble those of agamine lizards but at a small scale, forming a lamellate-spinulated pattern as previously described in adult epidermis. The embryonic epidermis matures by the dispersion of coarse filaments among keratin at the end of embryonic development and is shed around hatching. The presence of these matrix organelles in the embryonic epidermis of this primitive reptile further indicates that amniote epidermis acquired interkeratin matrix proteins early for land adaptation. Unlike the condition in lizards and snakes, a shedding complex is not formed in the epidermis of embryonic S. punctatus that is like that of the adult. Therefore, as in chelonians and crocodilians, the epidermis of S. punctatus also represents an initial stage that preceded the evolution of the shedding complex for moulting. PMID:17532799

Pluripotent cells in farm animals: state of the art and future perspectives.

PubMed

Nowak-Imialek, Monika; Niemann, Heiner

2012-01-01

Pluripotent cells, such as embryonic stem (ES) cells, embryonic germ cells and embryonic carcinoma cells are a unique type of cell because they remain undifferentiated indefinitely in in vitro culture, show self-renewal and possess the ability to differentiate into derivatives of the three germ layers. These capabilities make them a unique in vitro model for studying development, differentiation and for targeted modification of the genome. True pluripotent ESCs have only been described in the laboratory mouse and rat. However, rodent physiology and anatomy differ substantially from that of humans, detracting from the value of the rodent model for studies of human diseases and the development of cellular therapies in regenerative medicine. Recently, progress in the isolation of pluripotent cells in farm animals has been made and new technologies for reprogramming of somatic cells into a pluripotent state have been developed. Prior to clinical application of therapeutic cells differentiated from pluripotent stem cells in human patients, their survival and the absence of tumourigenic potential must be assessed in suitable preclinical large animal models. The establishment of pluripotent cell lines in farm animals may provide new opportunities for the production of transgenic animals, would facilitate development and validation of large animal models for evaluating ESC-based therapies and would thus contribute to the improvement of human and animal health. This review summarises the recent progress in the derivation of pluripotent and reprogrammed cells from farm animals. We refer to our recent review on this area, to which this article is complementary.

A protocol describing the use of a recombinant protein-based, animal product-free medium (APEL) for human embryonic stem cell differentiation as spin embryoid bodies.

PubMed

Ng, Elizabeth S; Davis, Richard; Stanley, Edouard G; Elefanty, Andrew G

2008-01-01

In order to promote the uniform and reproducible differentiation of human embryonic stem cells (HESCs) in response to exogenously added growth factors, we have developed a method (spin embryoid bodies (EBs)) that uses a recombinant protein-based, animal product-free medium in which HESCs are aggregated by centrifugation to form EBs. In this protocol we describe the formulation of this medium, denoted APEL (Albumin Polyvinylalcohol Essential Lipids), and its use in spin EB differentiation of HESCs. We also describe a more economical variant, BPEL (Bovine Serum Albumin (BSA) Polyvinylalchohol Essential Lipids), in which BSA replaces the recombinant human albumin. The integration of a medium that includes only defined and recombinant components with a defined number of cells to initiate EB formation results in a generally applicable, robust platform for growth factor-directed HESC differentiation.

Mechanisms of Microwave Induced Damage in Biologic Materials

DTIC Science & Technology

1992-10-01

that low level electromagnetic fields can cause developmental abnormalities in early stages of chick embryo development . In studies of the effects of...early embryonic development has led to a great deal of speculation about the safety of environmental exposure to such fields. Power lines, household...capable of covalent binding to embryonic or fetal macromolecules and nucleic acids, disrupting normal development . Individuals with low levels of

Comparison of defined culture systems for feeder cell free propagation of human embryonic stem cells

PubMed Central

Akopian, Veronika; Beil, Stephen; Benvenisty, Nissim; Brehm, Jennifer; Christie, Megan; Ford, Angela; Fox, Victoria; Gokhale, Paul J.; Healy, Lyn; Holm, Frida; Hovatta, Outi; Knowles, Barbara B.; Ludwig, Tenneille E.; McKay, Ronald D. G.; Miyazaki, Takamichi; Nakatsuji, Norio; Oh, Steve K. W.; Pera, Martin F.; Rossant, Janet; Stacey, Glyn N.; Suemori, Hirofumi

2010-01-01

There are many reports of defined culture systems for the propagation of human embryonic stem cells in the absence of feeder cell support, but no previous study has undertaken a multi-laboratory comparison of these diverse methodologies. In this study, five separate laboratories, each with experience in human embryonic stem cell culture, used a panel of ten embryonic stem cell lines (including WA09 as an index cell line common to all laboratories) to assess eight cell culture methods, with propagation in the presence of Knockout Serum Replacer, FGF-2, and mouse embryonic fibroblast feeder cell layers serving as a positive control. The cultures were assessed for up to ten passages for attachment, death, and differentiated morphology by phase contrast microscopy, for growth by serial cell counts, and for maintenance of stem cell surface marker expression by flow cytometry. Of the eight culture systems, only the control and those based on two commercial media, mTeSR1 and STEMPRO, supported maintenance of most cell lines for ten passages. Cultures grown in the remaining media failed before this point due to lack of attachment, cell death, or overt cell differentiation. Possible explanations for relative success of the commercial formulations in this study, and the lack of success with other formulations from academic groups compared to previously published results, include: the complex combination of growth factors present in the commercial preparations; improved development, manufacture, and quality control in the commercial products; differences in epigenetic adaptation to culture in vitro between different ES cell lines grown in different laboratories. PMID:20186512

Hypoxia delays hematopoiesis: retention of embryonic hemoglobin and erythrocytes in larval rainbow trout, Oncorhynchus mykiss, during chronic hypoxia exposure.

PubMed

Bianchini, Kristin; Wright, Patricia A

2013-12-01

In rainbow trout development, a switch occurs from high-affinity embryonic hemoglobin (Hb) and round, embryonic erythrocytes to lower-affinity adult Hb and oval, adult erythrocytes. Our study investigated the early ontogeny of rainbow trout blood properties and the hypoxia response. We hypothesized that hypoxia exposure would delay the ontogenetic turnover of Hb and erythrocytes because retention of high-affinity embryonic Hb would facilitate oxygen loading. To test this hypothesis we developed a method of efficiently extracting blood from individual embryos and larvae and optimized several techniques for measuring hematological parameters on microliter (0.5-2.0 μl) blood samples. In chronic hypoxia (30% of oxygen saturation), stage-matched embryos and larvae possessed half the Hb concentration, erythrocyte counts and hematocrit observed in normoxia. Hypoxia-reared larvae also had threefold to sixfold higher mRNA expression of the embryonic Hb α-1, β-1 and β-2 subunits relative to stage-matched normoxia-reared larvae. Furthermore, in hypoxia, the round embryonic erythrocytic shape persisted into later developmental stages. Despite these differences, Hb-oxygen affinity (P50), cooperativity and the Root effect were unaltered in hypoxia-reared O. mykiss. The data support our hypothesis that chronic hypoxia delays the ontogenetic turnover of Hb and erythrocytes, but without the predicted functional consequences (i.e. higher than expected P50). These results also suggest that the Hb-oxygen affinity is protected during development in chronic hypoxia to favor oxygen unloading at the tissues. We conclude that in early trout development, the blood-oxygen transport system responds very differently to chronic hypoxia relative to adults, possibly because respiration depends relatively more on oxygen diffusion than convection.

Functional optical coherence tomography for live dynamic analysis of mouse embryonic cardiogenesis

NASA Astrophysics Data System (ADS)

Wang, Shang; Lopez, Andrew L.; Larina, Irina V.

2018-02-01

Blood flow, heart contraction, and tissue stiffness are important regulators of cardiac morphogenesis and function during embryonic development. Defining how these factors are integrated is critically important to advance prevention, diagnostics, and treatment of congenital heart defects. Mammalian embryonic development is taking place deep within the female body, which makes cardiodynamic imaging and analysis during early developmental stages in humans inaccessible. With thousands of mutant lines available and well-established genetic manipulation tools, mouse is a great model to understand how biomechanical factors are integrated with molecular pathways to regulate cardiac function and development. Dynamic imaging and quantitative analysis of the biomechanics of live mouse embryos have become increasingly important, which demands continuous advancements in imaging techniques and live assessment approaches. This has been one of the major drives to keep pushing the frontier of embryonic imaging for better resolution, higher speed, deeper penetration, and more diverse and effective contrasts. Optical coherence tomography (OCT) has played a significant role in addressing such demands, and its features in non-labeling imaging, 3D capability, a large working distance, and various functional derivatives allow OCT to cover a number of specific applications in embryonic imaging. Recently, our group has made several technical improvements in using OCT to probe the biomechanical aspects of live developing mouse embryos at early stages. These include the direct volumetric structural and functional imaging of the cardiodynamics, four-dimensional quantitative Doppler imaging and analysis of the cardiac blood flow, and fourdimensional blood flow separation from the cardiac wall tissue in the beating embryonic heart. Here, we present a short review of these studies together with brief descriptions of the previous work that demonstrate OCT as a valuable and useful imaging tool for the research in developmental cardiology.

Identification of FGF-dependent genes in the Drosophila tracheal system.

PubMed

Stahl, Markus; Schuh, Reinhard; Adryan, Boris

2007-01-01

The embryonic development of the tracheal system of the fruit fly Drosophila provides a paradigm for genetic studies of branching morphogenesis. Efforts of many laboratories have identified Branchless (Bnl, a fibroblast growth factor homologue) and Breathless (Btl, the receptor homologue) as crucial factors at many stages of tracheal system development. The downstream targets of the Bnl/Btl signalling cascade, however, remain mostly unknown. Misexpression of the bnl gene results in specific tracheal phenotypes that lead to larval death. We characterised the transcriptional profiles of targeted over-expression of bnl in the embryonic trachea and of loss-of-function bnl(P1) mutant embryos. Gene expression data was mapped to high-throughput in situ hybridisation based ImaGO-annotation. Thus, we identified and confirmed by quantitative PCR 13 Bnl-dependent genes that are expressed in cells within and outside of the tracheal system.

Development of an in vitro culture method for stepwise differentiation of mouse embryonic stem cells and induced pluripotent stem cells into mature osteoclasts.

PubMed

Nishikawa, Keizo; Iwamoto, Yoriko; Ishii, Masaru

2014-05-01

The development of methods for differentiation of embryonic stem cells (ESCs) and induced pluripotent stem cell (iPSCs) into functional cells have helped to analyze the mechanism regulating cellular processes and to explore cell-based assays for drug discovery. Although several reports have demonstrated methods for differentiation of mouse ESCs into osteoclast-like cells, it remains unclear whether these methods are applicable for differentiation of iPSCs to osteoclasts. In this study, we developed a simple method for stepwise differentiation of mouse ESCs and iPSCs into bone-resorbing osteoclasts based upon a monoculture approach consisting of three steps. First, based on conventional hanging-drop methods, embryoid bodies (EBs) were produced from mouse ESCs or iPSCs. Second, EBs were cultured in medium supplemented with macrophage colony-stimulating factor (M-CSF), and differentiated to osteoclast precursors, which expressed CD11b. Finally, ESC- or iPSC-derived osteoclast precursors stimulated with receptor activator of nuclear factor-B ligand (RANKL) and M-CSF formed large multinucleated osteoclast-like cells that expressed tartrate-resistant acid phosphatase and were capable of bone resorption. Molecular analysis showed that the expression of osteoclast marker genes such as Nfatc1, Ctsk, and Acp5 are increased in a RANKL-dependent manner. Thus, our procedure is simple and easy and would be helpful for stem cell-based bone research.

Histology Atlas of the Developing Mouse Hepatobiliary System with Emphasis on Embryonic Days 9.5-18.5

PubMed Central

Crawford, Laura Wilding; Foley, Julie F.; Elmore, Susan A.

2012-01-01

Animal model phenotyping, in utero exposure toxiciy studies, and investigation into causes of embryonic, fetal, or perinatal deaths have required pathologists to recognize and diagnose developmental disorders in spontaneous and engineered mouse models of disease. In mammals, the liver is the main site of hematopoiesis during fetal development, has endocrine and exocrine functions important for maintaining homeostasis in fetal and adult life; and performs other functions including waste detoxification, production and removal of glucose, glycogen storage, triglyceride and fatty acid processing, and serum protein production. Due to its role in many critical functions, alterations in the size, morphology, or function(s) of the liver often lead to embryonic lethality. Many publications and websites describe individual aspects of hepatobiliary development at defined stages. However, no single resource provides a detailed histological evaluation of H&E-stained sections of the developing murine liver and biliary systems using high-magnification and high-resolution color images. The work herein provides a histology atlas of hepatobiliary development between embryonic days 9.5-18.5. Although the focus of this work is normal hepatobiliary development, common defects in liver development are also described as a reference for pathologists who may be asked to phenotype mice with congenital, inherited, or treatment-related hepatobiliary defects. PMID:20805319

Stage-dependent and locus-specific role of histone demethylase Jumonji D3 (JMJD3) in the embryonic stages of lung development.

PubMed

Li, Qingtian; Wang, Helen Y; Chepelev, Iouri; Zhu, Qingyuan; Wei, Gang; Zhao, Keji; Wang, Rong-Fu

2014-07-01

Histone demethylases have emerged as important players in developmental processes. Jumonji domain containing-3 (Jmjd3) has been identified as a key histone demethylase that plays a critical role in the regulation of gene expression; however, the in vivo function of Jmjd3 in embryonic development remains largely unknown. To this end, we generated Jmjd3 global and conditional knockout mice. Global deletion of Jmjd3 induces perinatal lethality associated with defective lung development. Tissue and stage-specific deletion revealed that Jmjd3 is dispensable in the later stage of embryonic lung development. Jmjd3 ablation downregulates the expression of genes critical for lung development and function, including AQP-5 and SP-B. Jmjd3-mediated alterations in gene expression are associated with locus-specific changes in the methylation status of H3K27 and H3K4. Furthermore, Jmjd3 is recruited to the SP-B promoter through interactions with the transcription factor Nkx2.1 and the epigenetic protein Brg1. Taken together, these findings demonstrate that Jmjd3 plays a stage-dependent and locus-specific role in the mouse lung development. Our study provides molecular insights into the mechanisms by which Jmjd3 regulates target gene expression in the embryonic stages of lung development.

Enzymatic Metabolism of Vitamin A in Developing Vertebrate Embryos

PubMed Central

Metzler, Melissa A.; Sandell, Lisa L.

2016-01-01

Embryonic development is orchestrated by a small number of signaling pathways, one of which is the retinoic acid (RA) signaling pathway. Vitamin A is essential for vertebrate embryonic development because it is the molecular precursor of the essential signaling molecule RA. The level and distribution of RA signaling within a developing embryo must be tightly regulated; too much, or too little, or abnormal distribution, all disrupt embryonic development. Precise regulation of RA signaling during embryogenesis is achieved by proteins involved in vitamin A metabolism, retinoid transport, nuclear signaling, and RA catabolism. The reversible first step in conversion of the precursor vitamin A to the active retinoid RA is mediated by retinol dehydrogenase 10 (RDH10) and dehydrogenase/reductase (SDR family) member 3 (DHRS3), two related membrane-bound proteins that functionally activate each other to mediate the interconversion of retinol and retinal. Alcohol dehydrogenase (ADH) enzymes do not contribute to RA production under normal conditions during embryogenesis. Genes involved in vitamin A metabolism and RA catabolism are expressed in tissue-specific patterns and are subject to feedback regulation. Mutations in genes encoding these proteins disrupt morphogenesis of many systems in a developing embryo. Together these observations demonstrate the importance of vitamin A metabolism in regulating RA signaling during embryonic development in vertebrates. PMID:27983671

Effect of temperature during embryonic development and first feeding of Trichogaster leeri larvae.

PubMed

Pereira, Samuel Louzada; de Andrade, Dalcio Ricardo; Radael, Marcella Costa; Fosse Filho, João Carlos; de Azevedo, Rafael Vieira; Mattos, Douglas da Cruz; Vidal Junior, Manuel Vazquez

2016-10-01

Temperature is an environmental factor that influences the development of fish, and when changed abruptly can lead to high mortality. Some species of fish are influenced by this factor, exhibiting a longer time for embryonic development and time to first feeding. This study aims to evaluate the effect of water temperature on embryonic and larval development up to first feeding, to describe the time in hours post fertilization (hpf) of the emergence of different structures and to determine the best hatching rate and survival of animals under different treatments. Five different egg incubation temperatures were used (24, 26, 28, 30 or 32°C, respectively). The eggs were observed at regular intervals of 30 min up to 24 h, every 2 h until 48 h and every 4 h until the display of first feeding in all treatments. Embryonic development was longer for eggs incubated at 24°C and the best results for hatching rate and survival of spawning efficiency were at 28°C. We recommend that incubation of Trichogaster leeri eggs is carried out at 28°C up to the first feeding of larvae.

How the embryonic chick brain twists.

PubMed

Chen, Zi; Guo, Qiaohang; Dai, Eric; Forsch, Nickolas; Taber, Larry A

2016-11-01

During early development, the tubular embryonic chick brain undergoes a combination of progressive ventral bending and rightward torsion, one of the earliest organ-level left-right asymmetry events in development. Existing evidence suggests that bending is caused by differential growth, but the mechanism for the predominantly rightward torsion of the embryonic brain tube remains poorly understood. Here, we show through a combination of in vitro experiments, a physical model of the embryonic morphology and mechanics analysis that the vitelline membrane (VM) exerts an external load on the brain that drives torsion. Our theoretical analysis showed that the force is of the order of 10 micronewtons. We also designed an experiment to use fluid surface tension to replace the mechanical role of the VM, and the estimated magnitude of the force owing to surface tension was shown to be consistent with the above theoretical analysis. We further discovered that the asymmetry of the looping heart determines the chirality of the twisted brain via physical mechanisms, demonstrating the mechanical transfer of left-right asymmetry between organs. Our experiments also implied that brain flexure is a necessary condition for torsion. Our work clarifies the mechanical origin of torsion and the development of left-right asymmetry in the early embryonic brain. © 2016 The Author(s).

Ca2+ signalling and early embryonic patterning during zebrafish development.

PubMed

Webb, Sarah E; Miller, Andrew L

2007-09-01

1. It has been proposed that Ca2+ signalling, in the form of pulses, waves and steady gradients, may play a crucial role in key pattern-forming events during early vertebrate development. 2. With reference to the embryo of the zebrafish (Danio rerio), herein we review the Ca2+ transients reported from the cleavage to segmentation periods. This time-window includes most of the major pattern-forming events of early development, which transform a single-cell zygote into a complex multicellular embryo with established primary germ layers and body axes. 3. Data are presented to support our proposal that intracellular Ca2+ waves are an essential feature of embryonic cytokinesis and that propagating intercellular Ca2+ waves (both long and short range) may play a crucial role in: (i) the establishment of the embryonic periderm and the coordination of cell movements during epiboly, convergence and extension; (ii) the establishment of the basic embryonic axes and germ layers; and (iii) definition of the morphological boundaries of specific tissue domains and embryonic structures, including future organ anlagen. 4. The potential downstream targets of these Ca2+ transients are also discussed, as well as how they may integrate with other pattern-forming signalling pathways known to modulate early developmental events.

The Evolutionary Economics of Embryonic-Sac Fluids in Squamate Reptiles.

PubMed

Bonnet, Xavier; Naulleau, Guy; Shine, Richard

2017-03-01

The parchment-shelled eggs of squamate reptiles take up substantial water from the nest environment, enabling the conversion of yolk into neonatal tissue and buffering the embryo against the possibility of subsequent dry weather. During development, increasing amounts of water are stored in the embryonic sacs (i.e., membranes around the embryo: amnion, allantois, and chorion). The evolution of viviparity (prolonged uterine retention of developing embryos) means that embryonic-sac fluid storage now imposes a cost (increased maternal burdening), confers less benefit (because the mother buffers fetal water balance), and introduces a potential conflict among uterine siblings (for access to finite water supplies). Our data on nine species of squamate reptiles and published information on three species show that the embryonic-sac fluids comprise around 33% of neonatal mass in viviparous species versus 94% in full-term eggs of oviparous squamates. Data on parturition in 149 vipers (Vipera aspis, a viviparous species) show that larger offspring store more fluids in their fetal sacs and that an increase in litter size is associated with a decrease in fluid-sac mass per offspring. Overall, the evolutionary transition from oviparity to viviparity may have substantially altered selective forces on offspring packaging and created competition among offspring for access to water reserves during embryonic development.

Embryonic development of Ampheres leucopheus and Iporangaia pustulosa (Arachnida: Opiliones: Gonyleptidae).

PubMed

Gnaspini, Pedro; Lerche, Cristiano Frederico

2010-09-15

The first studies concerning the embryonic development of harvestmen started in the late 19th century, and focused mostly on holarctic species, and only three species of the suborder Laniatores (the largest, among the four suborders considered presently) were studied. Moreover, the last studies on embryology of harvestmen were made during the late 1970s. This study focused on the embryonic development of Ampheres leucopheus (Gonyleptidae, Caelopyginae) and Iporangaia pustulosa (Gonyleptidae, Progonyleptoidellinae). The embryonic development was followed in the field, by taking daily photographs of different eggs during about 2 months. When laid, eggs of A. leucopheus and I. pustulosa have approximately 1.13 and 1.30 mm in diameter, respectively, and the second is embedded in a large amount of mucus. The eggs grow, mainly due to water absorption at the beginning of the process, and they reach a diameter of about 1.35 and 1.59 mm, respectively, close to hatching. It took, respectively, 29-56 days and 35-66 days from egg laying to hatching. For the description of the embryonic development, we use photographs from the field, SEM micrographs, and histological analysis. This allowed us, for instance, to document the progression of structures and pigmentation directly from live embryos in the field, and to record microstructures, such as the presence of perforations in the cuticle of the embryo in the place where eyes are developing. Yet, contrary to what was expected in the literature, we record an egg tooth in one of the studied laniatoreans. (c) 2010 Wiley-Liss, Inc.

Micromanaging cardiac regeneration: Targeted delivery of microRNAs for cardiac repair and regeneration

PubMed Central

Kamps, Jan AAM; Krenning, Guido

2016-01-01

The loss of cardiomyocytes during injury and disease can result in heart failure and sudden death, while the adult heart has a limited capacity for endogenous regeneration and repair. Current stem cell-based regenerative medicine approaches modestly improve cardiomyocyte survival, but offer neglectable cardiomyogenesis. This has prompted the need for methodological developments that crease de novo cardiomyocytes. Current insights in cardiac development on the processes and regulatory mechanisms in embryonic cardiomyocyte differentiation provide a basis to therapeutically induce these pathways to generate new cardiomyocytes. Here, we discuss the current knowledge on embryonic cardiomyocyte differentiation and the implementation of this knowledge in state-of-the-art protocols to the direct reprogramming of cardiac fibroblasts into de novo cardiomyocytes in vitro and in vivo with an emphasis on microRNA-mediated reprogramming. Additionally, we discuss current advances on state-of-the-art targeted drug delivery systems that can be employed to deliver these microRNAs to the damaged cardiac tissue. Together, the advances in our understanding of cardiac development, recent advances in microRNA-based therapeutics, and innovative drug delivery systems, highlight exciting opportunities for effective therapies for myocardial infarction and heart failure. PMID:26981212

Unique patterns of organization and migration of FGF-expressing cells during Drosophila morphogenesis.

PubMed

Du, Lijuan; Zhou, Amy; Patel, Akshay; Rao, Mishal; Anderson, Kelsey; Roy, Sougata

2017-07-01

Fibroblast growth factors (FGF) are essential signaling proteins that regulate diverse cellular functions in developmental and metabolic processes. In Drosophila, the FGF homolog, branchless (bnl) is expressed in a dynamic and spatiotemporally restricted pattern to induce branching morphogenesis of the trachea, which expresses the Bnl-receptor, breathless (btl). Here we have developed a new strategy to determine bnl- expressing cells and study their interactions with the btl-expressing cells in the range of tissue patterning during Drosophila development. To enable targeted gene expression specifically in the bnl expressing cells, a new LexA based bnl enhancer trap line was generated using CRISPR/Cas9 based genome editing. Analyses of the spatiotemporal expression of the reporter in various embryonic stages, larval or adult tissues and in metabolic hypoxia, confirmed its target specificity and versatility. With this tool, new bnl expressing cells, their unique organization and functional interactions with the btl-expressing cells were uncovered in a larval tracheoblast niche in the leg imaginal discs, in larval photoreceptors of the developing retina, and in the embryonic central nervous system. The targeted expression system also facilitated live imaging of simultaneously labeled Bnl sources and tracheal cells, which revealed a unique morphogenetic movement of the embryonic bnl- source. Migration of bnl- expressing cells may create a dynamic spatiotemporal pattern of the signal source necessary for the directional growth of the tracheal branch. The genetic tool and the comprehensive profile of expression, organization, and activity of various types of bnl-expressing cells described in this study provided us with an important foundation for future research investigating the mechanisms underlying Bnl signaling in tissue morphogenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

Developmental toxicity of treated municipal wastewater effluent on Bombina orientalis (Amphibia: Anura) embryos.

PubMed

Park, Chan Jin; Ahn, Hyo Min; Cho, Seong Chan; Kim, Tae-Hoon; Oh, Jong-Min; Ahn, Hong Kyu; Chun, Seung-Hoon; Gye, Myung Chan

2014-04-01

Amphibian populations have been decreasing in urban freshwater systems in Korea. To elucidate the biological safety of treated wastewater effluent (TWE) in the Tancheon basin, the capital area of Korea, a 7-d-exposure Bombina orientalis embryo developmental toxicity assay was examined during the breeding season. In March, there were no significant differences in embryonic survival or malformation among the water samples. In July, following monsoon precipitation, embryonic lethality in TWE was significantly higher than in the upstream water sample. Malformation in TWE and TWE-mixed waters was significantly higher than in the control and upstream water samples. Tail muscle height of tadpoles also significantly decreased in TWE and TWE-mixed waters. Heavy metals were not detected in any samples. Total nitrogen, total phosphorous, and chemical oxygen demand in TWE markedly increased together with a decrease in dissolved oxygen in July. The increase in organic and inorganic loading following precipitation could have made TWE and TWE-mixed water not suitable for embryonic development. Though being managed based on physicochemical criteria, the water quality of TWE may not be sufficient to assure normal development of amphibian embryos. An amphibian developmental toxicity assay would be helpful for the water-quality management of TWE and urban freshwater systems in Korea. © 2014 SETAC.

Conference Scene: Induced pluripotent cells: a new path for regenerative medicine. 7 October 2010, BioPark, Welwyn Garden City, Hertfordshire, UK.

PubMed

Crutzen, Hélène S G

2011-01-01

Embryonic stem cells and induced pluripotent stem (iPS) cells, which are embryonic stem-like cells derived from adult tissues, have the broadest differentiation potential. These cells are unique in their ability to self-renew, to be maintained in an undifferentiated state for long periods of culturing and to give rise to many different cell lineages including germ-line cells. They therefore represent an invaluable tool for facilitating research towards the realization of regenerative medicine. The recent developments in embryonic stem cell and iPS cell technology have allowed human cell models to be developed that will hopefully provide novel platforms for disease analysis not only at the basic science level, but also for drug discovery and screening, and other clinical applications. This 1-day conference, chaired by Professor Peter Andrews from the University of Sheffield, UK, and Dr Chris Denning from the University of Nottingham, UK, focused on generation of iPS cells, their differentiation into specific fates and applications to disease modeling. It consisted of 11 talks by UK-based and international researchers, and three posters; Ms Azra Fatima from Cologne University, Germany, won the competition for her poster on the derivation of iPS cells from a patient with arrhythmogenic right ventricular cardiomyopathy.

The transcriptional landscape of hematopoietic stem cell ontogeny

PubMed Central

McKinney-Freeman, Shannon; Cahan, Patrick; Li, Hu; Lacadie, Scott A.; Huang, Hsuan-Ting; Curran, Matthew; Loewer, Sabine; Naveiras, Olaia; Kathrein, Katie L.; Konantz, Martina; Langdon, Erin M.; Lengerke, Claudia; Zon, Leonard I.; Collins, James J.; Daley, George Q.

2012-01-01

Transcriptome analysis of adult hematopoietic stem cells (HSC) and their progeny has revealed mechanisms of blood differentiation and leukemogenesis, but a similar analysis of HSC development is lacking. Here, we acquired the transcriptomes of developing HSC purified from >2500 murine embryos and adult mice. We found that embryonic hematopoietic elements clustered into three distinct transcriptional states characteristic of the definitive yolk sac, HSCs undergoing specification, and definitive HSCs. We applied a network biology-based analysis to reconstruct the gene regulatory networks of sequential stages of HSC development and functionally validated candidate transcriptional regulators of HSC ontogeny by morpholino-mediated knock-down in zebrafish embryos. Moreover, we found that HSCs from in vitro differentiated embryonic stem cells closely resemble definitive HSC, yet lack a Notch-signaling signature, likely accounting for their defective lymphopoiesis. Our analysis and web resource (http://hsc.hms.harvard.edu) will enhance efforts to identify regulators of HSC ontogeny and facilitate the engineering of hematopoietic specification. PMID:23122293

Can the eastern red-backed salamander (Plethodon cinereus) persist in an acidified landscape?

Treesearch

Cheryl A. Bondi; Colin M. Beier; Peter K. Ducey; Gregory B. Lawrence; Scott Bailey

2016-01-01

Hardwood forests of eastern North America have experienced decades of acidic deposition, leading to soil acidification where base cation supply was insufficient to neutralize acid inputs. Negative impacts of soil acidity on amphibians include disrupted embryonic development, lower growth rates, and habitat loss. However, some amphibians exhibit intraspecific variation...

ChickScope: An Interactive MRI Classroom Curriculum Innovation for K-12.

ERIC Educational Resources Information Center

Bruce, B. C.; Carragher, B. O.; Damon, B. M.; Dawson, M. J.; Eurell, J. A.; Gregory, C. D.; Lauterbur, P. C.; Marjanovic, M. M.; Mason-Fossum, B.; Morris, H. D.; Potter, C. S.; Thakkar, U.

1997-01-01

Describes ChickScope, a 21-day chick embryonic development project, to demonstrate the remote control of a magnetic resonance imaging (MRI) instrument through the World Wide Web. Topics include remote instrumentation and the Web, teacher-based implementation, impact in elementary and secondary school classrooms, and future directions. (Author/LRW)

A qualified endorsement of embryonic stem cell research, based on two widely shared beliefs about the brain-diseased patients such research might benefit.

PubMed

Disilvestro, R

2008-07-01

Are there persuasive approaches to embryonic stem cell (ESC) research that appeal, not just to those fellow-citizens in one's own ideological camp, nor just to those undecided citizens in the middle, but to those citizens on the other side of the issue? I believe that there are such arguments and in this short paper I try to develop one of them. In particular, I argue that certain beliefs shared by some proponents and some opponents of ESC research--beliefs about the personal identity and moral status of those who are victims of terrible brain diseases--are beliefs that should lead us to adopt a qualified endorsement of ESC research.

Expression of the ephrin receptor B2 in the embryonic chicken bursa of Fabricius

USDA-ARS?s Scientific Manuscript database

Chicken B-cells develop in a specific organ, the bursa of Fabricius. To understand the bursal microenvironment guiding B-cell development, previous studies identified ephrin (Eph) receptor B2 (EphB2) gene transcripts in the embryonic bursa. We hypothesize that the EphB2 receptors and their ligands r...

Extra-embryonic tissue spreading directs early embryo morphogenesis in killifish

PubMed Central

Reig, Germán; Cerda, Mauricio; Sepúlveda, Néstor; Flores, Daniela; Castañeda, Victor; Tada, Masazumi; Härtel, Steffen; Concha, Miguel L.

2017-01-01

The spreading of mesenchymal-like cell layers is critical for embryo morphogenesis and tissue repair, yet we know little of this process in vivo. Here we take advantage of unique developmental features of the non-conventional annual killifish embryo to study the principles underlying tissue spreading in a simple cellular environment, devoid of patterning signals and major morphogenetic cell movements. Using in vivo experimentation and physical modelling we reveal that the extra-embryonic epithelial enveloping cell layer, thought mainly to provide protection to the embryo, directs cell migration and the spreading of embryonic tissue during early development. This function relies on the ability of embryonic cells to couple their autonomous random motility to non-autonomous signals arising from the expansion of the extra-embryonic epithelium, mediated by cell membrane adhesion and tension. Thus, we present a mechanism of extra-embryonic control of embryo morphogenesis that couples the mechanical properties of adjacent tissues in the early killifish embryo. PMID:28580937

Beyond the binding site: in vivo identification of tbx2, smarca5 and wnt5b as molecular targets of CNBP during embryonic development.

PubMed

Armas, Pablo; Margarit, Ezequiel; Mouguelar, Valeria S; Allende, Miguel L; Calcaterra, Nora B

2013-01-01

CNBP is a nucleic acid chaperone implicated in vertebrate craniofacial development, as well as in myotonic dystrophy type 2 (DM2) and sporadic inclusion body myositis (sIBM) human muscle diseases. CNBP is highly conserved among vertebrates and has been implicated in transcriptional regulation; however, its DNA binding sites and molecular targets remain elusive. The main goal of this work was to identify CNBP DNA binding sites that might reveal target genes involved in vertebrate embryonic development. To accomplish this, we used a recently described yeast one-hybrid assay to identify DNA sequences bound in vivo by CNBP. Bioinformatic analyses revealed that these sequences are G-enriched and show high frequency of putative G-quadruplex DNA secondary structure. Moreover, an in silico approach enabled us to establish the CNBP DNA-binding site and to predict CNBP putative targets based on gene ontology terms and synexpression with CNBP. The direct interaction between CNBP and candidate genes was proved by EMSA and ChIP assays. Besides, the role of CNBP upon the identified genes was validated in loss-of-function experiments in developing zebrafish. We successfully confirmed that CNBP up-regulates tbx2b and smarca5, and down-regulates wnt5b gene expression. The highly stringent strategy used in this work allowed us to identify new CNBP target genes functionally important in different contexts of vertebrate embryonic development. Furthermore, it represents a novel approach toward understanding the biological function and regulatory networks involving CNBP in the biology of vertebrates.

Extracellular Ca2+ Is Required for Fertilization in the African Clawed Frog, Xenopus laevis

PubMed Central

Duray, Alexis M.; Tembo, Maiwase; Beleny, David O.; Napolitano, Marc A.; Sauer, Monica L.; Wisner, Bennett W.

2017-01-01

Background The necessity of extracellular Ca2+ for fertilization and early embryonic development in the African clawed frog, Xenopus laevis, is controversial. Ca2+ entry into X. laevis sperm is reportedly required for the acrosome reaction, yet fertilization and embryonic development have been documented to occur in high concentrations of the Ca2+ chelator BAPTA. Here we sought to resolve this controversy. Methodology/principal finding Using the appearance of cleavage furrows as an indicator of embryonic development, we found that X. laevis eggs inseminated in a solution lacking added divalent cations developed normally. By contrast, eggs inseminated in millimolar concentrations of BAPTA or EGTA failed to develop. Transferring embryos to varying solutions after sperm addition, we found that extracellular Ca2+ is specifically required for events occurring within the first 30 minutes after sperm addition, but not after. We found that the fluorescently stained sperm were not able to penetrate the envelope of eggs inseminated in high BAPTA, whereas several had penetrated the vitelline envelope of eggs inseminated without a Ca2+ chelator, or with BAPTA and saturating CaCl2. Together these results indicate that fertilization does not occur in high concentrations of Ca2+ chelators. Finally, we found that the jelly coat includes >5 mM of readily diffusible Ca2+. Conclusions/Significance Taken together, these data are consistent with requirement of extracellular Ca2+ for fertilization. Based on our findings, we hypothesize that the jelly coat surrounding the egg acts as a reserve of readily available Ca2+ ions to foster fertilization in changing extracellular milieu. PMID:28114360

Beyond the Binding Site: In Vivo Identification of tbx2, smarca5 and wnt5b as Molecular Targets of CNBP during Embryonic Development

PubMed Central

Mouguelar, Valeria S.; Allende, Miguel L.; Calcaterra, Nora B.

2013-01-01

CNBP is a nucleic acid chaperone implicated in vertebrate craniofacial development, as well as in myotonic dystrophy type 2 (DM2) and sporadic inclusion body myositis (sIBM) human muscle diseases. CNBP is highly conserved among vertebrates and has been implicated in transcriptional regulation; however, its DNA binding sites and molecular targets remain elusive. The main goal of this work was to identify CNBP DNA binding sites that might reveal target genes involved in vertebrate embryonic development. To accomplish this, we used a recently described yeast one-hybrid assay to identify DNA sequences bound in vivo by CNBP. Bioinformatic analyses revealed that these sequences are G-enriched and show high frequency of putative G-quadruplex DNA secondary structure. Moreover, an in silico approach enabled us to establish the CNBP DNA-binding site and to predict CNBP putative targets based on gene ontology terms and synexpression with CNBP. The direct interaction between CNBP and candidate genes was proved by EMSA and ChIP assays. Besides, the role of CNBP upon the identified genes was validated in loss-of-function experiments in developing zebrafish. We successfully confirmed that CNBP up-regulates tbx2b and smarca5, and down-regulates wnt5b gene expression. The highly stringent strategy used in this work allowed us to identify new CNBP target genes functionally important in different contexts of vertebrate embryonic development. Furthermore, it represents a novel approach toward understanding the biological function and regulatory networks involving CNBP in the biology of vertebrates. PMID:23667590

A mesh generation and machine learning framework for Drosophila gene expression pattern image analysis

PubMed Central

2013-01-01

Background Multicellular organisms consist of cells of many different types that are established during development. Each type of cell is characterized by the unique combination of expressed gene products as a result of spatiotemporal gene regulation. Currently, a fundamental challenge in regulatory biology is to elucidate the gene expression controls that generate the complex body plans during development. Recent advances in high-throughput biotechnologies have generated spatiotemporal expression patterns for thousands of genes in the model organism fruit fly Drosophila melanogaster. Existing qualitative methods enhanced by a quantitative analysis based on computational tools we present in this paper would provide promising ways for addressing key scientific questions. Results We develop a set of computational methods and open source tools for identifying co-expressed embryonic domains and the associated genes simultaneously. To map the expression patterns of many genes into the same coordinate space and account for the embryonic shape variations, we develop a mesh generation method to deform a meshed generic ellipse to each individual embryo. We then develop a co-clustering formulation to cluster the genes and the mesh elements, thereby identifying co-expressed embryonic domains and the associated genes simultaneously. Experimental results indicate that the gene and mesh co-clusters can be correlated to key developmental events during the stages of embryogenesis we study. The open source software tool has been made available at http://compbio.cs.odu.edu/fly/. Conclusions Our mesh generation and machine learning methods and tools improve upon the flexibility, ease-of-use and accuracy of existing methods. PMID:24373308

Three-dimensional reconstruction of the embryonic pancreas in the grass snake Natrix natrix L. (Lepidosauria, Serpentes) based on histological studies.

PubMed

Kowalska, Magdalena; Hermyt, Mateusz; Rupik, Weronika

2017-04-01

The aim of this study was to evaluate two research hypotheses: H 0 -the embryonic pancreas in grass snakes develops in the same manner as in all previously investigated amniotes (from three buds) and its topographical localization within the adult body has no relation to its development; H 1 -the pancreas develops in a different manner and is related to the different topography of internal organs in snakes. For the evaluation of these hypotheses we used histological methods and three-dimensional (3D) reconstructions of the position of the pancreatic buds and surrounding organs at particular developmental stages and of the final position and shape of the pancreatic gland. Our results indicate that the pancreas primordium in the grass snake is formed by only two buds - a dorsal and a ventral one - that are not connected until the end of stage II. This differs from the majority of vertebrates investigated so far. The gall bladder of the grass snake embryos is connected with the liver only by a thin cystic duct, which also differs from many other vertebrates. Our histological study also indicates a different distribution of the endocrine cells in the embryonic pancreas of the grass snake because the first endocrine cells appeared in the dorsal part of the pancreas in a region located close to the spleen. During the entire developmental period no evidence of these cells was found in the ventral part of the pancreas. The endocrine cells form elongated, large and irregular-shaped islets. They can also form structures resembling "inverted acini". Such an arrangement is characteristic of snakes only. The differentiating pancreas penetrates the ventral part of the developing spleen and divides it into three separate parts at developmental stage IX. This is unique among vertebrates. At the end of the embryonic development (stage XI), the pancreas, the spleen and the gall bladder are located in close proximity and form the so-called triad. Our results suggest that the untypical topography of the organ systems in snakes may determine the unique development of the pancreas in these animals. Copyright © 2016 Elsevier GmbH. All rights reserved.

Impaired embryonic development in glucose-6-phosphate dehydrogenase-deficient Caenorhabditis elegans due to abnormal redox homeostasis induced activation of calcium-independent phospholipase and alteration of glycerophospholipid metabolism.

PubMed

Chen, Tzu-Ling; Yang, Hung-Chi; Hung, Cheng-Yu; Ou, Meng-Hsin; Pan, Yi-Yun; Cheng, Mei-Ling; Stern, Arnold; Lo, Szecheng J; Chiu, Daniel Tsun-Yee

2017-01-12

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a commonly pervasive inherited disease in many parts of the world. The complete lack of G6PD activity in a mouse model causes embryonic lethality. The G6PD-deficient Caenorhabditis elegans model also shows embryonic death as indicated by a severe hatching defect. Although increased oxidative stress has been implicated in both cases as the underlying cause, the exact mechanism has not been clearly delineated. In this study with C. elegans, membrane-associated defects, including enhanced permeability, defective polarity and cytokinesis, were found in G6PD-deficient embryos. The membrane-associated abnormalities were accompanied by impaired eggshell structure as evidenced by a transmission electron microscopic study. Such loss of membrane structural integrity was associated with abnormal lipid composition as lipidomic analysis revealed that lysoglycerophospholipids were significantly increased in G6PD-deficient embryos. Abnormal glycerophospholipid metabolism leading to defective embryonic development could be attributed to the increased activity of calcium-independent phospholipase A 2 (iPLA) in G6PD-deficient embryos. This notion is further supported by the fact that the suppression of multiple iPLAs by genetic manipulation partially rescued the embryonic defects in G6PD-deficient embryos. In addition, G6PD deficiency induced disruption of redox balance as manifested by diminished NADPH and elevated lipid peroxidation in embryos. Taken together, disrupted lipid metabolism due to abnormal redox homeostasis is a major factor contributing to abnormal embryonic development in G6PD-deficient C. elegans.

Effects of dieldrin treatment on physiological and biochemical aspects of the toad embryonic development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gauna, L.; Caballero de Castro, A.; Chifflet de Llamas, M.

1991-04-01

Dieldrin is a cylclodiene insecticide highly persistent in nature due to its chemical stability. The exposure of toad embryos to Dieldrin induces hyperactivity in the swimming larvae and inhibition of cholinesterases. However, the inhibition of these enzymes during early development is not life threatening. The present report provides a physiological and biochemical study of the noxious effect of Dieldrin on the toad embryonic development.

Heart Development, Diseases, and Regeneration　- New Approaches From Innervation, Fibroblasts, and Reprogramming.

PubMed

Ieda, Masaki

2016-09-23

It is well known that cardiac function is tightly controlled by neural activity; however, the molecular mechanism of cardiac innervation during development and the relationship with heart disease remain undetermined. My work has revealed the molecular networks that govern cardiac innervation and its critical roles in heart diseases such as silent myocardial ischemia and arrhythmias. Cardiomyocytes proliferate during embryonic development, but lose their proliferative capacity after birth. Cardiac fibroblasts are a major source of cells during fibrosis and induce cardiac hypertrophy after myocardial injury in the adult heart. Despite the importance of fibroblasts in the adult heart, the role of fibroblasts in embryonic heart development was previously not determined. I demonstrated that cardiac fibroblasts play important roles in myocardial growth and cardiomyocyte proliferation during embryonic development, and I identified key paracrine factors and signaling pathways. In contrast to embryonic cardiomyocytes, adult cardiomyocytes have little regenerative capacity, leading to heart failure and high mortality rates after myocardial infarction. Leveraging the knowledge of developmental biology, I identified cardiac reprogramming factors that can directly convert resident cardiac fibroblasts into cardiomyocytes for heart regeneration. These findings greatly improved our understanding of heart development and diseases, and provide a new strategy for heart regenerative therapy. (Circ J 2016; 80: 2081-2088).

Generation of induced pluripotent stem cells with high efficiency from human embryonic renal cortical cells.

PubMed

Yao, Ling; Chen, Ruifang; Wang, Pu; Zhang, Qi; Tang, Hailiang; Sun, Huaping

2016-01-01

Reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) emerges as a prospective therapeutic angle in regenerative medicine and a tool for drug screening. Although increasing numbers of iPSCs from different sources have been generated, there has been limited progress in yield of iPSC. Here, we show that four Yamanaka factors Oct4, Sox2, Klf4 and c-Myc can convert human embryonic renal cortical cells (hERCCs) to pluripotent stem cells with a roughly 40-fold higher reprogramming efficiency compared with that of adult human dermal fibroblasts. These iPSCs show pluripotency in vitro and in vivo, as evidenced by expression of pluripotency associated genes, differentiation into three embryonic germ layers by teratoma tests, as well as neuronal fate specification by embryoid body formation. Moreover, the four exogenous genes are effectively silenced in these iPSCs. This study highlights the use of hERCCs to generate highly functional human iPSCs which may aid the study of genetic kidney diseases and accelerate the development of cell-based regenerative therapy.

Uniform Embryoid Body Production and Enhanced Mesendoderm Differentiation with Murine Embryonic Stem Cells in a Rotary Suspension Bioreactor.

PubMed

Lei, Xiaohua; Deng, Zhili; Duan, Enkui

2016-01-01

Embryonic stem cells (ESCs) are capable of differentiating into almost all cell types in vitro and hold great promise for drug screening, developmental studies and have a huge potential in many therapeutic areas. ESCs can aggregate to form embryoid body (EB) in static suspension culture by spontaneous differentiation, which resembles an intact embryo; while static suspension culture cannot prevent agglomeration of cells and offers little control over the size and shape of EBs, it results in aggregation of EBs into large, irregular masses, which prejudice the efficiency of differentiation of cells. Recently, bioreactor-based platforms have been shown to not only offer a beneficial effect on increasing diffusion of nutrients and oxygen which promotes cell viability and proliferation but also display local biomechanical properties (e.g., low fluid shear stresses and hydrodynamic force) in tissue development and organogenesis. This chapter describes a protocol for using a rotary suspension bioreactor to produce embryoid bodies and process the differentiation of mouse embryonic stem cells (mESCs), and to assess the efficiency of EB differentiation in the bioreactor by real-time PCR and immunostaining.

Effect of the anti-androgenic endocrine disruptor vinclozolin on embryonic testis cord formation and postnatal testis development and function.

PubMed

Uzumcu, Mehmet; Suzuki, Hiroetsu; Skinner, Michael K

2004-01-01

Vinclozolin is a systemic dicarboximide fungicide that is used on fruits, vegetables, ornamental plants, and turf grass. Vinclozolin and its metabolites are known to be endocrine disruptors and act as androgen receptor antagonists. The hypothesis tested in the current study is that transient embryonic exposure to an anti-androgenic endocrine disruptor at the time of testis determination alters testis development and subsequently influences adult spermatogenic capacity and male reproduction. The effects of vinclozolin on embryonic testicular cord formation in vitro were examined, as well as the effects of transient in utero vinclozolin exposure on postnatal testis development and function. Embryonic day 13 (E13, sperm-positive vaginal smear day = E0) gonads were cultured in the absence or presence of vinclozolin (50-500microM). Vinclozolin treated gonads had significantly fewer cords (P < 0.05) and the histology of the cords that formed were abnormal as compared to vehicle-treated organs. Pregnant rats were exposed to vinclozolin (100 mg/kg/day) between embryonic days 8 and 14 (E8-E14) of development. Testis morphology and function were analyzed from postnatal day (P) 0, pubertal P20, and adult P60. No significant effect of vinclozolin on testis histology or germ cell viability was observed in P0 testis. The pubertal P20 testis from vinclozolin exposed animals had significantly higher numbers of apoptotic germ cells (P < 0.01), but testis weight was not affected. The adult P60 sperm motility was significantly lower in vinclozolin exposed males (P < 0.01). In addition, apoptotic germ cell number in testis of vinclozolin exposed animals was higher in adult P60 animals. Observations demonstrate that vinclozolin can effect embryonic testicular cord formation in vitro and that transient in utero exposure to vinclozolin increases apoptotic germ cell numbers in the testis of pubertal and adult animals. This correlated to reduced sperm motility in the adult. In conclusion, transient exposure to vinclozolin during the time of testis differentiation (i.e. cord formation) alters testis development and function. Observations indicate that transient exposure to an anti-androgenic endocrine disruptor during embryonic development causes delayed effects later in adult life on spermatogenic capacity.

Reactivation of the Nkx2.5 cardiac enhancer after myocardial infarction does not presage myogenesis.

PubMed

Deutsch, Marcus-André; Doppler, Stefanie A; Li, Xinghai; Lahm, Harald; Santamaria, Gianluca; Cuda, Giovanni; Eichhorn, Stefan; Ratschiller, Thomas; Dzilic, Elda; Dreßen, Martina; Eckart, Annekathrin; Stark, Konstantin; Massberg, Steffen; Bartels, Anna; Rischpler, Christoph; Gilsbach, Ralf; Hein, Lutz; Fleischmann, Bernd K; Wu, Sean M; Lange, Rüdiger; Krane, Markus

2018-03-20

The contribution of resident stem or progenitor cells to cardiomyocyte renewal after injury in adult mammalian hearts remains a matter of considerable debate. We evaluated a cell population in the adult mouse heart induced by myocardial infarction (MI) and characterized by an activated Nkx2.5 enhancer element that is specific for multipotent cardiac progenitor cells during embryonic development. We hypothesized that these MI induced cells (MICs) harbor cardiomyogenic properties similar to their embryonic counterparts. MICs reside in the heart and mainly localize to the infarction area and border zone. Interestingly, gene expression profiling of purified MICs one week after infarction revealed increased expression of stem cell markers and embryonic cardiac transcription factors in these cells as compared to the non-mycoyte cell fraction of adult hearts. A subsequent global transcriptome comparison with embryonic cardiac progenitor cells and fibroblasts and in vitro culture of MICs unveiled that (myo-) fibroblastic features predominated and that cardiac transcription factors were only expressed at background levels. Adult injury induced reactivation of a cardiac-specific Nkx2.5 enhancer element known to specifically mark myocardial progenitor cells during embryonic development does not reflect hypothesized embryonic cardiomyogenic properties. Our data suggest a decreasing plasticity of cardiac progenitor (-like) cell populations with increasing age. A re-expression of embryonic, stem or progenitor cell features in the adult heart must be interpreted very carefully with respect to the definition of cardiac resident progenitor cells. Albeit, the abundance of scar formation after cardiac injury suggests a potential to target predestinated activated profibrotic cells to push them towards cardiomyogenic differentiation to improve regeneration.

Embryonic development during chronic acceleration

NASA Technical Reports Server (NTRS)

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

1982-01-01

Experiments carried out on chicken eggs indicate that the embryo is affected during very early development, especially over the first four days, and during hatching. In the first four days, the brain develops as well as the anlage for all other organs. In addition, the heart commences to function and the extraembryonic membranes that compartmentalize the egg contents form. The latter require an appreciable extension and folding of tissue which may be disrupted by the mechanical load. Observations of embryonic abnormalities that occur during chronic acceleration suggest an inhibition of development of the axial skeleton, which is rarely seen otherwise, a general retardation of embryonic growth, and circulatory problems. The final stages of development (after 18 days) involve the uptake of fluids, the transition to aerial respiration, and the reorientation of the embryo into a normal hatching position. At 4 G mortality is very high during this period, with a majority of embryos failing to reorient into the normal hatching position.

Gravity and embryonic development

NASA Technical Reports Server (NTRS)

Young, R. S.

1976-01-01

The relationship between the developing embryo (both plant and animal) and a gravitational field has long been contemplated. The difficulty in designing critical experiments on the surface of the earth because of its background of 1 g, has been an obstacle to a resolution of the problem. Biological responses to gravity (particularly in plants) are obvious in many cases; however, the influence of gravity as an environmental input to the developing embryo is not as obvious and has proven to be extremely difficult to define. In spite of this, over the years numerous attempts have been made using a variety of embryonic materials to come to grips with the role of gravity in development. Three research tools are available: the centrifuge, the clinostat, and the orbiting spacecraft. Experimental results are now available from all three sources. Some tenuous conclusions are drawn, and an attempt at a unifying theory of gravitational influence on embryonic development is made.

Establishment of novel detection system for embryonic stem cell-derived hepatocyte-like cells based on nongenetic manipulation with indocyanine green.

PubMed

Yoshie, Susumu; Ito, Jun; Shirasawa, Sakiko; Yokoyama, Tadayuki; Fujimura, Yuu; Takeda, Kazuo; Mizuguchi, Masahiro; Matsumoto, Ken; Tomotsune, Daihachiro; Sasaki, Katsunori

2012-01-01

Hepatocytes derived from embryonic stem cells (ESCs) are expected to be useful for basic research and clinical applications. However, in several studies, genetic methods used to detect and obtain them are difficult and pose major safety problems. Therefore, in this study, we established a novel detection system for hepatocytes by using indocyanine green (ICG), which is selectively taken up by hepatocytes, based on nongenetic manipulation. ICG has maximum light absorption near 780 nm, and it fluoresces between 800 and 900 nm. Making use of these properties, we developed flow cytometry equipped with an excitation lazer of 785 nm and specific bandpass filters and successfully detected ESC-derived ICG-positive cells that were periodic acid-Schiff positive and expressed hepatocyte phenotypic mRNAs. These results demonstrate that this detection system based on nongenetic manipulation with ICG will lead to isolate hepatocytes generated from ESCs and provide the appropriate levels of stability, quality, and safety required for cell source for cell-based therapy and pharmaceutical studies such as toxicology.

Sperm and Spermatids Contain Different Proteins and Bind Distinct Egg Factors

PubMed Central

Teperek, Marta; Miyamoto, Kei; Simeone, Angela; Feret, Renata; Deery, Michael J.; Gurdon, John B.; Jullien, Jerome

2014-01-01

Spermatozoa are more efficient at supporting normal embryonic development than spermatids, their immature, immediate precursors. This suggests that the sperm acquires the ability to support embryonic development during spermiogenesis (spermatid to sperm maturation). Here, using Xenopus laevis as a model organism, we performed 2-D Fluorescence Difference Gel Electrophoresis (2D-DIGE) and mass spectrometry analysis of differentially expressed proteins between sperm and spermatids in order to identify factors that could be responsible for the efficiency of the sperm to support embryonic development. Furthermore, benefiting from the availability of egg extracts in Xenopus, we also tested whether the chromatin of sperm could attract different egg factors compared to the chromatin of spermatids. Our analysis identified: (1) several proteins which were present exclusively in sperm; but not in spermatid nuclei and (2) numerous egg proteins binding to the sperm (but not to the spermatid chromatin) after incubation in egg extracts. Amongst these factors we identified many chromatin-associated proteins and transcriptional repressors. Presence of transcriptional repressors binding specifically to sperm chromatin could suggest its preparation for the early embryonic cell cycles, during which no transcription is observed and suggests that sperm chromatin has a unique protein composition, which facilitates the recruitment of egg chromatin remodelling factors. It is therefore likely that the acquisition of these sperm-specific factors during spermiogenesis makes the sperm chromatin suitable to interact with the maternal factors and, as a consequence, to support efficient embryonic development. PMID:25244019

Deep RNA sequencing of pectoralis muscle transcriptomes during late-term embryonic to neonatal development in indigenous Chinese duck breeds

PubMed Central

Tao, Zhiyun; Liu, Hongxiang; Xu, Wenjuan; Zhang, Shuangjie; Li, Huifang

2017-01-01

Pectoral muscle (PM) comprises an important component of overall meat mass in ducks. However, PM has shown arrested or even reduced growth during late embryonic development, and the molecular mechanisms underlying PM growth during the late embryonic to neonatal period in ducks have not been addressed. In this study, we characterized potential candidate genes and signaling pathways related to PM development using RNA sequencing of PM samples selected at embryonic days (E) 21 and 27 and 5 days post-hatch (dph) in two duck breeds (Gaoyou and Jinding ducks). A total of 393 differentially expressed genes (DEGs) were identified, which showed higher or lower expression levels at E27 compared with E21 and 5 dph, reflecting the pattern of PM growth rates. Among these, 43 DEGs were common to all three time points in both duck breeds. These DEGs may thus be involved in regulating this developmental process. Specifically, KEGG pathway analysis of the 393 DEGs showed that genes involved with different metabolism pathways were highly expressed, while genes involved with cell cycle pathways showed lower expression levels at E27. These DEGs may thus be involved in the mechanisms responsible for the phenomenon of static or decreased breast muscle growth in duck breeds during the late embryonic period. These results increase the available genetic information for ducks and provide valuable resources for analyzing the mechanisms underlying the process of PM development. PMID:28771592

Deep RNA sequencing of pectoralis muscle transcriptomes during late-term embryonic to neonatal development in indigenous Chinese duck breeds.

PubMed

Zhu, Chunhong; Song, Weitao; Tao, Zhiyun; Liu, Hongxiang; Xu, Wenjuan; Zhang, Shuangjie; Li, Huifang

2017-01-01

Pectoral muscle (PM) comprises an important component of overall meat mass in ducks. However, PM has shown arrested or even reduced growth during late embryonic development, and the molecular mechanisms underlying PM growth during the late embryonic to neonatal period in ducks have not been addressed. In this study, we characterized potential candidate genes and signaling pathways related to PM development using RNA sequencing of PM samples selected at embryonic days (E) 21 and 27 and 5 days post-hatch (dph) in two duck breeds (Gaoyou and Jinding ducks). A total of 393 differentially expressed genes (DEGs) were identified, which showed higher or lower expression levels at E27 compared with E21 and 5 dph, reflecting the pattern of PM growth rates. Among these, 43 DEGs were common to all three time points in both duck breeds. These DEGs may thus be involved in regulating this developmental process. Specifically, KEGG pathway analysis of the 393 DEGs showed that genes involved with different metabolism pathways were highly expressed, while genes involved with cell cycle pathways showed lower expression levels at E27. These DEGs may thus be involved in the mechanisms responsible for the phenomenon of static or decreased breast muscle growth in duck breeds during the late embryonic period. These results increase the available genetic information for ducks and provide valuable resources for analyzing the mechanisms underlying the process of PM development.

Early events in xenograft development from the human embryonic stem cell line HS181--resemblance with an initial multiple epiblast formation.

PubMed

Gertow, Karin; Cedervall, Jessica; Jamil, Seema; Ali, Rouknuddin; Imreh, Marta P; Gulyas, Miklos; Sandstedt, Bengt; Ahrlund-Richter, Lars

2011-01-01

Xenografting is widely used for assessing in vivo pluripotency of human stem cell populations. Here, we report on early to late events in the development of mature experimental teratoma from a well-characterized human embryonic stem cell (HESC) line, HS181. The results show an embryonic process, increasingly chaotic. Active proliferation of the stem cell derived cellular progeny was detected already at day 5, and characterized by the appearance of multiple sites of engraftment, with structures of single or pseudostratified columnar epithelium surrounding small cavities. The striking histological resemblance to developing embryonic ectoderm, and the formation of epiblast-like structures was supported by the expression of the markers OCT4, NANOG, SSEA-4 and KLF4, but a lack of REX1. The early neural marker NESTIN was uniformly expressed, while markers linked to gastrulation, such as BMP-4, NODAL or BRACHYURY were not detected. Thus, observations on day 5 indicated differentiation comparable to the most early transient cell populations in human post implantation development. Confirming and expanding on previous findings from HS181 xenografts, these early events were followed by an increasingly chaotic development, incorporated in the formation of a benign teratoma with complex embryonic components. In the mature HS181 teratomas not all types of organs/tissues were detected, indicating a restricted differentiation, and a lack of adequate spatial developmental cues during the further teratoma formation. Uniquely, a kinetic alignment of rare complex structures was made to human embryos at diagnosed gestation stages, showing minor kinetic deviations between HS181 teratoma and the human counterpart.

The embryonic mir-35 family of microRNAs promotes multiple aspects of fecundity in Caenorhabditis elegans.

PubMed

McJunkin, Katherine; Ambros, Victor

2014-07-21

MicroRNAs guide many aspects of development in all metazoan species. Frequently, microRNAs are expressed during a specific developmental stage to perform a temporally defined function. The C. elegans mir-35-42 microRNAs are expressed abundantly in oocytes and early embryos and are essential for embryonic development. Here, we show that these embryonic microRNAs surprisingly also function to control the number of progeny produced by adult hermaphrodites. Using a temperature-sensitive mir-35-42 family mutant (a deletion of the mir-35-41 cluster), we demonstrate three distinct defects in hermaphrodite fecundity. At permissive temperatures, a mild sperm defect partially reduces hermaphrodite fecundity. At restrictive temperatures, somatic gonad dysfunction combined with a severe sperm defect sharply reduces fecundity. Multiple lines of evidence, including a late embryonic temperature-sensitive period, support a role for mir-35-41 early during development to promote subsequent sperm production in later larval stages. We further show that the predicted mir-35 family target sup-26 (suppressor-26) acts downstream of mir-35-41 in this process, suggesting that sup-26 de-repression in mir-35-41 deletion mutants may contribute to temperature-sensitive loss of fecundity. In addition, these microRNAs play a role in male fertility, promoting proper morphogenesis of male-specific mating structures. Overall, our results demonstrate that robust activity of the mir-35-42 family microRNAs not only is essential for embryonic development across a range of temperatures but also enables the worm to subsequently develop full reproductive capacity. Copyright © 2014 McJunkin and Ambros.

Neural Organization of the Optic Lobe Changes Steadily from Late Embryonic Stage to Adulthood in Cuttlefish Sepia pharaonis

PubMed Central

Liu, Yung-Chieh; Liu, Tsung-Han; Su, Chia-Hao; Chiao, Chuan-Chin

2017-01-01

The optic lobe is the largest structure in the cuttlefish brain. While the general morphology of the optic lobe in adult cuttlefish has been well described, the 3D structure and ontogenetic development of its neural organization have not been characterized. To correlate observed behavioral changes within the brain structure along the development of this animal, optic lobes from the late embryonic stage to adulthood were examined systematically in the present study. The MRI scan revealed that the so called “cell islands” in the medulla of the cephalopod's optic lobe (Young, 1962, 1974) are in fact a contiguous tree-like structure. Quantification of the neural organizational development of optic lobes showed that structural features of the cortex and radial column zone were established earlier than those of the tangential zone during embryonic and post-hatching stages. Within the cell islands, the density of nuclei was decreased while the size of nuclei was increased during the development. Furthermore, the visual processing area in the optic lobe showed a significant variation in lateralization during embryonic and juvenile stages. Our observation of a continuous increase in neural fibers and nucleus size in the tangential zone of the optic lobe from late embryonic stage to adulthood indicates that the neural organization of the optic lobe is modified along the development of cuttlefish. These findings thus support that the ontogenetic change of the optic lobe is responsible for their continuously increased complexity in body patterning and visuomotor behaviors. PMID:28798695

The miR-290-295 cluster as multi-faceted players in mouse embryonic stem cells.

PubMed

Yuan, Kai; Ai, Wen-Bing; Wan, Lin-Yan; Tan, Xiao; Wu, Jiang-Feng

2017-01-01

Increasing evidence indicates that embryonic stem cell specific microRNAs (miRNAs) play an essential role in the early development of embryo. Among them, the miR-290-295 cluster is the most highly expressed in the mouse embryonic stem cells and involved in various biological processes. In this paper, we reviewed the research progress of the function of the miR-290-295 cluster in embryonic stem cells. The miR-290-295 cluster is involved in regulating embryonic stem cell pluripotency maintenance, self-renewal, and reprogramming somatic cells to an embryonic stem cell-like state. Moreover, the miR-290-295 cluster has a latent pro-survival function in embryonic stem cells and involved in tumourigenesis and senescence with a great significance. Elucidating the interaction between the miR-290-295 cluster and other modes of gene regulation will provide us new ideas on the biology of pluripotent stem cells. In the near future, the broad prospects of the miRNA cluster will be shown in the stem cell field, such as altering cell identities with high efficiency through the transient introduction of tissue-specific miRNA cluster.

Density gradient electrophoresis of cultured human embryonic kidney cells

NASA Technical Reports Server (NTRS)

Plank, L. D.; Kunze, M. E.; Giranda, V.; Todd, P. W.

1985-01-01

Ground based confirmation of the electrophoretic heterogeneity of human embryonic kidney cell cultures, the general characterization of their electrophoretic migration, and observations on the general properties of cultures derived from electrophoretic subpopulations were studied. Cell migration in a density gradient electrophoresis column and cell electrophoretic mobility was determined. The mobility and heterogeneity of cultured human embryonic kidney cells with those of fixed rat erythrocytes as model test particle was compared. Electrophoretically separated cell subpopulations with respect to size, viability, and culture characteristics were examined.

Fluorescence lifetime imaging of induced pluripotent stem cells

NASA Astrophysics Data System (ADS)

Uchugonova, Aisada; Batista, Ana; König, Karsten

2014-02-01

The multiphoton FLIM tomograph MPTflex with its flexible scan head, articulated arm, and the tunable femtosecond laser source was employed to study cell monolayers and 3D cell clusters. FLIM was performed with 250 ps temporal resolution and submicron special resolution using time-correlated single photon counting. The autofluorescence based on NAD(P)H and flavins/flavoproteins has been measured in mouse embryonic fibroblasts, induced pluripotent stem cells (iPS cells) originated from mouse embryonic fibroblasts and non-proliferative mouse embryonic fibroblasts.

Isolation and characterization of the trophectoderm from the Arabian camel (Camelus dromedarius).

PubMed

Saadeldin, Islam M; Swelum, Ayman Abdel-Aziz; Elsafadi, Mona; Moumen, Abdullah F; Alzahrani, Faisal A; Mahmood, Amer; Alfayez, Musaad; Alowaimer, Abdullah N

2017-09-01

We isolated and characterized trophoblast from in vivo-derived camel embryos and compared with embryonic stem-like cells. Camel embryos were flushed on day 8 post-insemination and used to derive trophectoderm and embryonic stem-like cells under feeder-free culture conditions using a basement membrane matrix. Embryos were evaluated for the expression of POU5F1, MYC, KLF4, SOX2, CDX2, and KRT8 mRNA transcripts by relative quantitative polymerase chain reaction. Camel embryos grew and expanded to ∼4.5 mm and maintained their vesicular shape in vitro for 21 days post-insemination. Trophoblast and embryonic stem-like cell lines grew under feeder-free culture conditions and showed distinct morphological criteria and normal chromosomal counts. Embryonic stem-like cells showed positive staining in the alkaline phosphatase reaction. Trophoblast cells showed a significant increase in CDX2, KRT8, KLF4, and SOX2 expression compared with embryonic stem-like cells and whole embryos. Embryonic stem-like cells showed a significant decrease in CDX2 expression and increase in SOX2 and KRT8 expression compared to embryonic expression. POU5F1 and MYC expression showed no difference between embryos and both cell lines. We characterized embryo survival in vitro, particularly the derivation of trophectoderm and embryonic stem-like cells, providing a foundation for further analysis of early embryonic development and placentation in camels. Copyright © 2017 Elsevier Ltd. All rights reserved.

Unique differentiation profile of mouse embryonic stem cells in rotary and stirred tank bioreactors.

PubMed

Fridley, Krista M; Fernandez, Irina; Li, Mon-Tzu Alice; Kettlewell, Robert B; Roy, Krishnendu

2010-11-01

Embryonic stem (ES)-cell-derived lineage-specific stem cells, for example, hematopoietic stem cells, could provide a potentially unlimited source for transplantable cells, especially for cell-based therapies. However, reproducible methods must be developed to maximize and scale-up ES cell differentiation to produce clinically relevant numbers of therapeutic cells. Bioreactor-based dynamic culture conditions are amenable to large-scale cell production, but few studies have evaluated how various bioreactor types and culture parameters influence ES cell differentiation, especially hematopoiesis. Our results indicate that cell seeding density and bioreactor speed significantly affect embryoid body formation and subsequent generation of hematopoietic stem and progenitor cells in both stirred tank (spinner flask) and rotary microgravity (Synthecon™) type bioreactors. In general, high percentages of hematopoietic stem and progenitor cells were generated in both bioreactors, especially at high cell densities. In addition, Synthecon bioreactors produced more sca-1(+) progenitors and spinner flasks generated more c-Kit(+) progenitors, demonstrating their unique differentiation profiles. cDNA microarray analysis of genes involved in pluripotency, germ layer formation, and hematopoietic differentiation showed that on day 7 of differentiation, embryoid bodies from both bioreactors consisted of all three germ layers of embryonic development. However, unique gene expression profiles were observed in the two bioreactors; for example, expression of specific hematopoietic genes were significantly more upregulated in the Synthecon cultures than in spinner flasks. We conclude that bioreactor type and culture parameters can be used to control ES cell differentiation, enhance unique progenitor cell populations, and provide means for large-scale production of transplantable therapeutic cells.

ACTIONS OF THE ENDOCRINE DISRUPTOR METHOXYCHLOR AND ITS ESTROGENIC METABOLITE ON IN VITRO EMBRYONIC RAT SEMINIFEROUS CORD FORMATION AND PERINATAL TESTIS GROWTH. (R827405)

EPA Science Inventory

Abstract

The current study examines the actions of methoxychlor and its estrogenic metabolite, 2, 2-bis-(p-hydroxyphenyl)-1, 1, 1-trichloroethane (HPTE), on seminiferous cord formation and growth of the developing rat testis. The developing testis in the embryonic and ...

Generation of the Dimensional Embryology Application (App) for Visualization of Early Chick and Frog Embryonic Development

ERIC Educational Resources Information Center

Webb, Rebecca L.; Bilitski, James; Zerbee, Alyssa; Symans, Alexandra; Chop, Alexandra; Seitz, Brianne; Tran, Cindy

2015-01-01

The study of embryonic development of multiple organisms, including model organisms such as frogs and chicks, is included in many undergraduate biology programs, as well as in a variety of graduate programs. As our knowledge of biological systems increases and the amount of material to be taught expands, the time spent instructing students about…

Developmental origin of limb size variation in lizards.

PubMed

Andrews, Robin M; Skewes, Sable A

2017-05-01

In many respects, reptile hatchlings are fully functional, albeit miniature, adults. This means that the adult morphology must emerge during embryonic development. This insight emphasizes the connection between the mechanisms that generate phenotypic variation during embryonic development and the action of selection on post-hatching individuals. To determine when species-specific differences in limb and tail lengths emerge during embryonic development, we compared allometric patterns of early limb growth of four distantly related species of lizards. The major questions addressed were whether early embryonic limb and tail growth is characterized by the gradual (continuous allometry) or by the abrupt emergence (transpositional allometry) of size differences among species. Our observations supported transpositional allometry of both limbs and tails. Species-specific differences in limb and tail length were exhibited when limb and tail buds first protruded from the body wall. Genes known to be associated with early limb development of tetrapods are obvious targets for studies on the genetic mechanisms that determine interspecific differences in relative limb length. Broadly comparative studies of gene regulation would facilitate understanding of the mechanisms underlying adaptive variation in limb size, including limb reduction and loss, of squamate reptiles. © 2017 Wiley Periodicals, Inc.

Evidence of increased endometrial vascular permeability at the time of implantation in the short-nosed fruit bat, Cyanopterus sphinx.

PubMed

Pakrasi, Pranab Lal; Tiwari, Anjana

2007-09-01

Early embryonic development and implantation were studied in tropical short-nosed fruit bat Cyanopterus sphinx. We report preimplantation development and embryo implantation. Different stages of cleavage were observed in embryo by direct microscopic examination of fresh embryos after retrieving them either from the oviduct or the uterus at different days, up to the day of implantation. Generally, the embryos enter the uterus at the 8-cell stage. Embryonic development continued without any delay and blastocyst were formed showing attachment to the uterine epithelium at the mesometrial side of the uterus. A distinct blue band was formed in the uterus. The site of blastocyst attachment was visualized as a blue band following intravenous injection of pontamine blue. Implantation occurred 9+/-0.7 days after mating. This study reports that bat embryonic development can be studied like other laboratory animals and that this bat shows blue dye reaction, indicating the site and exact time of implantation. This blue dye reaction can be used to accurately find post-implantational delay. We prove conclusively that this species of tropical bat does not have any type of embryonic diapause.

TORC2 signaling antagonizes SKN-1 to induce C. elegans mesendodermal embryonic development

PubMed Central

Ruf, Vanessa; Holzem, Christina; Peyman, Tobias; Walz, Gerd; Blackwell, T. Keith; Neumann-Haefelin, Elke

2013-01-01

The evolutionarily conserved target of rapamycin (TOR) kinase controls fundamental metabolic processes to support cell and tissue growth. TOR functions within the context of two distinct complexes, TORC1 and TORC2. TORC2, with its specific component Rictor, has been recently implicated in aging and regulation of growth and metabolism. Here, we identify rict-1/Rictor as a regulator of embryonic development in C. elegans. The transcription factor skn-1 establishes development of the mesendoderm in embryos, and is required for cellular homeostasis and longevity in adults. Loss of maternal skn-1 function leads to misspecification of the mesendodermal precursor and failure to form intestine and pharynx. We found that genetic inactivation of rict-1 suppressed skn-1-associated lethality by restoring mesendodermal specification in skn-1 deficient embryos. Inactivation of other TORC2 but not TORC1 components also partially rescued skn-1 embryonic lethality. The SGK-1 kinase mediated these functions downstream of rict-1/TORC2, as a sgk-1 gain-of-function mutant suppressed the rict-1 mutant phenotype. These data indicate that TORC2 and SGK-1 antagonize SKN-1 during embryonic development. PMID:23973804

NG2 glia are required for vessel network formation during embryonic development

PubMed Central

Minocha, Shilpi; Valloton, Delphine; Brunet, Isabelle; Eichmann, Anne

2015-01-01

The NG2+ glia, also known as polydendrocytes or oligodendrocyte precursor cells, represent a new entity among glial cell populations in the central nervous system. However, the complete repertoire of their roles is not yet identified. The embryonic NG2+ glia originate from the Nkx2.1+ progenitors of the ventral telencephalon. Our analysis unravels that, beginning from E12.5 until E16.5, the NG2+ glia populate the entire dorsal telencephalon. Interestingly, their appearance temporally coincides with the establishment of blood vessel network in the embryonic brain. NG2+ glia are closely apposed to developing cerebral vessels by being either positioned at the sprouting tip cells or tethered along the vessel walls. Absence of NG2+ glia drastically affects the vascular development leading to severe reduction of ramifications and connections by E18.5. By revealing a novel and fundamental role for NG2+ glia, our study brings new perspectives to mechanisms underlying proper vessels network formation in embryonic brains. DOI: http://dx.doi.org/10.7554/eLife.09102.001 PMID:26651999

Oocyte exposure to ZnO nanoparticles inhibits early embryonic development through the γ-H2AX and NF-κB signaling pathways.

PubMed

Liu, Jing; Zhao, Yong; Ge, Wei; Zhang, Pengfei; Liu, Xinqi; Zhang, Weidong; Hao, Yanan; Yu, Shuai; Li, Lan; Chu, Meiqiang; Min, Lingjiang; Zhang, Hongfu; Shen, Wei

2017-06-27

The impacts of zinc oxide nanoparticles on embryonic development following oocyte stage exposure are unknown and the underlying mechanisms are sparsely understood. In the current investigation, intact nanoparticles were detected in ovarian tissue in vivo and cultured cells in vitro under zinc oxide nanoparticles treatment. Zinc oxide nanoparticles exposure during the oocyte stage inhibited embryonic development. Notably, in vitro culture data closely matched in vivo embryonic data, in that the impairments caused by Zinc oxide nanoparticles treatment passed through cell generations; and both gamma-H2AX and NF-kappaB pathways were involved in zinc oxide nanoparticles caused embryo-toxicity. Copper oxide and silicon dioxide nanoparticles have been used to confirm that particles are important for the toxicity of zinc oxide nanoparticles. The toxic effects of zinc oxide nanoparticles emanate from both intact nanoparticles and Zn2+. Our investigation along with others suggests that zinc oxide nanoparticles are toxic to the female reproductive system [ovaries (oocytes)] and subsequently embryo-toxic and that precaution should be taken regarding human exposure to their everyday use.

In vitro developmental model of the gastrointestinal tract from mouse embryonic stem cells.

PubMed

Torihashi, Shigeko; Kuwahara, Masaki; Kurahashi, Masaaki

2007-10-01

Mouse embryonic stem (ES) cells are pluripotent and retain their potential to form cells, tissues and organs originated from three embryonic germ layers. Recently, we developed in vitro organ--gut-like structures--from mouse ES cells. They had basically similar morphological features to a mouse gastrointestinal tract in vivo composed of three distinct layers (i.e., epithelium, connective tissue and musculature). Gut-like structures showed spontaneous contractions derived from pacemaker cells (interstitial cells of Cajal) in the musculature. We also examined their formation process and expression pattern of transcription factors crucial for gut organogenesis such as Id2, Sox17, HNF3beta/Foxa2 and GATA4. We found that they mimic the development of embryonic gut in vivo and showed a similar expression pattern of common transcription factors. They also maintain their developmental potential after transplantation to a renal capsule. Therefore, gut-like structures are suitable for in vitro models of gastrointestinal tracts and their development. In addition, we pointed out several unique features different from gut in vivo that provide useful and advantageous tools to investigate the developmental mechanism of the gastrointestinal tract.

Hematopoietic cell differentiation from embryonic and induced pluripotent stem cells

PubMed Central

2013-01-01

Pluripotent stem cells, both embryonic stem cells and induced pluripotent stem cells, are undifferentiated cells that can self-renew and potentially differentiate into all hematopoietic lineages, such as hematopoietic stem cells (HSCs), hematopoietic progenitor cells and mature hematopoietic cells in the presence of a suitable culture system. Establishment of pluripotent stem cells provides a comprehensive model to study early hematopoietic development and has emerged as a powerful research tool to explore regenerative medicine. Nowadays, HSC transplantation and hematopoietic cell transfusion have successfully cured some patients, especially in malignant hematological diseases. Owing to a shortage of donors and a limited number of the cells, hematopoietic cell induction from pluripotent stem cells has been regarded as an alternative source of HSCs and mature hematopoietic cells for intended therapeutic purposes. Pluripotent stem cells are therefore extensively utilized to facilitate better understanding in hematopoietic development by recapitulating embryonic development in vivo, in which efficient strategies can be easily designed and deployed for the generation of hematopoietic lineages in vitro. We hereby review the current progress of hematopoietic cell induction from embryonic stem/induced pluripotent stem cells. PMID:23796405

Oocyte exposure to ZnO nanoparticles inhibits early embryonic development through the γ-H2AX and NF-κB signaling pathways

PubMed Central

Liu, Jing; Zhao, Yong; Ge, Wei; Zhang, Pengfei; Liu, Xinqi; Zhang, Weidong; Hao, Yanan; Yu, Shuai; Li, Lan; Chu, Meiqiang; Min, Lingjiang; Zhang, Hongfu; Shen, Wei

2017-01-01

The impacts of zinc oxide nanoparticles on embryonic development following oocyte stage exposure are unknown and the underlying mechanisms are sparsely understood. In the current investigation, intact nanoparticles were detected in ovarian tissue in vivo and cultured cells in vitro under zinc oxide nanoparticles treatment. Zinc oxide nanoparticles exposure during the oocyte stage inhibited embryonic development. Notably, in vitro culture data closely matched in vivo embryonic data, in that the impairments caused by Zinc oxide nanoparticles treatment passed through cell generations; and both gamma-H2AX and NF-kappaB pathways were involved in zinc oxide nanoparticles caused embryo-toxicity. Copper oxide and silicon dioxide nanoparticles have been used to confirm that particles are important for the toxicity of zinc oxide nanoparticles. The toxic effects of zinc oxide nanoparticles emanate from both intact nanoparticles and Zn2+. Our investigation along with others suggests that zinc oxide nanoparticles are toxic to the female reproductive system [ovaries (oocytes)] and subsequently embryo-toxic and that precaution should be taken regarding human exposure to their everyday use. PMID:28487501

Metabolic circadian rhythms in embryonic turtles.

PubMed

Loudon, Fiona Kay; Spencer, Ricky-John; Strassmeyer, Alana; Harland, Karen

2013-07-01

Oviparous species are model organisms for investigating embryonic development of endogenous physiological circadian rhythms without the influence of maternal biorhythms. Recent studies have demonstrated that heart rates and metabolic rates of embryonic turtles are not constant or always maximal and can be altered in response to the presence of embryos at a more advanced stage of development within the nest. A first step in understanding the physiological mechanisms underpinning these responses in embryonic ectothermic organisms is to develop metabolic profiles (e.g., heart rate) at different temperatures throughout incubation. Heart beat and rhythmic patterns or changes in development may represent important signals or cues within a nest and may be vital to coordinate synchronous hatching well in advance of the final stages of incubation. We developed baseline embryonic heart-rate profiles of embryos of the short-necked Murray River turtle (Emydura macquarii) to determine the stage of embryogenesis that metabolic circadian rhythms become established, if at all. Eggs were incubated at constant temperatures (26°C and 30°C) and heart rates were monitored at 6-h intervals over 24 h every 7-11 days until hatching. Circadian heart rate rhythms were detected at the mid-gestation period and were maintained until hatching. Heart rates throughout the day varied by up to 20% over 24 h and were not related to time of day. This study demonstrated that endogenous metabolic circadian rhythms in developing embryos in turtle eggs establish earlier in embryogenesis than those documented in other vertebrate taxa during embryogenesis. Early establishment of circadian rhythms in heart rates may be critical for communication among embryos and synchrony in hatching and emergence from the nest.

Qualitative research of alternatively splice variants of fibronectin in different development stage of mice heart.

PubMed

Lu, Feng; Ma, Fang-Fang; Zhang, Wei; Li, Ying; Wei, Fei-Yu; Zhou, Lei

2015-12-01

Fibronectin (FN) plays vital roles in cell adhesion, differentiation, proliferation and migration. It is involved in the process of embryonic development and is highly conserved during evolution. The EIIIA and EIIIB of FN show a very high degree of homology among vertebrates. Embryos deleting both EIIIA and EIIIB displayed multiple embryonic cardiovascular defects, implying their crucial role during embryogenesis. The correlation of spliced EIIIB, EIIIA, and IIICS of FN to heart development was studied by observing their chronological expression in mice heart. C57 mice embryos at E11.5, E12.5, E13.5, E14.5, E15.5, E16.5, E17.5, E18.5, E19.5 days, postnatal day 1 (P1d), and adult male mice (3 months) were used. For each alternatively spliced FN1 domain (EIIIB, EIIIA and IIICS), primer pairs were designed for specific amplification. Total RNA was extracted from the heart tissue, reverse transcripted to cDNA, followed by RT-PCR with specific primers. The PCR amplification was verified by agarose gel electrophoresis, showing specific fragments of the expected sizes. In adult mice heart, only alternatively splice variants of EIIIA-, EIIIB-, IIICS+ were expressed. While in embryonic mice, spliced variant of EIIIA+/-, EIIIB+/-, IIICS+ were observed. The expression of EIIIA and EIIIB changed during heart development. FN is crucial for the normal development of the embryonic heart by modulating cardiac neural crest (CNC) proliferation and survival, and maintenance of CNC cells. FN1 gene seems to play a significant role by expression of highly conserved EIIIA and EIIIB in embryonic heart development.

Longitudinal Effects of Embryonic Exposure to Cocaine on Morphology, Cardiovascular Physiology, and Behavior in Zebrafish.

PubMed

Mersereau, Eric J; Boyle, Cody A; Poitra, Shelby; Espinoza, Ana; Seiler, Joclyn; Longie, Robert; Delvo, Lisa; Szarkowski, Megan; Maliske, Joshua; Chalmers, Sarah; Darland, Diane C; Darland, Tristan

2016-05-31

A sizeable portion of the societal drain from cocaine abuse results from the complications of in utero drug exposure. Because of challenges in using humans and mammalian model organisms as test subjects, much debate remains about the impact of in utero cocaine exposure. Zebrafish offer a number of advantages as a model in longitudinal toxicology studies and are quite sensitive physiologically and behaviorally to cocaine. In this study, we have used zebrafish to model the effects of embryonic pre-exposure to cocaine on development and on subsequent cardiovascular physiology and cocaine-induced conditioned place preference (CPP) in longitudinal adults. Larval fish showed a progressive decrease in telencephalic size with increased doses of cocaine. These treated larvae also showed a dose dependent response in heart rate that persisted 24 h after drug cessation. Embryonic cocaine exposure had little effect on overall health of longitudinal adults, but subtle changes in cardiovascular physiology were seen including decreased sensitivity to isoproterenol and increased sensitivity to cocaine. These longitudinal adult fish also showed an embryonic dose-dependent change in CPP behavior, suggesting an increased sensitivity. These studies clearly show that pre-exposure during embryonic development affects subsequent cocaine sensitivity in longitudinal adults.

[Regulation of in vitro and in vivo differentiation of mouse embryonic stem cells, embryonic germ cells, and teratocarcinoma cells by TGFb family signaling factors].

PubMed

Gordeeva, O F; Nikonova, T M; Lifantseva, N V

2009-01-01

The activity of specific signaling and transcription factors determines the cell fate in normal development and in tumor transformation. The transcriptional profiles of gene-components of different branches of TGFbeta family signaling pathways were studied in experimental models of initial stages of three-dimensional in vitro differentiation of embryonic stem cells, embryonic germ cells and teratocarcinoma cells and in teratomas and teratocarcinomas developed after their transplantation into immunodeficient Nude mice. Gene profile analysis of studied cell systems have revealed that expression patterns of ActivinA, Nodal, Lefty1, Lefty2, TGF TGFbeta1, BMP4, and GDF were identical in pluripotent stem cells whereas the mRNAs of all examined genes with the exception of Inhibin betaA/ActivinA were detected in the teratocarcinoma cells. These results indicate that differential activity of signaling pathways of the TGFbeta family factors regulates pluripotent state maintenance and pluripotent stem cell differentiation into the progenitors of three germ layers and extraembryonic structures and that normal expression pattern of TGFbeta family factors is rearranged in embryonic teratocarcinoma cells during tumor growth in vitro and in vivo.

Spatial distribution of endogenous retinoids in the murine embryonic mandible.

PubMed

Kronmiller, J E; Beeman, C S

1994-12-01

Retinoids play an important part in pattern formation during embryonic development. Exogenous retinoids alter the pattern of skeletal, neural and odontogenic tissues. Endogenous retinoids have been demonstrated previously in the murine embryonic mandible, reaching a concentration peak during the initiation of odontogenesis. It was now found that endogenous retinoids are present in a concentration gradient in the embryonic mouse mandible at the time of the initiation of the dental lamina. All-trans-retinoic acid was more concentrated in the incisor region and retinol in the molar region. These results, and the fact that exogenous retinoids produce supernumerary incisors and missing molars, suggest that all-trans-retinoic acid may instruct incisor morphology.

Embryonic Lethality of Mitochondrial Pyruvate Carrier 1 Deficient Mouse Can Be Rescued by a Ketogenic Diet

PubMed Central

Krznar, Petra; Hörl, Manuel; Ammar, Zeinab; Montessuit, Sylvie; Pierredon, Sandra; Zamboni, Nicola; Martinou, Jean-Claude

2016-01-01

Mitochondrial import of pyruvate by the mitochondrial pyruvate carrier (MPC) is a central step which links cytosolic and mitochondrial intermediary metabolism. To investigate the role of the MPC in mammalian physiology and development, we generated a mouse strain with complete loss of MPC1 expression. This resulted in embryonic lethality at around E13.5. Mouse embryonic fibroblasts (MEFs) derived from mutant mice displayed defective pyruvate-driven respiration as well as perturbed metabolic profiles, and both defects could be restored by reexpression of MPC1. Labeling experiments using 13C-labeled glucose and glutamine demonstrated that MPC deficiency causes increased glutaminolysis and reduced contribution of glucose-derived pyruvate to the TCA cycle. Morphological defects were observed in mutant embryonic brains, together with major alterations of their metabolome including lactic acidosis, diminished TCA cycle intermediates, energy deficit and a perturbed balance of neurotransmitters. Strikingly, these changes were reversed when the pregnant dams were fed a ketogenic diet, which provides acetyl-CoA directly to the TCA cycle and bypasses the need for a functional MPC. This allowed the normal gestation and development of MPC deficient pups, even though they all died within a few minutes post-delivery. This study establishes the MPC as a key player in regulating the metabolic state necessary for embryonic development, neurotransmitter balance and post-natal survival. PMID:27176894

Magnetic resonance imaging study of eye congenital birth defects in mouse model

PubMed Central

Tucker, Zachary; Mongan, Maureen; Meng, Qinghang; Xia, Ying

2017-01-01

Purpose Embryonic eyelid closure is a well-documented morphogenetic episode in mammalian eye development. Detection of eyelid closure defect in humans is a major challenge because eyelid closure and reopen occur entirely in utero. As a consequence, congenital eye defects that are associated with failure of embryonic eyelid closure remain unknown. To fill the gap, we developed a mouse model of defective eyelid closure. This preliminary work demonstrates that the magnetic resonance imaging (MRI) approach can be used for the detection of extraocular muscle abnormalities in the mouse model. Methods Mice with either normal (Map3k1+/−) or defective (Map3k1−/−) embryonic eyelid closure were used in this study. Images of the extraocular muscles were obtained with a 9.4 T high resolution microimaging MRI system. The extraocular muscles were identified, segmented, and measured in each imaging slice using an in-house program. Results In agreement with histological findings, the imaging data show that mice with defective embryonic eyelid closure develop less extraocular muscle than normal mice. In addition, the size of the eyeballs was noticeably reduced in mice with defective embryonic eyelid closure. Conclusions We demonstrated that MRI can potentially be used for the study of extraocular muscle in the mouse model of the eye open-at-birth defect, despite the lack of specificity of muscle group provided by the current imaging resolution. PMID:28848319

Effect of tidal overwash on the embryonic development of leatherback turtles in French Guiana.

PubMed

Caut, Stéphane; Guirlet, Elodie; Girondot, Marc

2010-05-01

In marine turtles, the physical conditions experienced by eggs during incubation affect embryonic development. In the leatherback, hatching success is known to be low in relation to other marine turtles as a result of high embryonic mortality. Moreover, the hatching success on Yalimapo in French Guiana, one major nesting beach for this species, is lower compared to other nesting sites. We assessed the rate of leatherback turtle embryonic mortality in order to investigate the tolerance of leatherback turtle clutches laid on Yalimapo beach to tidal overwash, and we highlight causes of poor hatching success. Of the 89 nests studied, 27 were overlapped by tide at least once during the incubation period (of which five nests were lost by erosion). The hatching success was on average significantly lower in overwashed nests than in non-overwashed, highlighting the existence of embryonic developmental arrest linked to tidal inundation. The stages of developmental arrest and their proportion are linked with time, frequency and level of overwash events. In the context of global warming and associated sea-level rise, understanding the detrimental effect of tidal inundation on the development of marine turtle nests is of interest in nesting sites where turtles are likely to be forced to nest closer to the tide line, thus exposing their nests to greater risk of nest overlap with sea and tidal inundation. Copyright 2009 Elsevier Ltd. All rights reserved.

Determining the impact of cell mixing on signaling during development.

PubMed

Uriu, Koichiro; Morelli, Luis G

2017-06-01

Cell movement and intercellular signaling occur simultaneously to organize morphogenesis during embryonic development. Cell movement can cause relative positional changes between neighboring cells. When intercellular signals are local such cell mixing may affect signaling, changing the flow of information in developing tissues. Little is known about the effect of cell mixing on intercellular signaling in collective cellular behaviors and methods to quantify its impact are lacking. Here we discuss how to determine the impact of cell mixing on cell signaling drawing an example from vertebrate embryogenesis: the segmentation clock, a collective rhythm of interacting genetic oscillators. We argue that comparing cell mixing and signaling timescales is key to determining the influence of mixing. A signaling timescale can be estimated by combining theoretical models with cell signaling perturbation experiments. A mixing timescale can be obtained by analysis of cell trajectories from live imaging. After comparing cell movement analyses in different experimental settings, we highlight challenges in quantifying cell mixing from embryonic timelapse experiments, especially a reference frame problem due to embryonic motions and shape changes. We propose statistical observables characterizing cell mixing that do not depend on the choice of reference frames. Finally, we consider situations in which both cell mixing and signaling involve multiple timescales, precluding a direct comparison between single characteristic timescales. In such situations, physical models based on observables of cell mixing and signaling can simulate the flow of information in tissues and reveal the impact of observed cell mixing on signaling. © 2017 Japanese Society of Developmental Biologists.

The histone demethylase Fbxl11/Kdm2a plays an essential role in embryonic development by repressing cell-cycle regulators.

PubMed

Kawakami, Eri; Tokunaga, Akinori; Ozawa, Manabu; Sakamoto, Reiko; Yoshida, Nobuaki

2015-02-01

Methylation and de-methylation of histone lysine residues play pivotal roles in mammalian early development; these modifications influence chromatin architecture and regulate gene transcription. Fbxl11 (F-box and leucine-rich repeat 11)/Kdm2a is a histone demethylase that selectively removes mono- and di-methylation from histone H3K36. Previously, two other histone H3K36 demethylases (Jmjd5 or Fbxl10) were analyzed based on the phenotypes of the corresponding knockout (KO) mice; the results of those studies implicated H3K36 demethylases in cell proliferation, apoptosis, and senescence (Fukuda et al., 2011; Ishimura et al., 2012). To elucidate the physiological role of Fbxl11, we generated and examined Fbxl11 KO mice. Fbxl11 was expressed throughout the body during embryogenesis, and the Fbxl11 KO mice exhibited embryonic lethality at E10.5-12.5, accompanied with severe growth defects leading to reduced body size. Furthermore, knockout of Fbxl11 decreased cell proliferation and increased apoptosis. The lack of Fbxl11 resulted in downregulation of the Polycomb group protein (PcG) Ezh2, PcG mediated H2A ubiquitination and upregulation of the cyclin-dependent kinase inhibitor p21Cip1. Taken together, our findings suggest that Fbxl11 plays an essential role in embryonic development and homeostasis by regulating cell proliferation and survival. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Preserving and Using Germplasm and Dissociated Embryonic Cells for Conserving Caribbean and Pacific Coral

PubMed Central

Hagedorn, Mary; Carter, Virginia; Martorana, Kelly; Paresa, Malia K.; Acker, Jason; Baums, Iliana B.; Borneman, Eric; Brittsan, Michael; Byers, Michael; Henley, Michael; Laterveer, Michael; Leong, Jo-Ann; McCarthy, Megan; Meyers, Stuart; Nelson, Brian D.; Petersen, Dirk; Tiersch, Terrence; Uribe, Rafael Cuevas; Woods, Erik; Wildt, David

2012-01-01

Coral reefs are experiencing unprecedented degradation due to human activities, and protecting specific reef habitats may not stop this decline, because the most serious threats are global (i.e., climate change), not local. However, ex situ preservation practices can provide safeguards for coral reef conservation. Specifically, modern advances in cryobiology and genome banking could secure existing species and genetic diversity until genotypes can be introduced into rehabilitated habitats. We assessed the feasibility of recovering viable sperm and embryonic cells post-thaw from two coral species, Acropora palmata and Fungia scutaria that have diffferent evolutionary histories, ecological niches and reproductive strategies. In vitro fertilization (IVF) of conspecific eggs using fresh (control) spermatozoa revealed high levels of fertilization (>90% in A. palmata; >84% in F. scutaria; P>0.05) that were unaffected by tested sperm concentrations. A solution of 10% dimethyl sulfoxide (DMSO) at cooling rates of 20 to 30°C/min most successfully cryopreserved both A. palmata and F. scutaria spermatozoa and allowed producing developing larvae in vitro. IVF success under these conditions was 65% in A. palmata and 53% in F. scutaria on particular nights; however, on subsequent nights, the same process resulted in little or no IVF success. Thus, the window for optimal freezing of high quality spermatozoa was short (∼5 h for one night each spawning cycle). Additionally, cryopreserved F. scutaria embryonic cells had∼50% post-thaw viability as measured by intact membranes. Thus, despite some differences between species, coral spermatozoa and embryonic cells are viable after low temperature (−196°C) storage, preservation and thawing. Based on these results, we have begun systematically banking coral spermatozoa and embryonic cells on a large-scale as a support approach for preserving existing bio- and genetic diversity found in reef systems. PMID:22413020

Adult Human Gingival Epithelial Cells as a Source for Whole-tooth Bioengineering

PubMed Central

Angelova Volponi, A.; Kawasaki, M.; Sharpe, P.T.

2013-01-01

Teeth develop from interactions between embryonic oral epithelium and neural-crest-derived mesenchyme. These cells can be separated into single-cell populations and recombined to form normal teeth, providing a basis for bioengineering new teeth if suitable, non-embryonic cell sources can be identified. We show here that cells can be isolated from adult human gingival tissue that can be expanded in vitro and, when combined with mouse embryonic tooth mesenchyme cells, form teeth. Teeth with developing roots can be produced from this cell combination following transplantation into renal capsules. These bioengineered teeth contain dentin and enamel with ameloblast-like cells and rests of Malassez of human origin. PMID:23458883

Embryonic Heart Progenitors and Cardiogenesis

PubMed Central

Brade, Thomas; Pane, Luna S.; Moretti, Alessandra; Chien, Kenneth R.; Laugwitz, Karl-Ludwig

2013-01-01

The mammalian heart is a highly specialized organ, comprised of many different cell types arising from distinct embryonic progenitor populations during cardiogenesis. Three precursor populations have been identified to contribute to different myocytic and nonmyocytic cell lineages of the heart: cardiogenic mesoderm cells (CMC), the proepicardium (PE), and cardiac neural crest cells (CNCCs). This review will focus on molecular cues necessary for proper induction, expansion, and lineage-specific differentiation of these progenitor populations during cardiac development in vivo. Moreover, we will briefly discuss how the knowledge gained on embryonic heart progenitor biology can be used to develop novel therapeutic strategies for the management of congenital heart disease as well as for improvement of cardiac function in ischemic heart disease. PMID:24086063

EMBRYONIC VASCULAR DISRUPTION ADVERSE OUTCOMES: LINKING HIGH THROUGHPUT SIGNALING SIGNATURES WITH FUNCTIONAL CONSEQUENCES

EPA Science Inventory

Embryonic vascular disruption is an important adverse outcome pathway (AOP) given the knowledge that chemical disruption of early cardiovascular system development leads to broad prenatal defects. High throughput screening (HTS) assays provide potential building blocks for AOP d...

EMBRYONIC PALATAL RESPONSES TO TERATOGENS IN SERUM-FREE ORGAN CULTURE

EPA Science Inventory

This study examines development of rat, mouse and human embryonic palates in submerged, serum-free organ culture. he concentration-response profiles for retinoic acid (RA), triamcinolone (TRI), hydrocortisone (HC), dexamethasone (DEX), and 2,3,7,11- tetrachlorodibenzo-p-dioxin (T...

Cell counting in whole mount tissue volumes using expansion OCT (Conference Presentation)

NASA Astrophysics Data System (ADS)

Liu, Yehe; Gu, Shi; Watanabe, Michiko; Rollins, Andrew M.; Jenkins, Michael W.

2017-02-01

Abnormal cell proliferation and migration during heart development can lead to severe congenital heart defects (CHDs). Studying the spatial distribution of cells during embryonic development helps our understanding of how the heart develops and the etiology of certain CHDs. However, imaging large groups of single cells in intact tissue volumes is challenging. No current technique can accomplish this task in both a time-efficient and cost-effective manner. OCT has potential with its large field of view and micron-scale resolution, but even the highest resolution OCT systems have poor contrast for counting cells and have a small field of view compared to conventional OCT. We propose using a conventional OCT system and processing the sample to enhance cellular contrast. Inspired by the recently developed Expansion Microscopy, we permeated whole-mount embryonic tissue with a superabsorbent monomer solution and polymerized into a hydrogel. When hydrated in DI water, the tissue-hydrogel complex was uniformly enlarged ( 5X in all dimensions) without distorting the microscopic structure. This had a twofold effect: it increased the resolution by a factor of 5 and decreased scattering, which allowed us to resolve cellular level features deep in the tissue with high contrast using conventional OCT. We noted that cell nuclei caused significantly more backscattering than the other subcellular structures after expansion. Based on this property, we were able to distinguish individual cell nuclei, and thus count cells, in expanded OCT images with simple intensity thresholding. We demonstrate the technique with embryonic quail hearts at various developmental stages.

Pubertal development and primary ovarian insufficiency in female survivors of embryonal brain tumors following risk-adapted craniospinal irradiation and adjuvant chemotherapy.

PubMed

DeWire, Mariko; Green, Daniel M; Sklar, Charles A; Merchant, Thomas E; Wallace, Dana; Lin, Tong; Vern-Gross, Tamara; Kun, Larry E; Krasin, Matthew J; Boyett, James M; Wright, Karen D; Wetmore, Cynthia; Broniscer, Alberto; Gajjar, Amar

2015-02-01

Female survivors of central nervous system (CNS) tumors are at an increased risk for gonadal damage and variations in the timing of puberty following radiotherapy and alkylating agent-based chemotherapy. Clinical and laboratory data were obtained from 30 evaluable female patients with newly diagnosed embryonal CNS tumors treated on a prospective protocol (SJMB 96) at St. Jude Children's Research Hospital (SJCRH). Pubertal development was evaluated by Tanner staging. Primary ovarian insufficiency (POI) was determined by Tanner staging and FSH level. Females with Tanner stage I-II and FSH > 15 mIU/ml, or Tanner stage III-V, FSH > 25 mIU/ml and FSH greater than LH were defined to have ovarian insufficiency. Recovery of ovarian function was defined as normalization of FSH without therapeutic intervention. Median length of follow-up post completion of therapy was 7.2 years (4.0-10.8 years). The cumulative incidence of pubertal onset was 75.6% by the age of 13. Precocious puberty was observed in 11.1% and delayed puberty in 11.8%. The cumulative incidence of POI was 82.8%, though recovery was observed in 38.5%. Treatment for primary CNS embryonal tumors may cause variations in the timing of pubertal development, impacting physical and psychosocial development. Female survivors are at risk for POI, a subset of whom will recover function over time. Further refinement of therapies is needed in order to reduce late ovarian insufficiency. Pediatr Blood Cancer 2015;62:329-334. © 2014 Wiley Periodicals, Inc. © 2014 Wiley Periodicals, Inc.

Precise chronology of differentiation of developing human primary dentition.

PubMed

Hu, Xuefeng; Xu, Shan; Lin, Chensheng; Zhang, Lishan; Chen, YiPing; Zhang, Yanding

2014-02-01

While correlation of developmental stage with embryonic age of the human primary dentition has been well documented, the available information regarding the differentiation timing of the primary teeth was largely based on the observation of initial mineralization and varies significantly. In this study, we aimed to document precise differentiation timing of the developing human primary dentition. We systematically examined the expression of odontogenic differentiation markers along with the formation of mineralized tissue in each developing maxillary and mandibular teeth from human embryos with well-defined embryonic age. We show that, despite that all primary teeth initiate development at the same time, odontogenic differentiation begins in the maxillary incisors at the 15th week and in the mandibular incisors at the 16th week of gestation, followed by the canine, the first primary premolar, and the second primary premolar at a week interval sequentially. Despite that the mandibular primary incisors erupt earlier than the maxillary incisors, this distal to proximal sequential differentiation of the human primary dentition coincides in general with the sequence of tooth eruption. Our results provide an accurate chronology of odontogenic differentiation of the developing human primary dentition, which could be used as reference for future studies of human tooth development.

Skull Development, Ossification Pattern, and Adult Shape in the Emerging Lizard Model Organism Pogona vitticeps: A Comparative Analysis With Other Squamates.

PubMed

Ollonen, Joni; Da Silva, Filipe O; Mahlow, Kristin; Di-Poï, Nicolas

2018-01-01

The rise of the Evo-Devo field and the development of multidisciplinary research tools at various levels of biological organization have led to a growing interest in researching for new non-model organisms. Squamates (lizards and snakes) are particularly important for understanding fundamental questions about the evolution of vertebrates because of their high diversity and evolutionary innovations and adaptations that portrait a striking body plan change that reached its extreme in snakes. Yet, little is known about the intricate connection between phenotype and genotype in squamates, partly due to limited developmental knowledge and incomplete characterization of embryonic development. Surprisingly, squamate models have received limited attention in comparative developmental studies, and only a few species examined so far can be considered as representative and appropriate model organism for mechanistic Evo-Devo studies. Fortunately, the agamid lizard Pogona vitticeps (central bearded dragon) is one of the most popular, domesticated reptile species with both a well-established history in captivity and key advantages for research, thus forming an ideal laboratory model system and justifying his recent use in reptile biology research. We first report here the complete post-oviposition embryonic development for P. vitticeps based on standardized staging systems and external morphological characters previously defined for squamates. Whereas the overall morphological development follows the general trends observed in other squamates, our comparisons indicate major differences in the developmental sequence of several tissues, including early craniofacial characters. Detailed analysis of both embryonic skull development and adult skull shape, using a comparative approach integrating CT-scans and gene expression studies in P. vitticeps as well as comparative embryology and 3D geometric morphometrics in a large dataset of lizards and snakes, highlights the extreme adult skull shape of P. vitticeps and further indicates that heterochrony has played a key role in the early development and ossification of squamate skull bones. Such detailed studies of embryonic character development, craniofacial patterning, and bone formation are essential for the establishment of well-selected squamate species as Evo-Devo model organisms. We expect that P. vitticeps will continue to emerge as a new attractive model organism for understanding developmental and molecular processes underlying tissue formation, morphology, and evolution.

Skull Development, Ossification Pattern, and Adult Shape in the Emerging Lizard Model Organism Pogona vitticeps: A Comparative Analysis With Other Squamates

PubMed Central

Ollonen, Joni; Da Silva, Filipe O.; Mahlow, Kristin; Di-Poï, Nicolas

2018-01-01

The rise of the Evo-Devo field and the development of multidisciplinary research tools at various levels of biological organization have led to a growing interest in researching for new non-model organisms. Squamates (lizards and snakes) are particularly important for understanding fundamental questions about the evolution of vertebrates because of their high diversity and evolutionary innovations and adaptations that portrait a striking body plan change that reached its extreme in snakes. Yet, little is known about the intricate connection between phenotype and genotype in squamates, partly due to limited developmental knowledge and incomplete characterization of embryonic development. Surprisingly, squamate models have received limited attention in comparative developmental studies, and only a few species examined so far can be considered as representative and appropriate model organism for mechanistic Evo-Devo studies. Fortunately, the agamid lizard Pogona vitticeps (central bearded dragon) is one of the most popular, domesticated reptile species with both a well-established history in captivity and key advantages for research, thus forming an ideal laboratory model system and justifying his recent use in reptile biology research. We first report here the complete post-oviposition embryonic development for P. vitticeps based on standardized staging systems and external morphological characters previously defined for squamates. Whereas the overall morphological development follows the general trends observed in other squamates, our comparisons indicate major differences in the developmental sequence of several tissues, including early craniofacial characters. Detailed analysis of both embryonic skull development and adult skull shape, using a comparative approach integrating CT-scans and gene expression studies in P. vitticeps as well as comparative embryology and 3D geometric morphometrics in a large dataset of lizards and snakes, highlights the extreme adult skull shape of P. vitticeps and further indicates that heterochrony has played a key role in the early development and ossification of squamate skull bones. Such detailed studies of embryonic character development, craniofacial patterning, and bone formation are essential for the establishment of well-selected squamate species as Evo-Devo model organisms. We expect that P. vitticeps will continue to emerge as a new attractive model organism for understanding developmental and molecular processes underlying tissue formation, morphology, and evolution. PMID:29643813

Reduced cell number in the hindgut epithelium disrupts hindgut left-right asymmetry in a mutant of pebble, encoding a RhoGEF, in Drosophila embryos.

PubMed

Nakamura, Mitsutoshi; Matsumoto, Kenjiroo; Iwamoto, Yuta; Muguruma, Takeshi; Nakazawa, Naotaka; Hatori, Ryo; Taniguchi, Kiichiro; Maeda, Reo; Matsuno, Kenji

2013-02-01

Animals often show left-right (LR) asymmetry in their body structures. In some vertebrates, the mechanisms underlying LR symmetry breaking and the subsequent signals responsible for LR asymmetric development are well understood. However, in invertebrates, the molecular bases of these processes are largely unknown. Therefore, we have been studying the genetic pathway of LR asymmetric development in Drosophila. The embryonic gut is the first organ that shows directional LR asymmetry during Drosophila development. We performed a genetic screen to identify mutations affecting LR asymmetric development of the embryonic gut. From this screen, we isolated pebble (pbl), which encodes a homolog of a mammalian RhoGEF, Ect2. The laterality of the hindgut was randomized in embryos homozygous for a null mutant of pbl. Pbl is a multi-functional protein required for cytokinesis and the epithelial-to-mesenchymal transition in Drosophila. Consistent with Pbl's role in cytokinesis, we found reduced numbers of cells in the hindgut epithelium in pbl homozygous embryos. The specific expression of pbl in the hindgut epithelium, but not in other tissues, rescued the LR defects and reduced cell number in embryonic pbl homozygotes. Embryos homozygous for string (stg), a mutant that reduces cell number through a different mechanism, also showed LR defects of the hindgut. However, the reduction in cell number in the pbl mutants was not accompanied by defects in the specification of hindgut epithelial tissues or their integrity. Based on these results, we speculate that the reduction in cell number may be one reason for the LR asymmetry defect of the pbl hindgut, although we cannot exclude contributions from other functions of Pbl, including regulation of the actin cytoskeleton through its RhoGEF activity. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Effects of an environmentally relevant polychlorinated biphenyl (PCB) mixture on embryonic survival and cardiac development in the domestic chicken.

PubMed

Carro, Tiffany; Dean, Karen; Ottinger, Mary Ann

2013-06-01

A 58-congener polychlorinated biphenyl (PCB) mixture based on contaminant analysis of spotted sandpiper eggs collected along the upper Hudson River, New York, USA, in 2004 was used to study in ovo PCB effects on cardiac development in the domestic chicken. Fertile eggs were injected prior to incubation with the following doses of the PCB mixture: untreated, sham, 0, 0.03, 0.08, 0.3, 0.5, 0.7, and 2.06 µg PCBs/g egg weight (toxic equivalent quotient [TEQ] range of 0.004-0.266 ng/g). In addition, there were untreated and sham-control groups. Embryonic development was monitored throughout incubation and chicks were necropsied at hatch. Hatchability followed a dose-dependent curve with significant (p < 0.05) mortality above the 0.5 µg PCBs/g egg weight treatment compared with controls. The median lethal dose (LD50) of this PCB mixture in hatchling chicks was estimated as 0.4 µg/g egg weight (0.052 ng TEQ/g egg wt) based on the lethality curve. Cardiac arrhythmia was observed at embryonic day 14 of development in embryos treated at concentrations of 0.5 µg/g egg weight and above. Histological analysis was utilized to characterize any cardiac abnormalities. Cardiomyopathies increased across treatments in a dose-dependent manner compared with control groups. Identified abnormalities included the absence of the trabeculated layer of the ventricular wall, ventricular dilation, thinning of the ventricular walls, malformation of the septal wall, and most commonly, absence of the compact layer of the ventricular wall. Chick heart width, depth, total area, compact layer depth, septal width, chamber area, and ventricular wall dimensions did not differ across treatments. The present study supports prior reports of adverse developmental effects of PCBs on cardiovascular systems in birds. Although the eggs hatched, measured cardiomyopathies suggest potential deleterious long-term impacts on individual health and fitness. Copyright © 2013 SETAC.

Vitamin K2 biosynthetic enzyme, UBIAD1 is essential for embryonic development of mice.

PubMed

Nakagawa, Kimie; Sawada, Natsumi; Hirota, Yoshihisa; Uchino, Yuri; Suhara, Yoshitomo; Hasegawa, Tomoka; Amizuka, Norio; Okamoto, Tadashi; Tsugawa, Naoko; Kamao, Maya; Funahashi, Nobuaki; Okano, Toshio

2014-01-01

UbiA prenyltransferase domain containing 1 (UBIAD1) is a novel vitamin K2 biosynthetic enzyme screened and identified from the human genome database. UBIAD1 has recently been shown to catalyse the biosynthesis of Coenzyme Q10 (CoQ10) in zebrafish and human cells. To investigate the function of UBIAD1 in vivo, we attempted to generate mice lacking Ubiad1, a homolog of human UBIAD1, by gene targeting. Ubiad1-deficient (Ubiad1(-/-)) mouse embryos failed to survive beyond embryonic day 7.5, exhibiting small-sized body and gastrulation arrest. Ubiad1(-/-) embryonic stem (ES) cells failed to synthesize vitamin K2 but were able to synthesize CoQ9, similar to wild-type ES cells. Ubiad1(+/-) mice developed normally, exhibiting normal growth and fertility. Vitamin K2 tissue levels and synthesis activity were approximately half of those in the wild-type, whereas CoQ9 tissue levels and synthesis activity were similar to those in the wild-type. Similarly, UBIAD1 expression and vitamin K2 synthesis activity of mouse embryonic fibroblasts prepared from Ubiad1(+/-) E15.5 embryos were approximately half of those in the wild-type, whereas CoQ9 levels and synthesis activity were similar to those in the wild-type. Ubiad1(-/-) mouse embryos failed to be rescued, but their embryonic lifespans were extended to term by oral administration of MK-4 or CoQ10 to pregnant Ubiad1(+/-) mice. These results suggest that UBIAD1 is responsible for vitamin K2 synthesis but may not be responsible for CoQ9 synthesis in mice. We propose that UBIAD1 plays a pivotal role in embryonic development by synthesizing vitamin K2, but may have additional functions beyond the biosynthesis of vitamin K2.

CITED1 Expression in Liver Development and Hepatoblastoma12

PubMed Central

Murphy, Andrew J; de Caestecker, Christian; Pierce, Janene; Boyle, Scott C; Ayers, Gregory D; Zhao, Zhiguo; Libes, Jaime M; Correa, Hernan; Walter, Teagan; Huppert, Stacey S; Perantoni, Alan O; de Caestecker, Mark P; Lovvorn, Harold N

2012-01-01

Hepatoblastoma, the most common pediatric liver cancer, consists of epithelial mixed embryonal/fetal (EMEF) and pure fetal histologic subtypes, with the latter exhibiting a more favorable prognosis. Few embryonal histology markers that yield insight into the biologic basis for this prognostic discrepancy exist. CBP/P-300 interacting transactivator 1 (CITED1), a transcriptional co-activator, is expressed in the self-renewing nephron progenitor population of the developing kidney and broadly in its malignant analog, Wilms tumor (WT). In this current study, CITED1 expression is detected in mouse embryonic liver initially on post-coitum day 10.5 (e10.5), begins to taper by e14.5, and is undetectable in e18.5 and adult livers. CITED1 expression is detected in regenerating murine hepatocytes following liver injury by partial hepatectomy and 3,5-diethoxycarbonyl-1,4-dihydrocollidine. Importantly, while CITED1 is undetectable in normal human adult livers, 36 of 41 (87.8%) hepatoblastoma specimens express CITED1, where it is enriched in EMEF specimens compared to specimens of pure fetal histology. CITED1 overexpression in Hep293TT human hepatoblastoma cells induces cellular proliferation and upregulates the Wnt inhibitors Kringle containing transmembrane protein 1 (KREMEN1) and CXXC finger protein 4 (CXXC4). CITED1 mRNA expression correlates with expression of CXXC4 and KREMEN1 in clinical hepatoblastoma specimens. These data show that CITED1 is expressed during a defined time course of liver development and is no longer expressed in the adult liver but is upregulated in regenerating hepatocytes following liver injury. Moreover, as in WT, this embryonic marker is reexpressed in hepatoblastoma and correlates with embryonal histology. These findings identify CITED1 as a novel marker of hepatic progenitor cells that is re-expressed following liver injury and in embryonic liver tumors. PMID:23308048

Lamin A/C Haploinsufficiency Modulates the Differentiation Potential of Mouse Embryonic Stem Cells

PubMed Central

Sehgal, Poonam; Chaturvedi, Pankaj; Kumaran, R. Ileng; Kumar, Satish; Parnaik, Veena K.

2013-01-01

Background Lamins are structural proteins that are the major determinants of nuclear architecture and play important roles in various nuclear functions including gene regulation and cell differentiation. Mutations in the human lamin A gene cause a spectrum of genetic diseases that affect specific tissues. Most available mouse models for laminopathies recapitulate disease symptoms for muscle diseases and progerias. However, loss of human lamin A/C also has highly deleterious effects on fetal development. Hence it is important to understand the impact of lamin A/C expression levels on embryonic differentiation pathways. Methodology and Principal Findings We have investigated the differentiation potential of mouse embryonic stem cells containing reduced levels of lamin A/C by detailed lineage analysis of embryoid bodies derived from these cells by in vitro culture. We initially carried out a targeted disruption of one allele of the mouse lamin A/C gene (Lmna). Undifferentiated wild-type and Lmna+/− embryonic stem cells showed similar expression of pluripotency markers and cell cycle profiles. Upon spontaneous differentiation into embryoid bodies, markers for visceral endoderm such as α-fetoprotein were highly upregulated in haploinsufficient cells. However, neuronal markers such as β-III tubulin and nestin were downregulated. Furthermore, we observed a reduction in the commitment of Lmna+/− cells into the myogenic lineage, but no discernible effects on cardiac, adipocyte or osteocyte lineages. In the next series of experiments, we derived embryonic stem cell clones expressing lamin A/C short hairpin RNA and examined their differentiation potential. These cells expressed pluripotency markers and, upon differentiation, the expression of lineage-specific markers was altered as observed with Lmna+/− embryonic stem cells. Conclusions We have observed significant effects on embryonic stem cell differentiation to visceral endoderm, neuronal and myogenic lineages upon depletion of lamin A/C. Hence our results implicate lamin A/C level as an important determinant of lineage-specific differentiation during embryonic development. PMID:23451281

Diversity and Complexity in Chromatin Recognition by TFII-I Transcription Factors in Pluripotent Embryonic Stem Cells and Embryonic Tissues

PubMed Central

Makeyev, Aleksandr V.; Enkhmandakh, Badam; Hong, Seung-Hyun; Joshi, Pujan; Shin, Dong-Guk; Bayarsaihan, Dashzeveg

2012-01-01

GTF2I and GTF2IRD1 encode a family of closely related transcription factors TFII-I and BEN critical in embryonic development. Both genes are deleted in Williams-Beuren syndrome, a complex genetic disorder associated with neurocognitive, craniofacial, dental and skeletal abnormalities. Although genome-wide promoter analysis has revealed the existence of multiple TFII-I binding sites in embryonic stem cells (ESCs), there was no correlation between TFII-I occupancy and gene expression. Surprisingly, TFII-I recognizes the promoter sequences enriched for H3K4me3/K27me3 bivalent domain, an epigenetic signature of developmentally important genes. Moreover, we discovered significant differences in the association between TFII-I and BEN with the cis-regulatory elements in ESCs and embryonic craniofacial tissues. Our data indicate that in embryonic tissues BEN, but not the highly homologous TFII-I, is primarily recruited to target gene promoters. We propose a “feed-forward model” of gene regulation to explain the specificity of promoter recognition by TFII-I factors in eukaryotic cells. PMID:22970219

Diversity and complexity in chromatin recognition by TFII-I transcription factors in pluripotent embryonic stem cells and embryonic tissues.

PubMed

Makeyev, Aleksandr V; Enkhmandakh, Badam; Hong, Seung-Hyun; Joshi, Pujan; Shin, Dong-Guk; Bayarsaihan, Dashzeveg

2012-01-01

GTF2I and GTF2IRD1 encode a family of closely related transcription factors TFII-I and BEN critical in embryonic development. Both genes are deleted in Williams-Beuren syndrome, a complex genetic disorder associated with neurocognitive, craniofacial, dental and skeletal abnormalities. Although genome-wide promoter analysis has revealed the existence of multiple TFII-I binding sites in embryonic stem cells (ESCs), there was no correlation between TFII-I occupancy and gene expression. Surprisingly, TFII-I recognizes the promoter sequences enriched for H3K4me3/K27me3 bivalent domain, an epigenetic signature of developmentally important genes. Moreover, we discovered significant differences in the association between TFII-I and BEN with the cis-regulatory elements in ESCs and embryonic craniofacial tissues. Our data indicate that in embryonic tissues BEN, but not the highly homologous TFII-I, is primarily recruited to target gene promoters. We propose a "feed-forward model" of gene regulation to explain the specificity of promoter recognition by TFII-I factors in eukaryotic cells.

Alterations in the developing testis transcriptome following embryonic vinclozolin exposure.

PubMed

Clement, Tracy M; Savenkova, Marina I; Settles, Matthew; Anway, Matthew D; Skinner, Michael K

2010-11-01

The current study investigates the direct effects of in utero vinclozolin exposure on the developing F1 generation rat testis transcriptome. Previous studies have demonstrated that exposure to vinclozolin during embryonic gonadal sex determination induces epigenetic modifications of the germ line and transgenerational adult onset disease states. Microarray analyses were performed to compare control and vinclozolin treated testis transcriptomes at embryonic days 13, 14 and 16. A total of 576 differentially expressed genes were identified and the major cellular functions and pathways associated with these altered transcripts were examined. The sets of regulated genes at the different development periods were found to be transiently altered and distinct. Categorization by major known functions of altered genes was performed. Specific cellular process and pathway analyses suggest the involvement of Wnt and calcium signaling, vascular development and epigenetic mechanisms as potential mediators of the direct F1 generation actions of vinclozolin. Copyright © 2010 Elsevier Inc. All rights reserved.

ALTERATIONS IN THE DEVELOPING TESTIS TRANSCRIPTOME FOLLOWING EMBRYONIC VINCLOZOLIN EXPOSURE

PubMed Central

Clement, Tracy M.; Savenkova, Marina I.; Settles, Matthew; Anway, Matthew D.; Skinner, Michael K.

2010-01-01

The current study investigates the direct effects of in utero vinclozolin exposure on the developing F1 generation rat testis transcriptome. Previous studies have demonstrated that exposure to vinclozolin during embryonic gonadal sex determination induces epigenetic modifications of the germ line and transgenerational adult onset disease states. Microarray analyses were performed to compare control and vinclozolin treated testis transcriptomes at embryonic day 13, 14 and 16. A total of 576 differentially expressed genes were identified and the major cellular functions and pathways associated with these altered transcripts were examined. The sets of regulated genes at the different development periods were found to be transiently altered and distinct. Categorization by major known functions of altered genes was performed. Specific cellular process and pathway analyses suggest the involvement of Wnt and calcium signaling, vascular development and epigenetic mechanisms as potential mediators of the direct F1 generation actions of vinclozolin. PMID:20566332

Characterizing the distribution of steroid sulfatase during embryonic development: when and where might metabolites of maternal steroids be reactivated?

PubMed

Paitz, Ryan T; Duffield, Kristin R; Bowden, Rachel M

2017-12-15

All vertebrate embryos are exposed to maternally derived steroids during development. In placental vertebrates, metabolism of maternal steroids by the placenta modulates embryonic exposure, but how exposure is regulated in oviparous vertebrates is less clear. Recent work in oviparous vertebrates has demonstrated that steroids are not static molecules, as they can be converted to more polar steroid sulfates by sulfotransferase enzymes. Importantly, these steroid sulfates can be converted back to the parent compound by the enzyme steroid sulfatase (STS). We investigated when and where STS was present during embryonic development in the red-eared slider turtle, Trachemys scripta We report that STS is present during all stages of development and in all tissues we examined. We conclude that STS activity may be particularly important for regulating maternal steroid exposure in oviparous vertebrates. © 2017. Published by The Company of Biologists Ltd.

Mirror-image duplication of the primary axis and heart in Xenopus embryos by the overexpression of Msx-1 gene.

PubMed

Chen, Y; Solursh, M

1995-10-01

The Msx-1 gene (formerly known as Hox-7) is a member of a discrete subclass of homeobox-containing genes. Examination of the expression pattern of Msx-1 in murine and avian embryos suggests that this gene may be involved in the regionalization of the medio-lateral axis during earlier development. We have examined the possible functions of Xenopus Msx-1 during early Xenopus embryonic development by overexpression of the Msx-1 gene. Overexpression of Msx-1 causes a left-right mirror-image duplication of primary axial structures, including notochord, neural tube, somites, suckers, and foregut. The embryonic developing heart is also mirror-image duplicated, including looping directions and polarity. These results indicate that Msx-1 may be involved in the mesoderm formation as well as left-right patterning in the early Xenopus embryonic development.

Watch-ing out for chick limb development.

PubMed

Pascoal, Susana; Palmeirim, Isabel

2007-09-01

Time control is a crucial issue during embryonic development. Nevertheless, little is known about how embryonic cells measure time. Until recently, the only molecular clock known to operate during vertebrate embryonic development was the somitogenesis clock, exclusively functioning in coordinating the precise timing of each new pair of somites formed from the presomitic mesoderm. We have recently evidenced that a similar molecular clock also underlies the timing at which autopod chondrogenic precursors are laid down to form a skeletal limb element. In addition, we herein suggest that the molecular clock is not the only parallelism that can be established between somitogenesis and limb-bud development. In an evolutionary perspective, we support the previously proposed idea that the molecular mechanisms involved in the segmentation of the body axis may have been partially reused in the mesoderm of the lateral plate, thereby allowing the emergence of paired appendages.

The effects of 1α, 25-dihydroxyvitamin D3 and transforming growth factor-β3 on bone development in an ex vivo organotypic culture system of embryonic chick femora.

PubMed

Smith, Emma L; Rashidi, Hassan; Kanczler, Janos M; Shakesheff, Kevin M; Oreffo, Richard O C

2015-01-01

Transforming growth factor-beta3 (TGF-β3) and 1α,25-dihydroxyvitamin D3 (1α,25 (OH) 2D3) are essential factors in chondrogenesis and osteogenesis respectively. These factors also play a fundamental role in the developmental processes and the maintenance of skeletal integrity, but their respective direct effects on these processes are not fully understood. Using an organotypic bone rudiment culture system the current study has examined the direct roles the osteotropic factors 1α,25 (OH)2D3 and TGF-β3 exert on the development and modulation of the three dimensional structure of the embryonic femur. Isolated embryonic chick femurs (E11) were organotypically cultured for 10 days in basal media, or basal media supplemented with either 1α,25 (OH) 2D3 (25 nM) or TGF-β3 (5 ng/mL & 15 ng/mL). Analyses of the femurs were undertaken using micro-computed tomography (μCT), histology and immunohistochemistry. 1α,25 (OH)2D3 supplemented cultures enhanced osteogenesis directly in the developing femurs with elevated levels of osteogenic markers such as type 1 collagen. In marked contrast organotypic femur cultures supplemented with TGF-β3 (5 ng/mL & 15 ng/mL) demonstrated enhanced chondrogenesis with a reduction in osteogenesis. These studies demonstrate the efficacy of the ex vivo organotypic embryonic femur culture employed to elucidate the direct roles of these molecules, 1α,25 (OH) 2D3 and TGF-β3 on the structural development of embryonic bone within a three dimensional framework. We conclude that 1α,25(OH)2D and TGF-β3 modify directly the various cell populations in bone rudiment organotypic cultures effecting tissue metabolism resulting in significant changes in embryonic bone growth and modulation. Understanding the roles of osteotropic agents in the process of skeletal development is integral to developing new strategies for the recapitulation of bone tissue in later life.

Nucleoli from two-cell embryos support the development of enucleolated germinal vesicle oocytes in the pig.

PubMed

Kyogoku, Hirohisa; Ogushi, Sugako; Miyano, Takashi

2012-11-01

Recent research has shown that nucleoli of oocytes at the germinal vesicle (GV) stage (GV nucleoli) are not necessary for oocyte maturation but are essential for early embryonic development. Nucleoli of 2-cell embryos (2-cell nucleoli) have morphology similar to that of nucleoli in oocytes at the GV stage. In this study, we examined the ability of 2-cell nucleoli to substitute for GV nucleoli in terms of supporting early embryonic development by nucleolus aspiration (enucleolation) and transfer into metaphase II (MII) oocytes or 2-cell embryos that were derived from enucleolated oocytes at the GV stage in the pig. When 2-cell embryos were centrifuged to move the lipid droplets to one side of the blastomere, multiple nucleoli in the nucleus fused into a single nucleolus. The nucleoli were then aspirated from the 2-cell embryos by micromanipulation. The injection of 2-cell nucleoli to GV enucleolated oocytes at the MII stage rescued the embryos from the early embryonic arrest, and the resulting oocytes developed to blastocysts. However, the injection of 2-cell and GV nucleoli to 2-cell embryos derived from GV enucleolated oocytes rarely restored the development to blastocysts. These results indicate that 2-cell nucleoli support early embryonic development as GV nucleoli and that the presence of nucleoli is essential for pig embryos before the 2-cell stage.

Knockout of the PKN Family of Rho Effector Kinases Reveals a Non-redundant Role for PKN2 in Developmental Mesoderm Expansion

PubMed Central

Quétier, Ivan; Marshall, Jacqueline J.T.; Spencer-Dene, Bradley; Lachmann, Sylvie; Casamassima, Adele; Franco, Claudio; Escuin, Sarah; Worrall, Joseph T.; Baskaran, Priththivika; Rajeeve, Vinothini; Howell, Michael; Copp, Andrew J.; Stamp, Gordon; Rosewell, Ian; Cutillas, Pedro; Gerhardt, Holger; Parker, Peter J.; Cameron, Angus J.M.

2016-01-01

Summary In animals, the protein kinase C (PKC) family has expanded into diversely regulated subgroups, including the Rho family-responsive PKN kinases. Here, we describe knockouts of all three mouse PKN isoforms and reveal that PKN2 loss results in lethality at embryonic day 10 (E10), with associated cardiovascular and morphogenetic defects. The cardiovascular phenotype was not recapitulated by conditional deletion of PKN2 in endothelial cells or the developing heart. In contrast, inducible systemic deletion of PKN2 after E7 provoked collapse of the embryonic mesoderm. Furthermore, mouse embryonic fibroblasts, which arise from the embryonic mesoderm, depend on PKN2 for proliferation and motility. These cellular defects are reflected in vivo as dependence on PKN2 for mesoderm proliferation and neural crest migration. We conclude that failure of the mesoderm to expand in the absence of PKN2 compromises cardiovascular integrity and development, resulting in lethality. PMID:26774483

Knockout of the PKN Family of Rho Effector Kinases Reveals a Non-redundant Role for PKN2 in Developmental Mesoderm Expansion.

PubMed

Quétier, Ivan; Marshall, Jacqueline J T; Spencer-Dene, Bradley; Lachmann, Sylvie; Casamassima, Adele; Franco, Claudio; Escuin, Sarah; Worrall, Joseph T; Baskaran, Priththivika; Rajeeve, Vinothini; Howell, Michael; Copp, Andrew J; Stamp, Gordon; Rosewell, Ian; Cutillas, Pedro; Gerhardt, Holger; Parker, Peter J; Cameron, Angus J M

2016-01-26

In animals, the protein kinase C (PKC) family has expanded into diversely regulated subgroups, including the Rho family-responsive PKN kinases. Here, we describe knockouts of all three mouse PKN isoforms and reveal that PKN2 loss results in lethality at embryonic day 10 (E10), with associated cardiovascular and morphogenetic defects. The cardiovascular phenotype was not recapitulated by conditional deletion of PKN2 in endothelial cells or the developing heart. In contrast, inducible systemic deletion of PKN2 after E7 provoked collapse of the embryonic mesoderm. Furthermore, mouse embryonic fibroblasts, which arise from the embryonic mesoderm, depend on PKN2 for proliferation and motility. These cellular defects are reflected in vivo as dependence on PKN2 for mesoderm proliferation and neural crest migration. We conclude that failure of the mesoderm to expand in the absence of PKN2 compromises cardiovascular integrity and development, resulting in lethality. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Characteristics and incidence of large eggs in Trichuris muris.

PubMed

Koyama, Koichi

2013-05-01

The production of small numbers of large eggs among the standard-sized eggs of Trichuris trichiura is well known. Large eggs have also been observed in Trichuris muris, but they have not been studied previously. This paper compares the characteristics of the large eggs (LEs, ≥74.5 μm long) and standard-sized eggs (SEs, <74.5 μm long) in cultures of T. muris. Among 112,554 cultured eggs, LEs occurred at very low frequency (0.03 %, i.e., about three large eggs per 10(4) cultured eggs). Embryonated eggs represented 93.72 % of SEs, but only 25.00 % of LEs were embryonated. Embryonated LEs and SEs contained fully matured larvae. An atypical category of unembryonated egg, which contained an incompletely developed larva, an abnormal larva, or granular components, was common among the LEs. However, similar atypical unembryonated SEs were rarely observed. These observations suggest that the LEs that occur very infrequently in T. muris result from an abnormality of embryonation (larval development).

Toxicological effects of mainstream whole smoke solutions on embryonic movements of the developing embryo.

PubMed

Ejaz, Sohail; Seok, Kim Bum; Woong, Lim Chae

2005-01-01

Cigarette smoking is unrivaled among developmental toxicants in terms of total adverse impact on the human population. Maternal tobacco use during pregnancy adversely affects prenatal and postnatal growth and increases the risk of behavioral and developmental defects in children and adolescents. In the current study, the effects of different preparations of nicotine and mainstream whole smoke solutions (MSWSS) on embryonic movements during neonatal development were examined in vivo, using the chicken embryo model, recorded in real-time by a video camera. It was observed that low doses of nicotine induced hyperactivity and higher doses induced hypoactivity. Accordingly, a significant (p < 0.01) decrease in movements was observed by application of 10 microg of nicotine and different preparations of MSWSS. A dose-dependent decrease in embryonic movements was observed, which did not recover by the end of experiment. It was concluded that nicotine could alter embryonic movements, which are important during embryogenesis for differentiation and maturation of the body systems.

RIPK3 Mediates Necroptosis during Embryonic Development and Postnatal Inflammation in Fadd-Deficient Mice.

PubMed

Zhao, Qun; Yu, XianJun; Zhang, HaiWei; Liu, YongBo; Zhang, XiXi; Wu, XiaoXia; Xie, Qun; Li, Ming; Ying, Hao; Zhang, Haibing

2017-04-25

RIPK3 mediates cell death and regulates inflammatory responses. Although genetic studies have suggested that RIPK3-MLKL-mediated necroptosis leads to embryonic lethality in Fadd or Caspase-8-deficient mice, the exact mechanisms are not fully understood. Here, we generated Ripk3 mutant mice by altering the RIPK3 kinase domain (Ripk3 Δ/Δ mice), thus abolishing its kinase activity. Ripk3 Δ/Δ cells were resistant to necroptosis stimulation in vitro, and Ripk3 Δ/Δ mice were protected from necroptotic diseases. Although the Ripk3 Δ/Δ mutation rescued embryonic lethality in Fadd -/- embryos, Fadd -/- Ripk3 Δ/Δ mice died within 1 day after birth due to massive inflammation. These results indicate that Ripk3 ablation rescues embryonic lethality in Fadd-deficient mice by suppressing two RIPK3-mediating processes: necroptosis during embryogenesis and inflammation during postnatal development in Fadd -/- mice. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

[Recent contributions to the establishment of the axes of the mammalian embryo].

PubMed

Catala, M

2002-06-01

The study of the establishment of embryonic axes during early development has shown that this process is a very early event (occurRing either during ovogenesis or during fertilization) for invertebrates and for lower vertebrates. In mammals, it was considered that this establishment appears late during development because of the great plasticity of blastomeres. Recent data in the mouse embryon show that the mammalian ovocyte is a polarized cell, the polar body corresponding to the animal pole of this cell. The blastomeres that are generated by the zygote divide asynchronously. The first that divides is the one which inherits the plasma cell membrane where fertilization takes place. This blastomere will preferentially give rise to the cells of the embryonic pole of the blastocyst whereas the other yields the cells of the abembryonic pole. The mammalian ovocyte is thus a polarized cell with an already established animal-vegetal axis. The point of sperm entry will determine the embryonic-abembryonic axis.

Redox environment in stem and differentiated cells: A quantitative approach.

PubMed

Lyublinskaya, O G; Ivanova, Ju S; Pugovkina, N A; Kozhukharova, I V; Kovaleva, Z V; Shatrova, A N; Aksenov, N D; Zenin, V V; Kaulin, Yu A; Gamaley, I A; Nikolsky, N N

2017-08-01

Stem cells are believed to maintain a specific intracellular redox status through a combination of enhanced removal capacity and limited production of ROS. In the present study, we challenge this assumption by developing a quantitative approach for the analysis of the pro- and antioxidant ability of human embryonic stem cells in comparison with their differentiated descendants, as well as adult stem and non-stem cells. Our measurements showed that embryonic stem cells are characterized by low ROS level, low rate of extracellular hydrogen peroxide removal and low threshold for peroxide-induced cytotoxicity. However, biochemical normalization of these parameters to cell volume/protein leads to matching of normalized values in stem and differentiated cells and shows that tested in the present study cells (human embryonic stem cells and their fibroblast-like progenies, adult mesenchymal stem cells, lymphocytes, HeLa) maintain similar intracellular redox status. Based on these observations, we propose to use ROS concentration averaged over the cell volume instead of ROS level as a measure of intracellular redox balance. We show that attempts to use ROS level for comparative analysis of redox status of morphologically different cells could lead to false conclusions. Methods for the assessment of ROS concentration based on flow cytometry analysis with the use of H 2 DCFDA dye and HyPer, genetically encoded probe for hydrogen peroxide, are discussed. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Induced Wnt5a expression perturbs embryonic outgrowth and intestinal elongation, but is well-tolerated in adult mice.

PubMed

Bakker, Elvira R M; Raghoebir, Lalini; Franken, Patrick F; Helvensteijn, Werner; van Gurp, Léon; Meijlink, Frits; van der Valk, Martin A; Rottier, Robbert J; Kuipers, Ernst J; van Veelen, Wendy; Smits, Ron

2012-09-01

Wnt5a is essential during embryonic development, as indicated by mouse Wnt5a knockout embryos displaying outgrowth defects of multiple structures including the gut. The dynamics of Wnt5a involvement in these processes is unclear, and perinatal lethality of Wnt5a knockout embryos has hampered investigation of Wnt5a during postnatal stages in vivo. Although in vitro studies have suggested a relevant role for Wnt5a postnatally, solid evidence for a significant impact of Wnt5a within the complexity of an adult organism is lacking. We generated a tightly-regulated inducible Wnt5a transgenic mouse model and investigated the effects of Wnt5a induction during different time-frames of embryonic development and in adult mice, focusing on the gastrointestinal tract. When induced in embryos from 10.5 dpc onwards, Wnt5a expression led to severe outgrowth defects affecting the gastrointestinal tracts, limbs, facial structures and tails, closely resembling the defects observed in Wnt5a knockout mice. However, Wnt5a induction from 13.5 dpc onwards did not cause this phenotype, indicating that the most critical period for Wnt5a in embryonic development is prior to 13.5 dpc. In adult mice, induced Wnt5a expression did not reveal abnormalities, providing the first in vivo evidence that Wnt5a has no major impact on mouse intestinal homeostasis postnatally. Protein expression of Wnt5a receptor Ror2 was strongly reduced in adult intestine compared to embryonic stages. Moreover, we uncovered a regulatory process where induction of Wnt5a causes downregulation of its receptor Ror2. Taken together, our results indicate a role for Wnt5a during a restricted time-frame of embryonic development, but suggest no impact during homeostatic postnatal stages. Copyright © 2012 Elsevier Inc. All rights reserved.

Forkhead box transcription factors in embryonic heart development and congenital heart disease.

PubMed

Zhu, Hong

2016-01-01

Embryonic heart development is a very complicated process regulated precisely by a network composed of many genes and signaling pathways in time and space. Forkhead box (Fox, FOX) proteins are a family of transcription factors characterized by the presence of an evolutionary conserved "forkhead"or "winged-helix" DNA-binding domain and able to organize temporal and spatial gene expression during development. They are involved in a wide variety of cellular processes, such as cell cycle progression, proliferation, differentiation, migration, metabolism and DNA damage response. An abundance of studies in model organisms and systems has established that Foxa2, Foxc1/c2, Foxh1 and Foxm1, Foxos and Foxps are important components of the signaling pathways that instruct cardiogenesis and embryonic heart development, playing paramount roles in heart development. The previous studies also have demonstrated that mutations in some of the forkhead box genes and the aberrant expression of forkhead box gene are heavily implicated in the congenital heart disease (CHD) of humans. This review primarily focuses on the current understanding of heart development regulated by forkhead box transcription factors and molecular genetic mechanisms by which forkhead box factors modulate heart development during embryogenesis and organogenesis. This review also summarizes human CHD related mutations in forkhead box genes as well as the abnormal expression of forkhead box gene, and discusses additional possible regulatory mechanisms of the forkhead box genes during embryonic heart development that warrant further investigation. Copyright © 2015 Elsevier Inc. All rights reserved.

The effects of triclosan on pluripotency factors and development of mouse embryonic stem cells and zebrafish.

PubMed

Chen, Xiaojiao; Xu, Bo; Han, Xiumei; Mao, Zhilei; Chen, Minjian; Du, Guizhen; Talbot, Prue; Wang, Xinru; Xia, Yankai

2015-04-01

Triclosan (TCS) poses potential risks to reproduction and development due to its endocrine-disrupting properties. However, the mechanism of TCS's effects on early embryonic development is little known. Embryonic stem cells (ESC) and zebrafish embryos provide valuable models for testing the toxic effects of environmental chemicals on early embryogenesis. In this study, mouse embryonic stem cells (mESC) were acutely exposed to TCS for 24 h, and general cytotoxicity and the effect of TCS on pluripotency were then evaluated. In addition, zebrafish embryos were exposed to TCS from 2- to 24-h post-fertilization (hpf), and their morphology was evaluated. In mESC, alkaline phosphatase staining was significantly decreased after treatment with the highest concentration of TCS (50 μM). Although the expression levels of Sox2 mRNA were not changed, the mRNA levels of Oct4 and Nanog in TCS-treated groups were significantly decreased compared to controls. In addition, the protein levels of Oct4, Sox2 and Nanog were significantly reduced in response to TCS treatment. MicroRNA (miR)-134, an expression inhibitor of pluripotency markers, was significantly increased in TCS-treated mESC. In zebrafish experiments, after 24 hpf of treatment, the controls had developed to the late stage of somitogenesis, while embryos exposed to 300 μg/L of TCS were still at the early stage of somitogenesis, and three genes (Oct4, Sox2 and Nanog) were upregulated in treated groups when compared with the controls. The two models demonstrated that TCS may affect early embryonic development by disturbing the expression of the pluripotency markers (Oct4, Sox2 and Nanog).

Bone matrix calcification during embryonic and postembryonic rat calvarial development assessed by SEM-EDX spectroscopy, XRD, and FTIR spectroscopy.

PubMed

Henmi, Akiko; Okata, Hiroshi; Anada, Takahisa; Yoshinari, Mariko; Mikami, Yasuto; Suzuki, Osamu; Sasano, Yasuyuki

2016-01-01

Bone mineral is constituted of biological hydroxyapatite crystals. In developing bone, the mineral crystal matures and the Ca/P ratio increases. However, how an increase in the Ca/P ratio is involved in maturation of the crystal is not known. The relationships among organic components and mineral changes are also unclear. The study was designed to investigate the process of calcification during rat calvarial bone development. Calcification was evaluated by analyzing the atomic distribution and concentration of Ca, P, and C with scanning electron microscopy (SEM)-energy-dispersive X-ray (EDX) spectroscopy and changes in the crystal structure with X-ray diffraction (XRD) and Fourier transform infrared (FTIR) spectroscopy. Histological analysis showed that rat calvarial bone formation started around embryonic day 16. The areas of Ca and P expanded, matching the region of the developing bone matrix, whereas the area of C became localized around bone. X-ray diffraction and FTIR analysis showed that the amorphous-like structure of the minerals at embryonic day 16 gradually transformed into poorly crystalline hydroxyapatite, whereas the proportion of mineral to protein increased until postnatal week 6. FTIR analysis also showed that crystallization of hydroxyapatite started around embryonic day 20, by which time SEM-EDX spectroscopy showed that the Ca/P ratio had increased and the C/Ca and C/P ratios had decreased significantly. The study suggests that the Ca/P molar ratio increases and the proportion of organic components such as proteins of the bone matrix decreases during the early stage of calcification, whereas crystal maturation continues throughout embryonic and postembryonic bone development.

Comparative analysis of miRNA expression during the development of insects of different metamorphosis modes and germ-band types.

PubMed

Ylla, Guillem; Piulachs, Maria-Dolors; Belles, Xavier

2017-10-11

Do miRNAs contribute to specify the germ-band type and the body structure in the insect embryo? Our goal was to address that issue by studying the changes in miRNA expression along the ontogeny of the German cockroach Blattella germanica, which is a short germ-band and hemimetabolan species. We sequenced small RNA libraries representing 11 developmental stages of B. germanica ontogeny (with especial emphasis on embryogenesis) and the changes in miRNA expression were examined. Data were compared with equivalent data for two long germ-band holometabolan species Drosophila melanogaster and Drosophila virilis, and the short germ-band holometabolan species Tribolium castaneum. The identification of B. germanica embryo small RNA sequences unveiled miRNAs not detected in previous studies, such as those of the MIR-309 family and 54 novel miRNAs. Four main waves of miRNA expression were recognized (with most miRNA changes occurring during the embryonic stages): the first from day 0 to day 1 of embryogenesis, the second during mid-embryogenesis (days 0-6), the third (with an acute expression peak) on day 2 of embryonic development, and the fourth during post-embryonic development. The second wave defined the boundaries of maternal-to-zygotic transition, with maternal mRNAs being cleared, presumably by Mir-309 and associated scavenger miRNAs. miRNAs follow well-defined patterns of expression over hemimetabolan ontogeny, patterns that are more diverse during embryonic development than during the nymphal stages. The results suggest that miRNAs play important roles in the developmental transitions between the embryonic stages of development (starting with maternal loading), during which they might influence the germ-band type and metamorphosis mode.

Distributional shift of urea production site from the extraembryonic yolk sac membrane to the embryonic liver during the development of cloudy catshark (Scyliorhinus torazame).

PubMed

Takagi, Wataru; Kajimura, Makiko; Tanaka, Hironori; Hasegawa, Kumi; Ogawa, Shuntaro; Hyodo, Susumu

2017-09-01

Urea is an essential osmolyte for marine cartilaginous fishes. Adult elasmobranchs and holocephalans are known to actively produce urea in the liver, muscle and other extrahepatic organs; however, osmoregulatory mechanisms in the developing cartilaginous fish embryo with an undeveloped urea-producing organ are poorly understood. We recently described the contribution of extraembryonic yolk sac membranes (YSM) to embryonic urea synthesis during the early developmental period of the oviparous holocephalan elephant fish (Callorhinchus milii). In the present study, to test whether urea production in the YSM is a general phenomenon among oviparous Chondrichthyes, we investigated gene expression and activities of ornithine urea cycle (OUC) enzymes together with urea concentrations in embryos of the elasmobranch cloudy catshark (Scyliorhinus torazame). The intracapsular fluid, in which the catshark embryo develops, had a similar osmolality to seawater, and embryos maintained a high concentration of urea at levels similar to that of adult plasma throughout development. Relative mRNA expressions and activities of catshark OUC enzymes were significantly higher in YSM than in embryos until stage 32. Concomitant with the development of the embryonic liver, the expression levels and activities of OUC enzymes were markedly increased in the embryo from stage 33, while those of the YSM decreased from stage 32. The present study provides further evidence that the YSM contributes to embryonic urea homeostasis until the liver and other extrahepatic organs become fully functional, and that urea-producing tissue shifts from the YSM to the embryonic liver in the late developmental period of oviparous marine cartilaginous fishes. Copyright © 2017 Elsevier Inc. All rights reserved.

Geographic variation in avian incubation periods and parental influences on embryonic temperature

USGS Publications Warehouse

Martin, T.E.; Auer, S.K.; Bassar, R.D.; Niklison, Alina M.; Lloyd, P.

2007-01-01

Theory predicts shorter embryonic periods in species with greater embryo mortality risk and smaller body size. Field studies of 80 passerine species on three continents yielded data that largely conflicted with theory; incubation (embryonic) periods were longer rather than shorter in smaller species, and egg (embryo) mortality risk explained some variation within regions, but did not explain larger differences in incubation periods among geographic regions. Incubation behavior of parents seems to explain these discrepancies. Bird embryos are effectively ectothermic and depend on warmth provided by parents sitting on the eggs to attain proper temperatures for development. Parents of smaller species, plus tropical and southern hemisphere species, commonly exhibited lower nest attentiveness (percent of time spent on the nest incubating) than larger and northern hemisphere species. Lower nest attentiveness produced cooler minimum and average embryonic temperatures that were correlated with longer incubation periods independent of nest predation risk or body size. We experimentally tested this correlation by swapping eggs of species with cool incubation temperatures with eggs of species with warm incubation temperatures and similar egg mass. Incubation periods changed (shortened or lengthened) as expected and verified the importance of egg temperature on development rate. Slower development resulting from cooler temperatures may simply be a cost imposed on embryos by parents and may not enhance offspring quality. At the same time, incubation periods of transferred eggs did not match host species and reflect intrinsic differences among species that may result from nest predation and other selection pressures. Thus, geographic variation in embryonic development may reflect more complex interactions than previously recognized. ?? 2007 The Author(s).

Redeployment of germ layers related TFs shows regionalized expression during two non-embryonic developments.

PubMed

Ricci, Lorenzo; Cabrera, Fabien; Lotito, Sonia; Tiozzo, Stefano

2016-08-01

In all non-vertebrate metazoan phyla, species that evolved non-embryonic developmental pathways as means of propagation or regeneration can be found. In this context, new bodies arise through asexual reproduction processes (such as budding) or whole body regeneration, that lack the familiar temporal and spatial cues classically associated with embryogenesis, like maternal determinants, or gastrulation. The molecular mechanisms underlying those non-embryonic developments (i.e., regeneration and asexual reproduction), and their relationship to those deployed during embryogenesis are poorly understood. We have addressed this question in the colonial ascidian Botryllus schlosseri, which undergoes an asexual reproductive process via palleal budding (PB), as well as a whole body regeneration by vascular budding (VB). We identified early regenerative structures during VB and then followed the fate of differentiating tissues during both non-embryonic developments (PB and VB) by monitoring the expression of genes known to play key functions in germ layer specification with well conserved expression patterns in solitary ascidian embryogenesis. The expression patterns of FoxA1, GATAa, GATAb, Otx, Bra, Gsc and Tbx2/3 were analysed during both PB and VB. We found that the majority of these transcription factors were expressed during both non-embryonic developmental processes, revealing a regionalization of the palleal and vascular buds. Knockdown of GATAa by siRNA in palleal buds confirmed that preventing the correct development of one of these regions blocks further tissue specification. Our results indicate that during both normal and injury-induced budding, a similar alternative developmental program operates via early commitment of epithelial regions. Copyright © 2016. Published by Elsevier Inc.

Embryonic exposure to model naphthenic acids delays growth and hatching in the pond snail Lymnaea stagnalis.

PubMed

Johnston, Christina U; Clothier, Lindsay N; Quesnel, Dean M; Gieg, Lisa M; Chua, Gordon; Hermann, Petra M; Wildering, Willem C

2017-02-01

Naphthenic acids (NAs), a class of structurally diverse carboxylic acids with often complex ring structures and large aliphatic tail groups, are important by-products of many petrochemical processes including the oil sands mining activity of Northern Alberta. While it is evident that NAs have both acute and chronic harmful effects on many organisms, many aspects of their toxicity remain to be clarified. Particularly, while substantive data sets have been collected on NA toxicity in aquatic prokaryote and vertebrate model systems, to date, nothing is known about the toxic effects of these compounds on the embryonic development of aquatic invertebrate taxa, including freshwater mollusks. This study examines under laboratory conditions the toxicity of NAs extracted from oil sands process water (OSPW) and the low-molecular weight model NAs cyclohexylsuccinic acid (CHSA), cyclohexanebutyric acid (CHBA), and 4-tert-butylcyclohexane carboxylic acid (4-TBCA) on embryonic development of the snail Lymnaea stagnalis, a common freshwater gastropod with a broad Palearctic distribution. Evidence is provided for concentration-dependent teratogenic effects of both OSPW-derived and model NAs with remarkably similar nominal threshold concentrations between 15 and 20 mg/L and 28d EC 50 of 31 mg/L. In addition, the data provide evidence for substantial toxicokinetic differences between CHSA, CHBA and 4-TBCA. Together, our study introduces Lymnaea stagnalis embryonic development as an effective model to assay NA-toxicity and identifies molecular architecture as a potentially important toxicokinetic parameter in the toxicity of low-molecular weight NA in embryonic development of aquatic gastropods. Copyright © 2016 Elsevier Ltd. All rights reserved.

Engineering human cell spheroids to model embryonic tissue fusion in vitro

PubMed Central

Wolf, Cynthia J.; Wood, Carmen; Ren, Hongzu; Grindstaff, Rachel; Padgett, William; Swank, Adam; MacMillan, Denise; Fisher, Anna; Winnik, Witold; Abbott, Barbara D.

2017-01-01

Epithelial-mesenchymal interactions drive embryonic fusion events during development, and perturbations of these interactions can result in birth defects. Cleft palate and neural tube defects can result from genetic defects or environmental exposures during development, yet very little is known about the effect of chemical exposures on fusion events during human development because of a lack of relevant and robust human in vitro assays of developmental fusion behavior. Given the etiology and prevalence of cleft palate and the relatively simple architecture and composition of the embryonic palate, we sought to develop a three-dimensional culture system that mimics the embryonic palate and could be used to study fusion behavior in vitro using human cells. We engineered size-controlled human Wharton’s Jelly stromal cell (HWJSC) spheroids and established that 7 days of culture in osteogenesis differentiation medium was sufficient to promote an osteogenic phenotype consistent with embryonic palatal mesenchyme. HWJSC spheroids supported the attachment of human epidermal keratinocyte progenitor cells (HPEKp) on the outer spheroid surface likely through deposition of collagens I and IV, fibronectin, and laminin by mesenchymal spheroids. HWJSC spheroids coated in HPEKp cells exhibited fusion behavior in culture, as indicated by the removal of epithelial cells from the seams between spheroids, that was dependent on epidermal growth factor signaling and fibroblast growth factor signaling in agreement with palate fusion literature. The method described here may broadly apply to the generation of three-dimensional epithelial-mesenchymal co-cultures to study developmental fusion events in a format that is amenable to predictive toxicology applications. PMID:28898253

Intermolecular Interactions of Homologs of Germ Plasm Components in Mammalian Germ Cells

PubMed Central

Fox, Mark S.; Clark, Amander T.; El Majdoubi, Mohammed; Vigne, Jean-Louis; Urano, Jun; Hostetler, Chris E.; Griswold, Michael D.; Weiner, Richard I.; Pera, Renee A. Reijo

2007-01-01

In some species such as flies, worms, frogs, and fish the key to forming and maintaining early germ cell populations is the assembly of germ plasm, microscopically-distinct egg cytoplasm that is rich in RNAs, RNA-binding proteins and ribosomes. Cells which inherit germ plasm are destined for the germ cell lineage. In contrast, in mammals, germ cells are formed and maintained later in development as a result of inductive signaling from one embryonic cell type to another. Research advances, using complementary approaches, including identification of key signaling factors that act during the initial stages of germ cell development, differentiation of germ cells in vitro from mouse and human embryonic stem cells and the demonstration, that homologs of germ plasm components are conserved in mammals, have shed light on key elements in the early development of mammalian germ cells. Here, we use FRET (Fluorescence Resonance Energy Transfer) to demonstrate that living mammalian germ cells possess specific RNA/protein complexes that contain germ plasm homologs, beginning in the earliest stages of development examined. Moreover, we demonstrate that although both human and mouse germ cells and embryonic stem cells express the same proteins, germ cell specific protein/protein interactions distinguish germ cells from precursor embryonic stem cells in vitro; interactions also determine sub-cellular localization of complex components. Finally, we suggest that assembly of similar protein complexes may be central to differentiation of diverse cell lineages and provide useful diagnostic tools for isolation of specific cell types from the assorted types differentiated from embryonic stem cells. PMID:16996493

Transcriptional role of androgen receptor in the expression of long non-coding RNA Sox2OT in neurogenesis

PubMed Central

Tosetti, Valentina; Sassone, Jenny; Ferri, Anna L. M.; Taiana, Michela; Bedini, Gloria; Nava, Sara; Brenna, Greta; Di Resta, Chiara; Pareyson, Davide; Di Giulio, Anna Maria; Carelli, Stephana

2017-01-01

The complex architecture of adult brain derives from tightly regulated migration and differentiation of precursor cells generated during embryonic neurogenesis. Changes at transcriptional level of genes that regulate migration and differentiation may lead to neurodevelopmental disorders. Androgen receptor (AR) is a transcription factor that is already expressed during early embryonic days. However, AR role in the regulation of gene expression at early embryonic stage is yet to be determinate. Long non-coding RNA (lncRNA) Sox2 overlapping transcript (Sox2OT) plays a crucial role in gene expression control during development but its transcriptional regulation is still to be clearly defined. Here, using Bicalutamide in order to pharmacologically inactivated AR, we investigated whether AR participates in the regulation of the transcription of the lncRNASox2OTat early embryonic stage. We identified a new DNA binding region upstream of Sox2 locus containing three androgen response elements (ARE), and found that AR binds such a sequence in embryonic neural stem cells and in mouse embryonic brain. Our data suggest that through this binding, AR can promote the RNA polymerase II dependent transcription of Sox2OT. Our findings also suggest that AR participates in embryonic neurogenesis through transcriptional control of the long non-coding RNA Sox2OT. PMID:28704421

Transcriptional role of androgen receptor in the expression of long non-coding RNA Sox2OT in neurogenesis.

PubMed

Tosetti, Valentina; Sassone, Jenny; Ferri, Anna L M; Taiana, Michela; Bedini, Gloria; Nava, Sara; Brenna, Greta; Di Resta, Chiara; Pareyson, Davide; Di Giulio, Anna Maria; Carelli, Stephana; Parati, Eugenio A; Gorio, Alfredo

2017-01-01

The complex architecture of adult brain derives from tightly regulated migration and differentiation of precursor cells generated during embryonic neurogenesis. Changes at transcriptional level of genes that regulate migration and differentiation may lead to neurodevelopmental disorders. Androgen receptor (AR) is a transcription factor that is already expressed during early embryonic days. However, AR role in the regulation of gene expression at early embryonic stage is yet to be determinate. Long non-coding RNA (lncRNA) Sox2 overlapping transcript (Sox2OT) plays a crucial role in gene expression control during development but its transcriptional regulation is still to be clearly defined. Here, using Bicalutamide in order to pharmacologically inactivated AR, we investigated whether AR participates in the regulation of the transcription of the lncRNASox2OTat early embryonic stage. We identified a new DNA binding region upstream of Sox2 locus containing three androgen response elements (ARE), and found that AR binds such a sequence in embryonic neural stem cells and in mouse embryonic brain. Our data suggest that through this binding, AR can promote the RNA polymerase II dependent transcription of Sox2OT. Our findings also suggest that AR participates in embryonic neurogenesis through transcriptional control of the long non-coding RNA Sox2OT.

Functional video-based analysis of 3D cardiac structures generated from human embryonic stem cells.

PubMed

Nitsch, Scarlett; Braun, Florian; Ritter, Sylvia; Scholz, Michael; Schroeder, Insa S

2018-05-01

Human embryonic stem cells (hESCs) differentiated into cardiomyocytes (CM) often develop into complex 3D structures that are composed of various cardiac cell types. Conventional methods to study the electrophysiology of cardiac cells are patch clamp and microelectrode array (MEAs) analyses. However, these methods are not suitable to investigate the contractile features of 3D cardiac clusters that detach from the surface of the culture dishes during differentiation. To overcome this problem, we developed a video-based motion detection software relying on the optical flow by Farnebäck that we call cBRA (cardiac beat rate analyzer). The beating characteristics of the differentiated cardiac clusters were calculated based on the local displacement between two subsequent images. Two differentiation protocols, which profoundly differ in the morphology of cardiac clusters generated and in the expression of cardiac markers, were used and the resulting CM were characterized. Despite these differences, beat rates and beating variabilities could be reliably determined using cBRA. Likewise, stimulation of β-adrenoreceptors by isoproterenol could easily be identified in the hESC-derived CM. Since even subtle changes in the beating features are detectable, this method is suitable for high throughput cardiotoxicity screenings. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Markers of murine embryonic and neural stem cells, neurons and astrocytes: reference points for developmental neurotoxicity testing

EPA Science Inventory

Developmental neurotoxicity (DNT) is a significant concern for environmental chemicals, as well as for food and drug constituents. The sensitivity of animal-based DNT models is unclear, and they are expensive and time consuming. Murine embryonic stem cells (mESC) recapitulate sev...

Scaffolding for Three-Dimensional Embryonic Vasculogenesis

NASA Astrophysics Data System (ADS)

Kraehenbuehl, Thomas P.; Aday, Sezin; Ferreira, Lino S.

Biomaterial scaffolds have great potential to support efficient vascular differentiation of embryonic stem cells. Vascular cell fate-specific biochemical and biophysical cues have been identified and incorporated into three-dimensional (3D) biomaterials to efficiently direct embryonic vasculogenesis. The resulting vascular-like tissue can be used for regenerative medicine applications, further elucidation of biophysical and biochemical cues governing vasculogenesis, and drug discovery. In this chapter, we give an overview on the following: (1) developmental cues for directed differentiation of human embryonic stem cells (hESCs) into vascular cells, (2) 3D vascular differentiation in embryoid bodies (EBs), (3) preparation of 3D scaffolds for the vascular differentiation of hESCs, and (4) the most significant studies combining scaffolding and hESCs for development of vascular-like tissue.

Maternal Germline-Specific Genes in the Asian Malaria Mosquito Anopheles stephensi: Characterization and Application for Disease Control

PubMed Central

Biedler, James K.; Qi, Yumin; Pledger, David; Macias, Vanessa M.; James, Anthony A.; Tu, Zhijian

2014-01-01

Anopheles stephensi is a principal vector of urban malaria on the Indian subcontinent and an emerging model for molecular and genetic studies of mosquito biology. To enhance our understanding of female mosquito reproduction, and to develop new tools for basic research and for genetic strategies to control mosquito-borne infectious diseases, we identified 79 genes that displayed previtellogenic germline-specific expression based on RNA-Seq data generated from 11 life stage–specific and sex-specific samples. Analysis of this gene set provided insights into the biology and evolution of female reproduction. Promoters from two of these candidates, vitellogenin receptor and nanos, were used in independent transgenic cassettes for the expression of artificial microRNAs against suspected mosquito maternal-effect genes, discontinuous actin hexagon and myd88. We show these promoters have early germline-specific expression and demonstrate 73% and 42% knockdown of myd88 and discontinuous actin hexagon mRNA in ovaries 48 hr after blood meal, respectively. Additionally, we demonstrate maternal-specific delivery of mRNA and protein to progeny embryos. We discuss the application of this system of maternal delivery of mRNA/miRNA/protein in research on mosquito reproduction and embryonic development, and for the development of a gene drive system based on maternal-effect dominant embryonic arrest. PMID:25480960

Identification of compounds that modulate retinol signaling using a cell-based qHTS assay

PubMed Central

Chen, Yanling; Sakamuru, Srilatha; Huang, Ruili; Reese, David H.; Xia, Menghang

2016-01-01

In vertebrates, the retinol (vitamin A) signaling pathway (RSP) controls the biosynthesis and catabolism of all-trans retinoic acid (atRA), which regulates transcription of genes essential for embryonic development. Chemicals that interfere with the RSP to cause abnormal intracellular levels of atRA are potential developmental toxicants. To assess chemicals for the ability to interfere with retinol signaling, we have developed a cell-based RARE (Retinoic Acid Response Element) reporter gene assay to identify RSP disruptors. To validate this assay in a quantitative high-throughput screening (qHTS) platform, we screened the Library of Pharmacologically Active Compounds (LOPAC) in both agonist and antagonist modes. The screens detected known RSP agonists, demonstrating assay reliability, and also identified novel RSP agonists including kenpaullone, niclosamide, PD98059 and SU4312, and RSP antagonists including Bay 11-7085, LY294002, 3,4-Methylenedioxy-β-nitrostyrene, and topoisomerase inhibitors (camptothecin, topotecan, amsacrine hydrochloride, and idarubicin). When evaluated in the P19 pluripotent cell, these compounds were found to affect the expression of the Hoxa1 gene that is essential for embryo body patterning. These results show that the RARE assay is an effective qHTS approach for screening large compound libraries to identify chemicals that have the potential to adversely affect embryonic development through interference with retinol signaling. PMID:26820057

Comparative hazard analysis and toxicological modeling of diverse nanomaterials using the embryonic zebrafish (EZ) metric of toxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harper, Bryan; Thomas, Dennis G.; Chikkagoudar, Satish

The integration of rapid assays, large data sets, informatics and modeling can overcome current barriers in understanding nanomaterial structure-toxicity relationships by providing a weight-of-the-evidence mechanism to generate hazard rankings for nanomaterials. Here we present the use of a rapid, low-cost assay to perform screening-level toxicity evaluations of nanomaterials in vivo. Calculated EZ Metric scores, a combined measure of morbidity and mortality, were established at realistic exposure levels and used to develop a predictive model of nanomaterial toxicity. Hazard ranking and clustering analysis of 68 diverse nanomaterials revealed distinct patterns of toxicity related to both core composition and outermost surface chemistrymore » of nanomaterials. The resulting clusters guided the development of a predictive model of gold nanoparticle toxicity to embryonic zebrafish. In addition, our findings suggest that risk assessments based on the size and core composition of nanomaterials alone may be wholly inappropriate, especially when considering complex engineered nanomaterials. These findings reveal the need to expeditiously increase the availability of quantitative measures of nanomaterial hazard and broaden the sharing of that data and knowledge to support predictive modeling. In addition, research should continue to focus on methodologies for developing predictive models of nanomaterial hazard based on sub-lethal responses to low dose exposures.« less

Comparative hazard analysis and toxicological modeling of diverse nanomaterials using the embryonic zebrafish (EZ) metric of toxicity

DOE PAGES

Harper, Bryan; Thomas, Dennis G.; Chikkagoudar, Satish; ...

2015-06-04

The integration of rapid assays, large data sets, informatics and modeling can overcome current barriers in understanding nanomaterial structure-toxicity relationships by providing a weight-of-the-evidence mechanism to generate hazard rankings for nanomaterials. Here we present the use of a rapid, low-cost assay to perform screening-level toxicity evaluations of nanomaterials in vivo. Calculated EZ Metric scores, a combined measure of morbidity and mortality, were established at realistic exposure levels and used to develop a predictive model of nanomaterial toxicity. Hazard ranking and clustering analysis of 68 diverse nanomaterials revealed distinct patterns of toxicity related to both core composition and outermost surface chemistrymore » of nanomaterials. The resulting clusters guided the development of a predictive model of gold nanoparticle toxicity to embryonic zebrafish. In addition, our findings suggest that risk assessments based on the size and core composition of nanomaterials alone may be wholly inappropriate, especially when considering complex engineered nanomaterials. These findings reveal the need to expeditiously increase the availability of quantitative measures of nanomaterial hazard and broaden the sharing of that data and knowledge to support predictive modeling. In addition, research should continue to focus on methodologies for developing predictive models of nanomaterial hazard based on sub-lethal responses to low dose exposures.« less

Utilization of different anti-viral mechanisms by mammalian embryonic stem cells and differentiated cells.

PubMed

Guo, Yan-Lin

2017-01-01

Embryonic stem cells (ESCs) have received tremendous attention because of their potential applications in regenerative medicine. Over the past two decades, intensive research has not only led to the generation of various types of cells from ESCs that can be potentially used for the treatment of human diseases but also led to the formation of new concepts and breakthroughs that have significantly impacted our understanding of basic cell biology and developmental biology. Recent studies have revealed that ESCs and other types of pluripotent cells do not have a functional interferon (IFN)-based anti-viral mechanism, challenging the idea that the IFN system is developed as the central component of anti-viral innate immunity in all types of cells in vertebrates. This finding also provided important insight into a question that has been uncertain for a long time: whether or not the RNA interference (RNAi) anti-viral mechanism operates in mammalian cells. An emerging paradigm is that mammals may have adapted distinct anti-viral mechanisms at different stages of organismal development; the IFN-based system is mainly used by differentiated somatic cells, while the RNAi anti-viral mechanism may be used in ESCs. This paper discusses the molecular basis and biological implications for mammals to have different anti-viral mechanisms during development.

An unbiased approach to identify genes involved in development in a turtle with temperature-dependent sex determination.

PubMed

Chojnowski, Jena L; Braun, Edward L

2012-07-15

Many reptiles exhibit temperature-dependent sex determination (TSD). The initial cue in TSD is incubation temperature, unlike genotypic sex determination (GSD) where it is determined by the presence of specific alleles (or genetic loci). We used patterns of gene expression to identify candidates for genes with a role in TSD and other developmental processes without making a priori assumptions about the identity of these genes (ortholog-based approach). We identified genes with sexually dimorphic mRNA accumulation during the temperature sensitive period of development in the Red-eared slider turtle (Trachemys scripta), a turtle with TSD. Genes with differential mRNA accumulation in response to estrogen (estradiol-17β; E(2)) exposure and developmental stages were also identified. Sequencing 767 clones from three suppression-subtractive hybridization libraries yielded a total of 581 unique sequences. Screening a macroarray with a subset of those sequences revealed a total of 26 genes that exhibited differential mRNA accumulation: 16 female biased and 10 male biased. Additional analyses revealed that C16ORF62 (an unknown gene) and MALAT1 (a long noncoding RNA) exhibited increased mRNA accumulation at the male producing temperature relative to the female producing temperature during embryonic sexual development. Finally, we identified four genes (C16ORF62, CCT3, MMP2, and NFIB) that exhibited a stage effect and five genes (C16ORF62, CCT3, MMP2, NFIB and NOTCH2) showed a response to E(2) exposure. Here we report a survey of genes identified using patterns of mRNA accumulation during embryonic development in a turtle with TSD. Many previous studies have focused on examining the turtle orthologs of genes involved in mammalian development. Although valuable, the limitations of this approach are exemplified by our identification of two genes (MALAT1 and C16ORF62) that are sexually dimorphic during embryonic development. MALAT1 is a noncoding RNA that has not been implicated in sexual differentiation in other vertebrates and C16ORF62 has an unknown function. Our results revealed genes that are candidates for having roles in turtle embryonic development, including TSD, and highlight the need to expand our search parameters beyond protein-coding genes.

Deficiencies in the uterine environment and failure to support embryo development

USDA-ARS?s Scientific Manuscript database

Pregnancy failure in livestock can result from failure to fertilize the oocyte or embryonic loss during gestation. Although fertilization failure occurs, embryonic mortality has a greater contribution to pregnancy failure. The focus of this review is on cattle and factors affecting, and mechanisms r...

AN EMBRYONIC CHICK PANCREAS ORGAN CULTURE MODEL: CHARACTERIZATION AND NEURAL CONTROL OF EXOCRINE RELEASE

EPA Science Inventory

An embryonic chick (Gallus domesticus) whole-organ pancreas culture system was developed for use as an in vitro model to study cholinergic regulation of exocrine pancreatic function. The culture system was examined for characteristic exocrine function and viability by measuring e...

Embryonic development of the grass pufferfish (Takifugu niphobles): From egg to larvae.

PubMed

Gallego, V; Yoshida, M; Kurokawa, D; Asturiano, J F; Fraser, G J

2017-03-01

Tetraodontidae (pufferfish) family members carry the smallest genomes among vertebrates, and these pocket-sized genomes have directly contributed to our understanding of the structure and evolution of higher animals. The grass pufferfish (Takifugu niphobles) could be considered a potential new model organism for comparative genomics and development due to the potential access to embryos, and availability of sequence data for two similar genomes: that of spotted green pufferfish (Tetraodon nigroviridis) and Fugu (Takifugu rubripes). In this study, we provide the first description of the normal embryonic development of T. niphobles, by drawing comparisons with the closely related species cited above. Embryos were obtained by in vitro fertilization of eggs, and subsequent development was monitored at a constant temperature consistent with natural conditions. T. niphobles development was divided into seven periods of embryogenesis: the zygote, cleavage, blastula, gastrula, segmentation, pharyngula, and hatching periods; and stages subdividing these periods are defined based on morphological characteristics. The developmental stage series described in this study aims to provide the utilization of T. niphobles as an experimental model organism for comparative developmental studies. Copyright © 2016 Elsevier Inc. All rights reserved.

Microscopic analysis of Spodoptera frugiperda (Lepidoptera: Noctuidae) embryonic development before and after treatment with azadirachtin, lufenuron, and deltamethrin.

PubMed

Correia, Alicely A; Wanderley-Teixeira, Valéria; Teixeira, Alvaro A C; Oliveira, José V; Gonçalves, Gabriel G A; Cavalcanti, MaríIia G S; Brayner, Fábio A; Alves, Luiz C

2013-04-01

The botanical insecticides, growth regulators, and pyrethroids have an effect on the biology of Spodoptera frugiperda (Smith). However, no emphasis has been given to the effect of these insecticides on embryonic development of insects, in histological level. Thus, this research aimed to examine by light and scanning electron microscopy S. frugiperda eggs and to describe the embryonic development, before and after immersion treatment, using commercial concentrations and lower concentrations than commercial ones, of the compounds lufenuron (Match), azadirachtin (AzaMax), and deltamethrin (Decis-positive control). For light microscopy semithin sections of eggs were used, and for scanning electron microscopy, images of the surface of eggs, treated and untreated with insecticides. The morphological characteristics of S. frugiperda eggs, in general, were similar to those described in the literature for most of the insects in the order Lepidoptera. Spherical eggs slightly flattened at the poles, with chorion, yolk, vitelline membrane, and embryo formation. In both microscopic analysis, we observed that insecticides acted immediately and independent of concentration, resulting absence, or incomplete embryo, presented yolk granules widely dispersed, without vitellophage formation, chorion disintegration, disorganized blastoderm, presenting vacuoles, yolk region with amorphous cells, and formation of completely uncharacterized appendages. Thus, we conclude that the compounds lufenuron and azadirachtin interfere on S. frugiperda embryonic development.

Knockdown of Fanconi anemia genes in human embryonic stem cells reveals early developmental defects in the hematopoietic lineage.

PubMed

Tulpule, Asmin; Lensch, M William; Miller, Justine D; Austin, Karyn; D'Andrea, Alan; Schlaeger, Thorsten M; Shimamura, Akiko; Daley, George Q

2010-04-29

Fanconi anemia (FA) is a genetically heterogeneous, autosomal recessive disorder characterized by pediatric bone marrow failure and congenital anomalies. The effect of FA gene deficiency on hematopoietic development in utero remains poorly described as mouse models of FA do not develop hematopoietic failure and such studies cannot be performed on patients. We have created a human-specific in vitro system to study early hematopoietic development in FA using a lentiviral RNA interference (RNAi) strategy in human embryonic stem cells (hESCs). We show that knockdown of FANCA and FANCD2 in hESCs leads to a reduction in hematopoietic fates and progenitor numbers that can be rescued by FA gene complementation. Our data indicate that hematopoiesis is impaired in FA from the earliest stages of development, suggesting that deficiencies in embryonic hematopoiesis may underlie the progression to bone marrow failure in FA. This work illustrates how hESCs can provide unique insights into human development and further our understanding of genetic disease.

Modulation of ovarian steroidogenesis by adiponectin during delayed embryonic development of Cynopterus sphinx.

PubMed

Anuradha; Krishna, Amitabh

2014-09-01

The aim of present study was to evaluate role of adiponectin in ovarian steroidogenesis during delayed embryonic development of Cynopterus sphinx. This study showed significantly low circulating adiponectin level and a decline in expression of adiponectin receptor 1 (AdipoR1) in the ovary during the period of delayed embryonic development as compared with the normal development. The adiponectin treatment in vivo during the period of delayed development caused significantly increased in circulating progesterone and estradiol levels together with increased expression of AdipoR1 in the ovary. The in vitro study confirmed the stimulatory effect of adiponectin on progesterone synthesis. Both in vivo and in vitro studies showed that the effects of adiponectin on ovarian steroidogenesis were mediated through increased expression of luteinizing hormone-receptor, steroidogenic acute regulatory protein and 3β-hydroxyl steroid dehydrogenase enzyme. The adiponectin treatment may also promote progesterone synthesis by modulating ovarian angiogenesis, cell survival and rate of apoptosis. Copyright © 2014 Elsevier Ltd. All rights reserved.

A staging table for the embryonic development of the brownbanded bamboo shark (Chiloscyllium punctatum)

PubMed Central

Onimaru, Koh; Motone, Fumio; Kiyatake, Itsuki; Nishida, Kiyonori

2018-01-01

Background: Studying cartilaginous fishes (chondrichthyans) has helped us understand vertebrate evolution and diversity. However, resources such as genome sequences, embryos, and detailed staging tables are limited for species within this clade. To overcome these limitations, we have focused on a species, the brownbanded bamboo shark (Chiloscyllium punctatum), which is a relatively common aquarium species that lays eggs continuously throughout the year. In addition, because of its relatively small genome size, this species is promising for molecular studies. Results: To enhance biological studies of cartilaginous fishes, we establish a normal staging table for the embryonic development of the brownbanded bamboo shark. Bamboo shark embryos take around 118 days to reach the hatching period at 25°C, which is approximately 1.5 times as fast as the small‐spotted catshark (Scyliorhinus canicula) takes. Our staging table divides the embryonic period into 38 stages. Furthermore, we found culture conditions that allow early embryos to grow in partially opened egg cases. Conclusions: In addition to the embryonic staging table, we show that bamboo shark embryos exhibit relatively fast embryonic growth and are amenable to culture, key characteristics that enhance their experimental utility. Therefore, the present study is a foundation for cartilaginous fish research. Developmental Dynamics 247:712–723, 2018. © 2017 Wiley Periodicals, Inc. PMID:29396887

The embryonic origin of the ampullate silk glands of the spider Cupiennius salei.

PubMed

Hilbrant, Maarten; Damen, Wim G M

2015-05-01

Silk production in spiders is considered a key innovation, and to have been vital for the diversification of the clade. The evolutionary origin of the organs involved in spider silk production, however, and in particular of the silk glands, is poorly understood. Homologies have been proposed between these and other glands found in arachnids, but lacking knowledge of the embryonic development of spider silk glands hampers an evaluation of hypotheses. This study focuses on the embryonic origin of the largest silk glands of the spider Cupiennius salei, the major and minor ampullate glands. We show how the ampullate glands originate from ectodermal invaginations on the embryonic spinneret limb buds, in relation to morphogenesis of these buds. Moreover, we visualize the subsequent growth of the ampullate glands in sections of the early postembryonic stages. The invaginations are shown to correlate with expression of the proneural gene CsASH2, which is remarkable since it has been proposed that spider silk glands and their nozzles originate from sensory bristles. Hence, by confirming the ectodermal origin of spider silk glands, and by describing the (post-)embryonic morphogenesis of the ampullate glands, this work provides a starting point for further investigating into the genetic program that underlies their development. Copyright © 2015 Elsevier Ltd. All rights reserved.

Autophagy in Human Embryonic Stem Cells

PubMed Central

Tra, Thien; Gong, Lan; Kao, Lin-Pin; Li, Xue-Lei; Grandela, Catarina; Devenish, Rodney J.; Wolvetang, Ernst; Prescott, Mark

2011-01-01

Autophagy (macroautophagy) is a degradative process that involves the sequestration of cytosolic material including organelles into double membrane vesicles termed autophagosomes for delivery to the lysosome. Autophagy is essential for preimplantation development of mouse embryos and cavitation of embryoid bodies. The precise roles of autophagy during early human embryonic development, remain however largely uncharacterized. Since human embryonic stem cells constitute a unique model system to study early human embryogenesis we investigated the occurrence of autophagy in human embryonic stem cells. We have, using lentiviral transduction, established multiple human embryonic stem cell lines that stably express GFP-LC3, a fluorescent marker for the autophagosome. Each cell line displays both a normal karyotype and pluripotency as indicated by the presence of cell types representative of the three germlayers in derived teratomas. GFP expression and labelling of autophagosomes is retained after differentiation. Baseline levels of autophagy detected in cultured undifferentiated hESC were increased or decreased in the presence of rapamycin and wortmannin, respectively. Interestingly, autophagy was upregulated in hESCs induced to undergo differentiation by treatment with type I TGF-beta receptor inhibitor SB431542 or removal of MEF secreted maintenance factors. In conclusion we have established hESCs capable of reporting macroautophagy and identify a novel link between autophagy and early differentiation events in hESC. PMID:22110659

In Situ Histochemical Localisation of Alkaloids and Acetogenins in the Endosperm and Embryonic Axis of Annona Macroprophyllata Donn. Sm. Seeds During Germination

PubMed Central

Brechú-Franco, A.E.; Laguna-Hernández, G.; De la Cruz-Chacón, I.; González-Esquinca, A.R.

2016-01-01

Currently, the Annonaceae family is characterised by the production of acetogenins (ACGs), and also by the biosynthesis of alkaloids, primarily benzylisoquinolines derived from tyrosine. The objective of this study was to confirm the presence of alkaloids and acetogenins in the idioblasts of the endosperm and the embryonic axis of A. macroprophyllata seeds in germination. The Dragendorff, Dittmar, Ellram, and Lugol reagents were used to test for alkaloids, and Kedde’s reagent was used to determine the presence of acetogenins in fresh sections of the endosperm and embryonic axis of seeds after twelve days of germination. A positive reaction was observed for all the reagents, and the presence of alkaloids and acetogenins was confirmed in the idioblasts of the endosperm and those involved in the differentiation of the embryonic axis of the developing seedling. We concluded that the idioblasts store both metabolites, acetogenins and alkaloids. Beginning at differentiation, the idioblasts of the embryonic axis simultaneously biosynthesise acetogenins and alkaloids that are characteristic of the species during the development of the seedling. The method used here can be applied to histochemically confirm the presence of acetogenins and alkaloids in tissues and structures of the plant in different stages of its life cycle. PMID:26972713

Quantitation of two endogenous lactose-inhibitable lectins in embryonic and adult chicken tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Beyer, E.C.; Barondes, S.H.

Two lactose-binding lectins from chicken tissues, chicken-lactose-lectin-I (CLL-I) and chicken-lactose-lectin-II (CLL-II) were quantified with a radioimmunoassay in extracts of a number of developing and adult chicken tissues. Both lectins could be measured in the same extract without separation, because they showed no significant immunological cross- reactivity. Many embryonic and adult tissues, including brain, heart, intestine, kidney, liver, lung, muscle, pancreas, and spleen, contained one or both lectins, although their concentrations differed markedly. For example, embryonic muscle, the richest source of CLL-I contained only traces of CLL-II whereas embryonic kidney, a very rich source of CLL-II contained substantial CLL-I. In bothmore » muscle and kidney, lectin levels in adulthood were much lower than in the embryonic state. In contrast, CLL-I in liver and CLL-II in intestine were 10-fold to 30-fold more concentrated in the adult than in the 15-d embryo. CLL-I and CLL-II from several tissues were purified by affinity chromatography and their identity in the various tissues was confirmed by polyacrylamide gel electrophoresis, isoelectric focusing, and peptide mapping. The results suggest that these lectins might have different functions in the many developing and adult tissues in which they are found.« less

Optical coherence microscopy as a novel, non-invasive method for the 4D live imaging of early mammalian embryos.

PubMed

Karnowski, Karol; Ajduk, Anna; Wieloch, Bartosz; Tamborski, Szymon; Krawiec, Krzysztof; Wojtkowski, Maciej; Szkulmowski, Maciej

2017-06-23

Imaging of living cells based on traditional fluorescence and confocal laser scanning microscopy has delivered an enormous amount of information critical for understanding biological processes in single cells. However, the requirement for a high numerical aperture and fluorescent markers still limits researchers' ability to visualize the cellular architecture without causing short- and long-term photodamage. Optical coherence microscopy (OCM) is a promising alternative that circumvents the technical limitations of fluorescence imaging techniques and provides unique access to fundamental aspects of early embryonic development, without the requirement for sample pre-processing or labeling. In the present paper, we utilized the internal motion of cytoplasm, as well as custom scanning and signal processing protocols, to effectively reduce the speckle noise typical for standard OCM and enable high-resolution intracellular time-lapse imaging. To test our imaging system we used mouse and pig oocytes and embryos and visualized them through fertilization and the first embryonic division, as well as at selected stages of oogenesis and preimplantation development. Because all morphological and morphokinetic properties recorded by OCM are believed to be biomarkers of oocyte/embryo quality, OCM may represent a new chapter in imaging-based preimplantation embryo diagnostics.

Self-organization of vertebrate mesoderm based on simple boundary conditions.

PubMed

Green, Jeremy B A; Dominguez, Isabel; Davidson, Lance A

2004-11-01

Embryonic development requires cell movements whose coordination is robust and reproducible. A dramatic example is the primary body axis of vertebrates: despite perturbation, cells in prospective axial tissue coordinate their movements to make an elongated body axis. The spatial cues coordinating these movements are not known. We show here that cells deprived of preexisting spatial cues by physical dissociation and reaggregation nonetheless organize themselves into an axis. Activin-induced cells that are reaggregated into a flat disc initially round up into a ball before elongating perpendicular to the disc. Manipulations of the geometry of the disc and immunofluorescence micrography reveal that the edge of the disc provides a circumferential alignment zone. This finding indicates that physical boundaries provide alignment cues and that circumferential "hoop stress" drives the axial extrusion in a manner resembling late-involuting mesoderm of Xenopus and archenteron elongation in other deuterostome species such as sea urchins. Thus, a population of cells finds its own midline based on the form of the population's boundaries using an edge-aligning mechanism. This process provides a remarkably simple organizing principle that contributes to the reliability of embryonic development as a whole. (c) 2004 Wiley-Liss, Inc.

Impacts of chemical modification on the toxicity of diverse nanocellulose materials to developing zebrafish

PubMed Central

Harper, Bryan J.; Clendaniel, Alicea; Sinche, Federico; Way, Daniel; Hughes, Michael; Schardt, Jenna; Simonsen, John; Stefaniak, Aleksandr B.

2016-01-01

Cellulose is an abundant and renewable resource currently being investigated for utility in nanomaterial form for various promising applications ranging from medical and pharmaceutical uses to mechanical reinforcement and biofuels. The utility of nanocellulose and wide implementation ensures increasing exposure to humans and the environment as nanocellulose-based technologies advance. Here, we investigate how differences in aspect ratio and changes to surface chemistry, as well as synthesis methods, influence the biocompatibility of nanocellulose materials using the embryonic zebrafish. Investigations into the toxicity of neutral, cationic and anionic surface functionalities revealed that surface chemistry had a minimal influence on the overall toxicity of nanocellulose materials. Higher aspect ratio cellulose nanofibers produced by mechanical homogenization were, in some cases, more toxic than other cellulose-based nanofibers or nanocrystals produced by chemical synthesis methods. Using fluorescently labeled nanocellulose we were able to show that nanocellulose uptake did occur in embryonic zebrafish during development. We conclude that the benign nature of nanocellulose materials makes them an ideal platform to systematically investigate the inherent surface features driving nanomaterial toxicity in order to create safer design principles for engineered nanoparticles. PMID:27468180

Paternal identity impacts embryonic development for two species of freshwater fish.

PubMed

Siddique, Mohammad Abdul Momin; Linhart, Otomar; Krejszeff, Sławomir; Żarski, Daniel; Pitcher, Trevor E; Politis, Sebastian Nikitas; Butts, Ian Anthony Ernest

2017-05-01

Paternal, compared to maternal, contributions were believed to have only a limited influence on embryonic development and larval fitness traits in fishes. Therefore, the perspective of male influence on early life history traits has come under scrutiny. This study was conducted to determine parental effects on the rate of eyed embryos of Ide Leuciscus idus and Northern pike Esox lucius. Five sires and five dams from each species were crossed using a quantitative genetic breeding design and the resulting 25 sib groups of each species were reared to the embryonic eyed stage. We then partition variation in embryonic phenotypic performance to maternal, paternal, and parental interactions using the Restricted Maximum Likelihood (REML) model. Results showed that paternal, maternal, and the paternal×maternal interaction terms were highly significant for both species; clearly demonstrating that certain family combinations were more compatible than others. Paternal effects explained 20.24% of the total variance, which was 2-fold higher than the maternal effects (10.73%) in Ide, while paternal effects explained 18.9% of the total variance, which was 15-fold higher than the maternal effects (1.3%) in Northern pike. Together, these results indicate that male effects are of major importance during embryonic development for these species. Furthermore, this study demonstrates that genetic compatibility between sires and dams plays an important role and needs to be taken into consideration for reproduction of these and likely other economically important fish species. Copyright © 2016 Elsevier Inc. All rights reserved.

Fundal variations in the eyes of the osteoglossomorph fishes.

PubMed

Saidel, W M; Braford, M R

1985-01-01

The appearance of the fundus of the eye varies among the families of osteoglossomorph teleosts. In this study, four different fundal patterns were observed: (i) an anteroposterior (or horizontal) embryonic fissure with a septal falciform process (family Osteoglossidae); (ii) an embryonic fissure from the optic disc nasoventrally with a falciform process (family Arapaimidae); (iii) an embryonic fissure nasoventrally from the optic disc without a falciform process (family Hiodontidae); and (iv) neither an embryonic fissure nor a falciform process (families Notopteridae and Mormyridae). The distribution of these various forms among the osteoglossomorph fishes is consistent with the recent cladogram for the Osteoglossomorpha [Lauder and Liem, 1983] which was based on many characters. The embryonic fissure in adult Amia calva was also examined. Its existence in adult Amia, in most Osteoglossomorpha, and in many non-euteleostean bony fishes suggests that its persistence in the adult stage is a primitive trait of bony fishes, and its absence in the Notopteroidei (with the exception of Hiodon) is a derived condition.

The Evolution of Lineage-Specific Regulatory Activities in the Human Embryonic Limb

PubMed Central

Cotney, Justin; Leng, Jing; Yin, Jun; Reilly, Steven K.; DeMare, Laura E.; Emera, Deena; Ayoub, Albert E.; Rakic, Pasko; Noonan, James P.

2013-01-01

SUMMARY The evolution of human anatomical features likely involved changes in gene regulation during development. However, the nature and extent of human-specific developmental regulatory functions remain unknown. We obtained a genome-wide view of cis-regulatory evolution in human embryonic tissues by comparing the histone modification H3K27ac, which provides a quantitative readout of promoter and enhancer activity, during human, rhesus, and mouse limb development. Based on increased H3K27ac, we find that 13% of promoters and 11% of enhancers have gained activity on the human lineage since the human-rhesus divergence. These gains largely arose by modification of ancestral regulatory activities in the limb or potential co-option from other tissues and are likely to have heterogeneous genetic causes. Most enhancers that exhibit gain of activity in humans originated in mammals. Gains at promoters and enhancers in the human limb are associated with increased gene expression, suggesting they include molecular drivers of human morphological evolution. PMID:23827682

Large-scale time-lapse microscopy of Oct4 expression in human embryonic stem cell colonies.

PubMed

Bhadriraju, Kiran; Halter, Michael; Amelot, Julien; Bajcsy, Peter; Chalfoun, Joe; Vandecreme, Antoine; Mallon, Barbara S; Park, Kye-Yoon; Sista, Subhash; Elliott, John T; Plant, Anne L

2016-07-01

Identification and quantification of the characteristics of stem cell preparations is critical for understanding stem cell biology and for the development and manufacturing of stem cell based therapies. We have developed image analysis and visualization software that allows effective use of time-lapse microscopy to provide spatial and dynamic information from large numbers of human embryonic stem cell colonies. To achieve statistically relevant sampling, we examined >680 colonies from 3 different preparations of cells over 5days each, generating a total experimental dataset of 0.9 terabyte (TB). The 0.5 Giga-pixel images at each time point were represented by multi-resolution pyramids and visualized using the Deep Zoom Javascript library extended to support viewing Giga-pixel images over time and extracting data on individual colonies. We present a methodology that enables quantification of variations in nominally-identical preparations and between colonies, correlation of colony characteristics with Oct4 expression, and identification of rare events. Copyright © 2016. Published by Elsevier B.V.

How frequent and important is behavioral thermoregulation by embryonic reptiles?

PubMed

Shine, Richard; Du, Wei-Guo

2018-03-22

The debate about behavioral thermoregulation inside reptile eggs centers on the frequency (and hence, biological significance) of the phenomenon, not about its validity. Both sides of the debate agree that large eggs in shallow nests laid in sun-exposed soil will experience clines in mean temperature and (especially) diel thermal variance; that embryos in the middle phase of development have the ability to reposition themselves, and room to do so; and that small changes in developmental temperatures can influence offspring fitness. Equally, all protagonists agree that thermal clines will be too low in some other kinds of nests, and that embryonic repositioning is impossible very early and very late in development. Based on an array of other fitness-enhancing behaviors exhibited by tetrapod embryos, and general principles for recognizing adaptation, we conclude that behavioral thermoregulation inside the egg likely is adaptive in some but not all reptile species. We identify productive directions for empirical research to resolve points of contention. © 2018 Wiley Periodicals, Inc.

Spatial transcriptomic survey of human embryonic cerebral cortex by single-cell RNA-seq analysis.

PubMed

Fan, Xiaoying; Dong, Ji; Zhong, Suijuan; Wei, Yuan; Wu, Qian; Yan, Liying; Yong, Jun; Sun, Le; Wang, Xiaoye; Zhao, Yangyu; Wang, Wei; Yan, Jie; Wang, Xiaoqun; Qiao, Jie; Tang, Fuchou

2018-06-04

The cellular complexity of human brain development has been intensively investigated, although a regional characterization of the entire human cerebral cortex based on single-cell transcriptome analysis has not been reported. Here, we performed RNA-seq on over 4,000 individual cells from 22 brain regions of human mid-gestation embryos. We identified 29 cell sub-clusters, which showed different proportions in each region and the pons showed especially high percentage of astrocytes. Embryonic neurons were not as diverse as adult neurons, although they possessed important features of their destinies in adults. Neuron development was unsynchronized in the cerebral cortex, as dorsal regions appeared to be more mature than ventral regions at this stage. Region-specific genes were comprehensively identified in each neuronal sub-cluster, and a large proportion of these genes were neural disease related. Our results present a systematic landscape of the regionalized gene expression and neuron maturation of the human cerebral cortex.

Melatonin Inhibits Embryonic Salivary Gland Branching Morphogenesis by Regulating Both Epithelial Cell Adhesion and Morphology

PubMed Central

Miura, Jiro; Sakai, Manabu; Uchida, Hitoshi; Nakamura, Wataru; Nohara, Kanji; Maruyama, Yusuke; Hattori, Atsuhiko; Sakai, Takayoshi

2015-01-01

Many organs, including salivary glands, lung, and kidney, are formed by epithelial branching during embryonic development. Branching morphogenesis occurs via either local outgrowths or the formation of clefts that subdivide epithelia into buds. This process is promoted by various factors, but the mechanism of branching morphogenesis is not fully understood. Here we have defined melatonin as a potential negative regulator or “brake” of branching morphogenesis, shown that the levels of it and its receptors decline when branching morphogenesis begins, and identified the process that it regulates. Melatonin has various physiological functions, including circadian rhythm regulation, free-radical scavenging, and gonadal development. Furthermore, melatonin is present in saliva and may have an important physiological role in the oral cavity. In this study, we found that the melatonin receptor is highly expressed on the acinar epithelium of the embryonic submandibular gland. We also found that exogenous melatonin reduces salivary gland size and inhibits branching morphogenesis. We suggest that this inhibition does not depend on changes in either proliferation or apoptosis, but rather relates to changes in epithelial cell adhesion and morphology. In summary, we have demonstrated a novel function of melatonin in organ formation during embryonic development. PMID:25876057

MiRNA-mediated regulation of cell signaling and homeostasis in the early mouse embryo.

PubMed

Pernaute, Barbara; Spruce, Thomas; Rodriguez, Tristan A; Manzanares, Miguel

2011-02-15

At the time of implantation the mouse embryo is composed of three tissues the epiblast, trophectoderm and primitive endoderm. As development progresses the epiblast goes on to form the foetus whilst the trophectoderm and primitive endoderm give rise to extra-embryonic structures with important roles in embryo patterning and nutrition. Dramatic changes in gene expression occur during early embryo development and these require regulation at different levels. miRNAs are small non coding RNAs that have emerged over the last decade as important post-transcriptional repressors of gene expression. The roles played by miRNAs during early mammalian development are only starting to be elucidated. In order to gain insight into the function of miRNAs in the different lineages of the early mouse embryo we have analysed in depth the phenotype of embryos and extra-embryonic stem cells mutant for the miRNA maturation protein Dicer. This study revealed that miRNAs are involved in regulating cell signaling and homeostasis in the early embryo. Specifically, we identified a role for miRNAs in regulating the Erk signaling pathway in the extra-embryonic endoderm, cell cycle progression in extra-embryonic tissues and apoptosis in the epiblast.

Expression of the Drosophila homeobox gene, Distal-less supports an ancestral role in neural development

PubMed Central

Plavicki, Jessica S.; Squirrell, Jayne M.; Eliceiri, Kevin W.; Boekhoff-Falk, Grace

2015-01-01

Background Distal-less (Dll) encodes a homeodomain transcription factor expressed in developing appendages of organisms throughout metazoan phylogeny. Based on earlier observations in the limbless nematode Caenorhabditis elegans and the primitive chordate amphioxus, it was proposed that Dll had an ancestral function in nervous system development. Consistent with this hypothesis, Dll is necessary for the development of both peripheral and central components of the Drosophila olfactory system. Furthermore, vertebrate homologs of Dll, the Dlx genes, play critical roles in mammalian brain development. Results Using fluorescent immunohistochemistry of fixed samples and multiphoton microscopy of living Drosophila embryos we show that Dll is expressed in the embryonic, larval and adult CNS and PNS in embryonic and larval neurons, brain and ventral nerve cord (VNC) glia, as well as in PNS structures associated with chemosensation. In adult flies, Dll expression is expressed in the optic lobes, central brain regions and the antennal lobes. Conclusions Characterization of Dll expression in the developing nervous system supports a role of Dll in neural development and function and establishes an important basis for determining the specific functional roles of Dll in Drosophila development and for comparative studies of Drosophila Dll functions with those of its vertebrate counterparts. PMID:26472170

Genome editing reveals a role for OCT4 in human embryogenesis.

PubMed

Fogarty, Norah M E; McCarthy, Afshan; Snijders, Kirsten E; Powell, Benjamin E; Kubikova, Nada; Blakeley, Paul; Lea, Rebecca; Elder, Kay; Wamaitha, Sissy E; Kim, Daesik; Maciulyte, Valdone; Kleinjung, Jens; Kim, Jin-Soo; Wells, Dagan; Vallier, Ludovic; Bertero, Alessandro; Turner, James M A; Niakan, Kathy K

2017-10-05

Despite their fundamental biological and clinical importance, the molecular mechanisms that regulate the first cell fate decisions in the human embryo are not well understood. Here we use CRISPR-Cas9-mediated genome editing to investigate the function of the pluripotency transcription factor OCT4 during human embryogenesis. We identified an efficient OCT4-targeting guide RNA using an inducible human embryonic stem cell-based system and microinjection of mouse zygotes. Using these refined methods, we efficiently and specifically targeted the gene encoding OCT4 (POU5F1) in diploid human zygotes and found that blastocyst development was compromised. Transcriptomics analysis revealed that, in POU5F1-null cells, gene expression was downregulated not only for extra-embryonic trophectoderm genes, such as CDX2, but also for regulators of the pluripotent epiblast, including NANOG. By contrast, Pou5f1-null mouse embryos maintained the expression of orthologous genes, and blastocyst development was established, but maintenance was compromised. We conclude that CRISPR-Cas9-mediated genome editing is a powerful method for investigating gene function in the context of human development.

Role of acetylcholine receptors in proliferation and differentiation of P19 embryonal carcinoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Resende, R.R.; Alves, A.S.; Britto, L.R.G

2008-04-15

Coordinated proliferation and differentiation of progenitor cells is the base for production of appropriate numbers of neurons and glia during neuronal development in order to establish normal brain functions. We have used murine embryonal carcinoma P19 cells as an in vitro model for early differentiation to study participation of nicotinic (nAChR) and muscarinic acetylcholine (mAChR) receptors in the proliferation of neural progenitor cells and their differentiation to neurons. We have previously shown that functional nicotinic acetylcholine receptors (nAChRs) already expressed in embryonic cells mediate elevations in cytosolic free calcium concentration ([Ca{sup 2+}]{sub i}) via calcium influx through nAChR channels whereasmore » intracellular stores contribute to nAChR- and mAChR-mediated calcium fluxes in differentiated cells [Resende et al., Cell Calcium 43 (2008) 107-121]. In the present study, we have demonstrated that nicotine provoked inhibition of proliferation in embryonic cells as determined by BrdU labeling. However, in neural progenitor cells nicotine stimulated proliferation which was reversed in the presence of inhibitors of calcium mobilization from intracellular stores, indicating that liberation of intracellular calcium contributed to this proliferation induction. Muscarine induced proliferation stimulation in progenitor cells by activation of G{alpha}{sub q/11}-coupled M{sub 1}, M{sub 3} and M{sub 5} receptors and intracellular calcium stores, whereas G{alpha}{sub i/o}-protein coupled M{sub 2} receptor activity mediated neuronal differentiation.« less

FACS-based isolation, propagation and characterization of mouse embryonic cardiomyocytes based on VCAM-1 surface marker expression.

PubMed

Pontén, Annica; Walsh, Stuart; Malan, Daniela; Xian, Xiaojie; Schéele, Susanne; Tarnawski, Laura; Fleischmann, Bernd K; Jovinge, Stefan

2013-01-01

Purification of cardiomyocytes from the embryonic mouse heart, embryonic stem (ES) or induced pluripotent stem cells (iPS) is a challenging task and will require specific isolation procedures. Lately the significance of surface markers for the isolation of cardiac cell populations with fluorescence activated cell sorting (FACS) has been acknowledged, and the hunt for cardiac specific markers has intensified. As cardiomyocytes have traditionally been characterized by their expression of specific transcription factors and structural proteins, and not by specific surface markers, this constitutes a significant bottleneck. Lately, Flk-1, c-kit and the cellular prion protein have been reported to specify cardiac progenitors, however, no surface markers have so far been reported to specify a committed cardiomyocyte. Herein show for the first time, that embryonic cardiomyocytes can be isolated with 98% purity, based on their expression of vascular cell adhesion molecule-1 (VCAM-1). The FACS-isolated cells express phenotypic markers for embryonic committed cardiomyocytes but not cardiac progenitors. An important aspect of FACS is to provide viable cells with retention of functionality. We show that VCAM-1 positive cardiomyocytes can be isolated with 95% viability suitable for in vitro culture, functional assays or expression analysis. In patch-clamp experiments we provide evidence of functionally intact cardiomyocytes of both atrial and ventricular subtypes. This work establishes that cardiomyocytes can be isolated with a high degree of purity and viability through FACS, based on specific surface marker expression as has been done in the hematopoietic field for decades. Our FACS protocol represents a significant advance in which purified populations of cardiomyocytes may be isolated and utilized for downstream applications, such as purification of ES-cell derived cardiomyocytes.

FACS-Based Isolation, Propagation and Characterization of Mouse Embryonic Cardiomyocytes Based on VCAM-1 Surface Marker Expression

PubMed Central

Pontén, Annica; Walsh, Stuart; Malan, Daniela; Xian, Xiaojie; Schéele, Susanne; Tarnawski, Laura; Fleischmann, Bernd K.; Jovinge, Stefan

2013-01-01

Purification of cardiomyocytes from the embryonic mouse heart, embryonic stem (ES) or induced pluripotent stem cells (iPS) is a challenging task and will require specific isolation procedures. Lately the significance of surface markers for the isolation of cardiac cell populations with fluorescence activated cell sorting (FACS) has been acknowledged, and the hunt for cardiac specific markers has intensified. As cardiomyocytes have traditionally been characterized by their expression of specific transcription factors and structural proteins, and not by specific surface markers, this constitutes a significant bottleneck. Lately, Flk-1, c-kit and the cellular prion protein have been reported to specify cardiac progenitors, however, no surface markers have so far been reported to specify a committed cardiomyocyte. Herein show for the first time, that embryonic cardiomyocytes can be isolated with 98% purity, based on their expression of vascular cell adhesion molecule-1 (VCAM-1). The FACS-isolated cells express phenotypic markers for embryonic committed cardiomyocytes but not cardiac progenitors. An important aspect of FACS is to provide viable cells with retention of functionality. We show that VCAM-1 positive cardiomyocytes can be isolated with 95% viability suitable for in vitro culture, functional assays or expression analysis. In patch-clamp experiments we provide evidence of functionally intact cardiomyocytes of both atrial and ventricular subtypes. This work establishes that cardiomyocytes can be isolated with a high degree of purity and viability through FACS, based on specific surface marker expression as has been done in the hematopoietic field for decades. Our FACS protocol represents a significant advance in which purified populations of cardiomyocytes may be isolated and utilized for downstream applications, such as purification of ES-cell derived cardiomyocytes. PMID:24386094

Embryonic metabolism of the ornithischian dinosaurs Protoceratops andrewsi and Hypacrosaurus stebingeri and implications for calculations of dinosaur egg incubation times

NASA Astrophysics Data System (ADS)

Lee, Scott A.

2017-04-01

The embryonic metabolisms of the ornithischian dinosaurs Protoceratops andrewsi and Hypacrosaurus stebingeri have been determined and are in the range observed in extant reptiles. The average value of the measured embryonic metabolic rates for P. andrewsi and H. stebingeri are then used to calculate the incubation times for 21 dinosaurs from both Sauischia and Ornithischia using a mass growth model based on conservation of energy. The calculated incubation times vary from about 70 days for Archaeopteryx lithographica to about 180 days for Alamosaurus sanjuanensis. Such long incubation times seem unlikely, particularly for the sauropods and large theropods. Incubation times are also predicted with the assumption that the saurischian dinosaurs had embryonic metabolisms in the range observed in extant birds.

Embryonic metabolism of the ornithischian dinosaurs Protoceratops andrewsi and Hypacrosaurus stebingeri and implications for calculations of dinosaur egg incubation times.

PubMed

Lee, Scott A

2017-04-01

The embryonic metabolisms of the ornithischian dinosaurs Protoceratops andrewsi and Hypacrosaurus stebingeri have been determined and are in the range observed in extant reptiles. The average value of the measured embryonic metabolic rates for P. andrewsi and H. stebingeri are then used to calculate the incubation times for 21 dinosaurs from both Sauischia and Ornithischia using a mass growth model based on conservation of energy. The calculated incubation times vary from about 70 days for Archaeopteryx lithographica to about 180 days for Alamosaurus sanjuanensis. Such long incubation times seem unlikely, particularly for the sauropods and large theropods. Incubation times are also predicted with the assumption that the saurischian dinosaurs had embryonic metabolisms in the range observed in extant birds.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kalousek, D.K.; Fitch, N.; Paradice, B.

Topics covered in this book include a general review of normal embryonic and fetal development; abortion and the basic approach to the examination of aborted embryos and fetuses; and pathologic findings detected on examination of products of conception. The authors illustrate specific morphologic lesions and the variable expression of genetic syndromes in the embryonic and fetal periods.

EFFECT OF TRANSIENT EMBRYONIC IN VIVO EXPOSURE TO THE ENDOCRINE DISRUPTOR METHOXYCHLOR ON EMBRYONIC AND POSTNATAL TESTIS DEVELOPMENT. (R827405)

EPA Science Inventory

The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

A New Wnt1-CRE TomatoRosa Embryonic Stem Cell Line: A Tool for Studying Neural Crest Cell Integration Capacity.

PubMed

Acuna-Mendoza, Soledad; Martin, Sabrina; Kuchler-Bopp, Sabine; Ribes, Sandy; Thalgott, Jérémy; Chaussain, Catherine; Creuzet, Sophie; Lesot, Hervé; Lebrin, Franck; Poliard, Anne

2017-12-01

Neural crest (NC) cells are a migratory, multipotent population giving rise to numerous lineages in the embryo. Their plasticity renders attractive their use in tissue engineering-based therapies, but further knowledge on their in vivo behavior is required before clinical transfer may be envisioned. We here describe the isolation and characterization of a new mouse embryonic stem (ES) line derived from Wnt1-CRE-R26 Rosa TomatoTdv blastocyst and show that it displays the characteristics of typical ES cells. Further, these cells can be efficiently directed toward an NC stem cell-like phenotype as attested by concomitant expression of NC marker genes and Tomato fluorescence. As native NC progenitors, they are capable of differentiating toward typical derivative phenotypes and interacting with embryonic tissues to participate in the formation of neo-structures. Their specific fluorescence allows purification and tracking in vivo. This cellular tool should facilitate a better understanding of the mechanisms driving NC fate specification and help identify the key interactions developed within a tissue after in vivo implantation. Altogether, this novel model may provide important knowledge to optimize NC stem cell graft conditions, which are required for efficient tissue repair.

A genetic screen for temperature-sensitive cell-division mutants of Caenorhabditis elegans.

PubMed Central

O'Connell, K F; Leys, C M; White, J G

1998-01-01

A novel screen to isolate conditional cell-division mutants in Caenorhabditis elegans has been developed. The screen is based on the phenotypes associated with existing cell-division mutations: some disrupt postembryonic divisions and affect formation of the gonad and ventral nerve cord-resulting in sterile, uncoordinated animals-while others affect embryonic divisions and result in lethality. We obtained 19 conditional mutants that displayed these phenotypes when shifted to the restrictive temperature at the appropriate developmental stage. Eighteen of these mutations have been mapped; 17 proved to be single alleles of newly identified genes, while 1 proved to be an allele of a previously identified gene. Genetic tests on the embryonic lethal phenotypes indicated that for 13 genes, embryogenesis required maternal expression, while for 6, zygotic expression could suffice. In all cases, maternal expression of wild-type activity was found to be largely sufficient for embryogenesis. Cytological analysis revealed that 10 mutants possessed embryonic cell-division defects, including failure to properly segregate DNA, failure to assemble a mitotic spindle, late cytokinesis defects, prolonged cell cycles, and improperly oriented mitotic spindles. We conclude that this approach can be used to identify mutations that affect various aspects of the cell-division cycle. PMID:9649522

Notochord-derived BMP antagonists inhibit endothelial cell generation and network formation.

PubMed

Bressan, Michael; Davis, Patricia; Timmer, John; Herzlinger, Doris; Mikawa, Takashi

2009-02-01

Embryonic blood vessel formation is initially mediated through the sequential differentiation, migration, and assembly of endothelial cells (ECs). While many molecular signals that promote vascular development have been identified, little is known about suppressors of this process. In higher vertebrates, including birds and mammals, the vascular network forms throughout the embryonic disk with the exception of a region along the midline. We have previously shown that the notochord is responsible for the generation and maintenance of the avascular midline and that BMP antagonists expressed by this embryonic tissue, including Noggin and Chordin, can mimic this inhibitory role. Here we report that the notochord suppresses the generation of ECs from the mesoderm both in vivo and in vitro. We also report that the notochord diminishes the ability of mature ECs to organize into a primitive plexus. Furthermore, Noggin mimics notochord-based inhibition by preventing mesodermal EC generation and mature EC network formation. These findings suggest that the mesoderm surrounding the midline is competent to give rise to ECs and to form blood vessels, but that notochord derived-BMP antagonists suppress EC differentiation and maturation processes leading to inhibition of midline vessel formation.

F4/80+ Host Macrophages Are a Barrier to Murine Embryonic Stem Cell-Derived Hematopoietic Progenitor Engraftment In Vivo.

PubMed

Thompson, Heather L; van Rooijen, Nico; McLelland, Bryce T; Manilay, Jennifer O

2016-01-01

Understanding how embryonic stem cells and their derivatives interact with the adult host immune system is critical to developing their therapeutic potential. Murine embryonic stem cell-derived hematopoietic progenitors (ESHPs) were generated via coculture with the bone marrow stromal cell line, OP9, and then transplanted into NOD.SCID.Common Gamma Chain (NSG) knockout mice, which lack B, T, and natural killer cells. Compared to control mice transplanted with adult lineage-negative bone marrow (Lin - BM) progenitors, ESHP-transplanted mice attained a low but significant level of donor hematopoietic chimerism. Based on our previous studies, we hypothesized that macrophages might contribute to the low engraftment of ESHPs in vivo . Enlarged spleens were observed in ESHP-transplanted mice and found to contain higher numbers of host F4/80 + macrophages compared to BM-transplanted controls. In vivo depletion of host macrophages using clodronate-loaded liposomes improved the ESHP-derived hematopoietic chimerism in the spleen but not in the BM. F4/80 + macrophages demonstrated a striking propensity to phagocytose ESHP targets in vitro . Taken together, these results suggest that macrophages are a barrier to both syngeneic and allogeneic ESHP engraftment in vivo .

[Influence of the activator of transcription GAL4 on growth and development of embryos and embryonic cells in primary cultures of sand dollar].

PubMed

Odintsova, N A; Kiselev, K V; Bulgakov, V P; Kol'tsova, E A; Iakovlev, K V

2003-01-01

In order to solve many tasks of biotechnology, constant lines of the cells of marine invertebrates with a high growth potential are required, which are absent at present. We used the universal activator of transcription gal4 to change the degree of expression of genes of growth factors in embryonic sea urchin cells and, thereby, increase their proliferative activity. The fertilized sea urchin eggs and dissociated embryonic cells at the blastula stage were treated with plasmids containing both the functional gene gal4 and the gene devoid of the regions encoding the activator domain. The transfection of embryonic sea urchin eggs with the functional gene led to cell dedifferentiation and formation of tumor-like structures in the embryos or increased number of embryonic cells in culture. In the cells obtained from the transfected embryos, the pigments were found within two months of cultivation, whose absorption spectrum coincided with that of echinochrome.

Ethanol Inactivated Mouse Embryonic Fibroblasts Maintain the Self-Renew and Proliferation of Human Embryonic Stem Cells.

PubMed

Huang, Boxian; Ning, Song; Zhuang, Lili; Jiang, Chunyan; Cui, Yugui; Fan, Guoping; Qin, Lianju; Liu, Jiayin

2015-01-01

Conventionally, mouse embryonic fibroblasts (MEFs) inactivated by mitomycin C or irradiation were applied to support the self-renew and proliferation of human embryonic stem cells (hESCs). To avoid the disadvangtages of mitomycin C and irradiation, here MEFs were treated by ethanol (ET). Our data showed that 10% ET-inactivated MEFs (eiMEFs) could well maintain the self-renew and proliferation of hESCs. hESCs grown on eiMEFs expressed stem cell markers of NANOG, octamer-binding protein 4 (OCT4), stage-specific embryonic antigen-4 (SSEA4) and tumour related antigen-1-81 (TRA-1-81), meanwhile maintained normal karyotype after long time culture. Also, hESCs cocultured with eiMEFs were able to form embryoid body (EB) in vitro and develop teratoma in vivo. Moreover, eiMEFs could keep their nutrient functions after long time cryopreservation. Our results indicate that the application of eiMEF in hESCs culture is safe, economical and convenient, thus is a better choice.

Human embryonic stem cell-derived pancreatic endoderm alleviates diabetic pathology and improves reproductive outcome in C57BL/KsJ-Lep(db/+) gestational diabetes mellitus mice.

PubMed

Xing, Baoheng; Wang, Lili; Li, Qin; Cao, Yalei; Dong, Xiujuan; Liang, Jun; Wu, Xiaohua

2015-07-01

Gestational diabetes mellitus is a condition commonly encountered during mid to late pregnancy with pathologic manifestations including hyperglycemia, hyperinsulinemia, insulin resistance, and fetal maldevelopment. The cause of gestational diabetes mellitus can be attributed to both genetic and environmental factors, hence complicating its diagnosis and treatment. Pancreatic progenitors derived from human embryonic stem cells were shown to be able to effectively treat diabetes in mice. In this study, we have developed a system of treating diabetes using human embryonic stem cell-derived pancreatic endoderm in a mouse model of gestational diabetes mellitus. Human embryonic stem cells were differentiated in vitro into pancreatic endoderm, which were then transplanted into db/+ mice suffering from gestational diabetes mellitus. The transplant greatly improved glucose metabolism and reproductive outcome of the females compared with the control groups. Our findings support the feasibility of using differentiated human embryonic stem cells for treating gestational diabetes mellitus patients. Copyright © 2015 Elsevier Inc. All rights reserved.

Flt1/VEGFR1 heterozygosity causes transient embryonic edema.

PubMed

Otowa, Yasunori; Moriwaki, Kazumasa; Sano, Keigo; Shirakabe, Masanori; Yonemura, Shigenobu; Shibuya, Masabumi; Rossant, Janet; Suda, Toshio; Kakeji, Yoshihiro; Hirashima, Masanori

2016-06-02

Vascular endothelial growth factor-A is a major player in vascular development and a potent vascular permeability factor under physiological and pathological conditions by binding to a decoy receptor Flt1 and its primary receptor Flk1. In this study, we show that Flt1 heterozygous (Flt1(+/-)) mouse embryos grow up to adult without life-threatening abnormalities but exhibit a transient embryonic edema around the nuchal and back regions, which is reminiscent of increased nuchal translucency in human fetuses. Vascular permeability is enhanced and an intricate infolding of the plasma membrane and huge vesicle-like structures are seen in Flt1(+/-) capillary endothelial cells. Flk1 tyrosine phosphorylation is elevated in Flt1(+/-) embryos, but Flk1 heterozygosity does not suppress embryonic edema caused by Flt1 heterozygosity. When Flt1 mutants are crossed with Aspp1(-/-) mice which exhibit a transient embryonic edema with delayed formation and dysfunction of lymphatic vessels, only 5.7% of Flt1(+/-); Aspp1(-/-) mice survive, compared to expected ratio (25%). Our results demonstrate that Flt1 heterozygosity causes a transient embryonic edema and can be a risk factor for embryonic lethality in combination with other mutations causing non-lethal vascular phenotype.

Lack of centrioles and primary cilia in STIL−/− mouse embryos

PubMed Central

David, Ahuvit; Liu, Fengying; Tibelius, Alexandra; Vulprecht, Julia; Wald, Diana; Rothermel, Ulrike; Ohana, Reut; Seitel, Alexander; Metzger, Jasmin; Ashery-Padan, Ruth; Meinzer, Hans-Peter; Gröne, Hermann-Josef; Izraeli, Shai; Krämer, Alwin

2014-01-01

Although most animal cells contain centrosomes, consisting of a pair of centrioles, their precise contribution to cell division and embryonic development is unclear. Genetic ablation of STIL, an essential component of the centriole replication machinery in mammalian cells, causes embryonic lethality in mice around mid gestation associated with defective Hedgehog signaling. Here, we describe, by focused ion beam scanning electron microscopy, that STIL−/− mouse embryos do not contain centrioles or primary cilia, suggesting that these organelles are not essential for mammalian development until mid gestation. We further show that the lack of primary cilia explains the absence of Hedgehog signaling in STIL−/− cells. Exogenous re-expression of STIL or STIL microcephaly mutants compatible with human survival, induced non-templated, de novo generation of centrioles in STIL−/− cells. Thus, while the abscence of centrioles is compatible with mammalian gastrulation, lack of centrioles and primary cilia impairs Hedgehog signaling and further embryonic development. PMID:25486474

Lack of centrioles and primary cilia in STIL(-/-) mouse embryos.

PubMed

David, Ahuvit; Liu, Fengying; Tibelius, Alexandra; Vulprecht, Julia; Wald, Diana; Rothermel, Ulrike; Ohana, Reut; Seitel, Alexander; Metzger, Jasmin; Ashery-Padan, Ruth; Meinzer, Hans-Peter; Gröne, Hermann-Josef; Izraeli, Shai; Krämer, Alwin

2014-01-01

Although most animal cells contain centrosomes, consisting of a pair of centrioles, their precise contribution to cell division and embryonic development is unclear. Genetic ablation of STIL, an essential component of the centriole replication machinery in mammalian cells, causes embryonic lethality in mice around mid gestation associated with defective Hedgehog signaling. Here, we describe, by focused ion beam scanning electron microscopy, that STIL(-/-) mouse embryos do not contain centrioles or primary cilia, suggesting that these organelles are not essential for mammalian development until mid gestation. We further show that the lack of primary cilia explains the absence of Hedgehog signaling in STIL(-/-) cells. Exogenous re-expression of STIL or STIL microcephaly mutants compatible with human survival, induced non-templated, de novo generation of centrioles in STIL(-/-) cells. Thus, while the abscence of centrioles is compatible with mammalian gastrulation, lack of centrioles and primary cilia impairs Hedgehog signaling and further embryonic development.

Behavioral development in embryonic and early juvenile cuttlefish (Sepia officinalis).

PubMed

O'Brien, Caitlin E; Mezrai, Nawel; Darmaillacq, Anne-Sophie; Dickel, Ludovic

2017-03-01

Though a mollusc, the cuttlefish Sepia officinalis possesses a sophisticated brain, advanced sensory systems, and a large behavioral repertoire. Cuttlefish provide a unique perspective on animal behavior due to their phylogenic distance from more traditional (vertebrate) models. S. officinalis is well-suited to addressing questions of behavioral ontogeny. As embryos, they can perceive and learn from their environment and experience no direct parental care. A marked progression in learning and behavior is observed during late embryonic and early juvenile development. This improvement is concomitant with expansion and maturation of the vertical lobe, the cephalopod analog of the mammalian hippocampus. This review synthesizes existing knowledge regarding embryonic and juvenile development in this species in an effort to better understand cuttlefish behavior and animal behavior in general. It will serve as a guide to future researchers and encourage greater awareness of the utility of this species to behavioral science. © 2016 Wiley Periodicals, Inc.

High-Throughput Screening Assay for Embryoid Body Differentiation of Human Embryonic Stem Cells

PubMed Central

Outten, Joel T.; Gadue, Paul; French, Deborah L.; Diamond, Scott L.

2012-01-01

Serum-free human pluripotent stem cell media offer the potential to develop reproducible clinically applicable differentiation strategies and protocols. The vast array of possible growth factor and cytokine combinations for media formulations makes differentiation protocol optimization both labor and cost-intensive. This unit describes a 96-well plate, 4-color flow cytometry-based screening assay to optimize pluripotent stem cell differentiation protocols. We provide conditions both to differentiate human embryonic stem cells (hESCs) to the three primary germ layers, ectoderm, endoderm, and mesoderm, and to utilize flow cytometry to distinguish between them. This assay exhibits low inter-well variability and can be utilized to efficiently screen a variety of media formulations, reducing cost, incubator space, and labor. Protocols can be adapted to a variety of differentiation stages and lineages. PMID:22415836

Studies of teratomas in mice: possibilities for the future production of animal models.

PubMed Central

Lehman, J. M.

1980-01-01

The murine teratoma-teratocarcinoma has become an interesting model for the study of neoplastic transformation, developmental biology, and possibly a useful system for genetic studies. These tumors arise spontaneously in 129 strain mice and can be induced in other strains by transplanting early embryos or portions of embryos into extrauterine sites. The majority of these tumors are benign, but some are capable of transplantation due to the presence of the stem cell, embryonal carcinoma, which is a multipotential cell able to proliferate and also differentiate into tissues and cell types representative of all the embryonic germ layers. It has been elegantly shown by transplantation of embryonal carcinoma cells into blastocysts which are then placed into a pseudopregnant mouse that a normal mouse is obtained composed of cells from the host blastocyst and also cells from the malignant embryonal carcinoma. Therefore, under this set of circumstances, embryonal carcinoma cells are induced to functionally differentiate into multiple cell and tissue types which are benign and able to contribute to the development of a mouse. The adaptation of the embryonal carcinoma cell to tissue culture has allowed the manipulation of these cells with subsequent selection of mutant cells which can be further transplanted into blastocysts to obtain a mouse which contains these mutant cells. If the mutant cells have populated the germ line, it may be possible to obtain a stock of mice with the lesion present in all cells. This system may be exploitable for studies in neoplasia, developmental biology, and with proper selection procedures, allow the development of new genetic strains of mice. PMID:7457573

Derivation and characterization of Chinese human embryonic stem cell line with high potential to differentiate into pancreatic and hepatic cells.

PubMed

Shi, Cheng; Shen, Huan; Jiang, Wei; Song, Zhi-Hua; Wang, Cheng-Yan; Wei, Li-Hui

2011-04-01

Human embryonic stem cells have prospective uses in regenerative medicine and drug screening. Every human embryonic stem cell line has its own genetic background, which determines its specific ability for differentiation as well as susceptibility to drugs. It is necessary to compile many human embryonic stem cell lines with various backgrounds for future clinical use, especially in China due to its large population. This study contributes to isolating new Chinese human embryonic stem cell lines with clarified directly differentiation ability. Donated embryos that exceeded clinical use in our in vitro fertilization-embryo transfer (IVF-ET) center were collected to establish human embryonic stem cells lines with informed consent. The classic growth factors of basic fibroblast growth factor (bFGF) and recombinant human leukaemia inhibitory factor (hLIF) for culturing embryonic stem cells were used to capture the stem cells from the plated embryos. Mechanical and enzymetic methods were used to propagate the newly established human embryonic stem cells line. The new cell line was checked for pluripotent characteristics with detecting the expression of stemness genes and observing spontaneous differentiation both in vitro and in vivo. Finally similar step-wise protocols from definitive endoderm to target specific cells were used to check the cell line's ability to directly differentiate into pancreatic and hepatic cells. We generated a new Chinese human embryonic stem cells line, CH1. This cell line showed the same characteristics as other reported Chinese human embryonic stem cells lines: normal morphology, karyotype and pluripotency in vitro and in vivo. The CH1 cells could be directly differentiated towards pancreatic and hepatic cells with equal efficiency compared to the H1 cell line. This newly established Chinese cell line, CH1, which is pluripotent and has high potential to differentiate into pancreatic and hepatic cells, will provide a useful tool for embryo development research, along with clinical treatments for diabetes and some hepatic diseases.

Adiposity associated changes in serum glucose and adiponectin levels modulate ovarian steroidogenesis during delayed embryonic development in the fruit bat, Cynopterus sphinx.

PubMed

Anuradha; Krishna, Amitabh

2018-06-01

The aim of the present study was to evaluate the mechanism by which embryonic development in Cynopterus sphinx is impaired during the period of increased accumulation of white adipose tissue during winter scarcity of food. The change in the mass of white adipose tissue during adipogenesis showed significant positive correlation with the circulating glucose level. But increase in circulating glucose level during the adipogenesis showed negative correlation with circulating progesterone and adiponectin levels. The in vivo study showed increased glucose uptake by the adipose tissue during adipogenesis due to increased expression of insulin receptor (IR) and glucose transporter (GLUT) 4 proteins. This study showed decline in the adiponectin level during fat accumulation. In the in vitro study, ovary treated with high doses of glucose showed impaired progesterone synthesis. This is due to decreased glucose uptake mediated decrease in the expression of luteinizing hormone-receptor, steroidogenic acute regulatory protein, IR, GLUT4 and AdipoR1 proteins. But the ovary treated with adiponectin either alone or with higher concentration of glucose showed improvement in progesterone synthesis due to increased expression of IR, GLUT4 and AdipoR1 mediated increased glucose uptake. In conclusion, increased circulating glucose level prior to winter dormancy preferably transported to white adipose tissue for fat accumulation diverting glucose away from the ovary. Consequently the decreased availability of adiponectin and glucose to the ovary and utero-embryonic unit may be responsible for impaired progesterone synthesis and delayed embryonic development. The delayed embryonic development in Cynopterus sphinx may have evolved, in part, as a mechanism to prevent pregnancy loss during the period of decreased energy availability. Copyright © 2018 Elsevier Inc. All rights reserved.

Variation in maternal effects and embryonic development rates among passerine species

USGS Publications Warehouse

Martin, T.E.; Schwabl, H.

2008-01-01

Embryonic development rates are reflected by the length of incubation period in birds, and these vary substantially among species within and among geographical regions. The incubation periods are consistently shorter in North America (Arizona study site) than in tropical (Venezuela) and subtropical (Argentina) South America based on the study of 83 passerine species in 17 clades. Parents, mothers in particular, may influence incubation periods and resulting offspring quality through proximate pathways, while variation in maternal strategies among species can result from selection by adult and offspring mortality. Parents of long-lived species, as is common in the tropics and subtropics, may be under selection to minimize costs to themselves during incubation. Indeed, time spent incubating is often lower in the tropical and subtropical species than the related north temperate species, causing cooler average egg temperatures in the southern regions. Decreased egg temperatures result in longer incubation periods and reflect a cost imposed on offspring by parents because energy cost to the embryo and risk of offspring predation are both increased. Mothers may adjust egg size and constituents as a means to partially offset such costs. For example, reduced androgen concentrations in egg yolks may slow development rates, but may enhance offspring quality through physiological trade-offs that may be particularly beneficial in longer-lived species, as in the tropics and subtropics. We provide initial data to show that yolks of tropical birds contain substantially lower concentrations of growth-promoting androgens than north temperate relatives. Thus, maternal (and parental) effects on embryonic development rates may include contrasting and complementary proximate influences on offspring quality and deserve further field study among species. ?? 2007 The Royal Society.

Variation in maternal effects and embryonic development rates among passerine species.

PubMed

Martin, Thomas E; Schwabl, Hubert

2008-05-12

Embryonic development rates are reflected by the length of incubation period in birds, and these vary substantially among species within and among geographical regions. The incubation periods are consistently shorter in North America (Arizona study site) than in tropical (Venezuela) and subtropical (Argentina) South America based on the study of 83 passerine species in 17 clades. Parents, mothers in particular, may influence incubation periods and resulting offspring quality through proximate pathways, while variation in maternal strategies among species can result from selection by adult and offspring mortality. Parents of long-lived species, as is common in the tropics and subtropics, may be under selection to minimize costs to themselves during incubation. Indeed, time spent incubating is often lower in the tropical and subtropical species than the related north temperate species, causing cooler average egg temperatures in the southern regions. Decreased egg temperatures result in longer incubation periods and reflect a cost imposed on offspring by parents because energy cost to the embryo and risk of offspring predation are both increased. Mothers may adjust egg size and constituents as a means to partially offset such costs. For example, reduced androgen concentrations in egg yolks may slow development rates, but may enhance offspring quality through physiological trade-offs that may be particularly beneficial in longer-lived species, as in the tropics and subtropics. We provide initial data to show that yolks of tropical birds contain substantially lower concentrations of growth-promoting androgens than north temperate relatives. Thus, maternal (and parental) effects on embryonic development rates may include contrasting and complementary proximate influences on offspring quality and deserve further field study among species.

Comparative Chondrogenesis of Human Cell Sources in 3D Scaffolds

PubMed Central

Tıg̑lı, R. Seda; Ghosh, Sourabh; Laha, Michael M.; Shevde, Nirupama K.; Daheron, Laurence; Gimble, Jeffrey; Gümüşdereliog̑lu, Menemşe; Kaplan, David L.

2009-01-01

Cartilage tissue can be engineered by starting from a diversity of cell sources, including stem-cell based and primary cell-based platforms. Selecting an appropriate cell source for the process of cartilage tissue engineering or repair is critical and challenging due to the variety of cell options available. In this study, cellular responses of isolated human chondrocytes, human embryonic stem cells and mesenchymal stem cells (MSCs) derived from three sources, human embryonic stem cells, bone marrow and adipose tissue, were assessed for chondrogenic potential in 3D culture. All cell sources were characterized by FACS analysis to compare expression of some surface markers. The cells were differentiated in two different biomaterial matrices, silk and chitosan scaffolds, in the presence and absence of bone morphogenetic protein 6 (BMP-6) along with the standard chondrogenic differentiating factors. Embryonic stem cells derived MSCs showed unique characteristics with preserved chondrogenic phenotype in both scaffolds with regard to chondrogenesis, as determined by real time RT-PCR, histological and microscopic analyses. After 4 weeks of cultivation, embryonic stem cells derived MSCs were promising for chondrogenesis, particularly in the silk scaffolds with BMP-6. The results suggest that cell source differences are important to consider with regard to chondrogenic outcomes and with the variables addressed here, the human embryonic stem cells derived MSCs were the preferred cell source. PMID:19382119

Present state and future perspectives of using pluripotent stem cells in toxicology research

PubMed Central

Löser, Peter

2011-01-01

The use of novel drugs and chemicals requires reliable data on their potential toxic effects on humans. Current test systems are mainly based on animals or in vitro–cultured animal-derived cells and do not or not sufficiently mirror the situation in humans. Therefore, in vitro models based on human pluripotent stem cells (hPSCs) have become an attractive alternative. The article summarizes the characteristics of pluripotent stem cells, including embryonic carcinoma and embryonic germ cells, and discusses the potential of pluripotent stem cells for safety pharmacology and toxicology. Special attention is directed to the potential application of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) for the assessment of developmental toxicology as well as cardio- and hepatotoxicology. With respect to embryotoxicology, recent achievements of the embryonic stem cell test (EST) are described and current limitations as well as prospects of embryotoxicity studies using pluripotent stem cells are discussed. Furthermore, recent efforts to establish hPSC-based cell models for testing cardio- and hepatotoxicity are presented. In this context, methods for differentiation and selection of cardiac and hepatic cells from hPSCs are summarized, requirements and implications with respect to the use of these cells in safety pharmacology and toxicology are presented, and future challenges and perspectives of using hPSCs are discussed. PMID:21225242

A simple in vitro culture system for tracheal cartilage development.

PubMed

Park, Jinhyung; Zhang, Jennifer J R; Choi, Ruth; Trinh, Irene; Kim, Peter C W

2010-02-01

Semi-circular tracheal cartilage is a critical determinant of maintaining architectural integrity of the respiratory airway. The current effort to understand the morphogenesis of tracheal cartilage is challenged by the lack of appropriate model systems. Here we report an in vitro tracheal cartilage system using embryonic tracheal–lung explants to recapitulate in vivo tracheal cartilage developmental processes. With modifications of a current lung culture protocol, we report a consistent in vitro technique of culturing tracheal cartilage from primitive mouse embryonic foregut for the first time. This tracheal culture system not only induces the formation of tracheal cartilage from the mouse embryonic foregut but also allows for the proper patterning of the developed tracheal cartilage. Furthermore, we show that this culture technique can be applied to culturing other types of cartilage in vertebrae, limbs, and ribs. We believe that this novel application of our in vitro culture system will facilitate the manipulation of cartilage development under various conditions and thus enabling us to advance our current limited knowledge on cartilage biology and development.

Oviposition behaviors and ontogenetic embryonic characteristics of the western tarnished plant bug, Lygus hesperus

USDA-ARS?s Scientific Manuscript database

Lygus hesperus Knight (Hemiptera: Miridae) is a key pest of fruit, vegetable, and field crops in the western United States but many aspects of L. hesperus ecology are poorly documented. A sound understanding of oviposition behavior and characterization of the phases of embryonic development would be...

Carotid-vertebrobasilar Anastomoses with Reference to Their Segmental Property.

PubMed

Namba, Katsunari

2017-06-15

The primitive carotid-vertebrobasilar anastomoses are primitive embryonic cerebral vessels that temporarily provide arterial supply from the internal carotid artery to the longitudinal neural artery, the future vertebrobasilar artery in the hindbrain. Four types known are the trigeminal, otic, hypoglossal, and proatlantal intersegmental arteries. The arteries are accompanied by their corresponding nerves and resemble an intersegmental pattern. These vessels exist in the very early period of cerebral arterial development and rapidly involute within a week. Occasionally, persistence of the carotid to vertebrobasilar anastomosis is discovered in the adult period, and is considered as the vestige of the corresponding primitive embryonic vessel. The embryonic development and the segmental property of the primitive carotid-vertebrobasilar anastomoses are discussed. This is followed by a brief description of the persisting anastomoses in adults.

X-chromosome dosage as a modulator of pluripotency, signalling and differentiation?

PubMed

Schulz, Edda G

2017-11-05

Already during early embryogenesis, before sex-specific hormone production is initiated, sex differences in embryonic development have been observed in several mammalian species. Typically, female embryos develop more slowly than their male siblings. A similar phenotype has recently been described in differentiating murine embryonic stem cells, where a double dose of the X-chromosome halts differentiation until dosage-compensation has been achieved through X-chromosome inactivation. On the molecular level, several processes associated with early differentiation of embryonic stem cells have been found to be affected by X-chromosome dosage, such as the transcriptional state of the pluripotency network, the activity pattern of several signal transduction pathways and global levels of DNA-methylation. This review provides an overview of the sex differences described in embryonic stem cells from mice and discusses a series of X-linked genes that are associated with pluripotency, signalling and differentiation and their potential involvement in mediating the observed X-dosage-dependent effects.This article is part of the themed issue 'X-chromosome inactivation: a tribute to Mary Lyon'. © 2017 The Author(s).

Mutagenesis Screen Identifies agtpbp1 and eps15L1 as Essential for T lymphocyte Development in Zebrafish.

PubMed

Seiler, Christoph; Gebhart, Nichole; Zhang, Yong; Shinton, Susan A; Li, Yue-sheng; Ross, Nicola L; Liu, Xingjun; Li, Qin; Bilbee, Alison N; Varshney, Gaurav K; LaFave, Matthew C; Burgess, Shawn M; Balciuniene, Jorune; Balciunas, Darius; Hardy, Richard R; Kappes, Dietmar J; Wiest, David L; Rhodes, Jennifer

2015-01-01

Genetic screens are a powerful tool to discover genes that are important in immune cell development and function. The evolutionarily conserved development of lymphoid cells paired with the genetic tractability of zebrafish make this a powerful model system for this purpose. We used a Tol2-based gene-breaking transposon to induce mutations in the zebrafish (Danio rerio, AB strain) genome, which served the dual purpose of fluorescently tagging cells and tissues that express the disrupted gene and provided a means of identifying the disrupted gene. We identified 12 lines in which hematopoietic tissues expressed green fluorescent protein (GFP) during embryonic development, as detected by microscopy. Subsequent analysis of young adult fish, using a novel approach in which single cell suspensions of whole fish were analyzed by flow cytometry, revealed that 8 of these lines also exhibited GFP expression in young adult cells. An additional 15 lines that did not have embryonic GFP+ hematopoietic tissue by microscopy, nevertheless exhibited GFP+ cells in young adults. RT-PCR analysis of purified GFP+ populations for expression of T and B cell-specific markers identified 18 lines in which T and/or B cells were fluorescently tagged at 6 weeks of age. As transposon insertion is expected to cause gene disruption, these lines can be used to assess the requirement for the disrupted genes in immune cell development. Focusing on the lines with embryonic GFP+ hematopoietic tissue, we identified three lines in which homozygous mutants exhibited impaired T cell development at 6 days of age. In two of the lines we identified the disrupted genes, agtpbp1 and eps15L1. Morpholino-mediated knockdown of these genes mimicked the T cell defects in the corresponding mutant embryos, demonstrating the previously unrecognized, essential roles of agtpbp1 and eps15L1 in T cell development.

Effects of the EVCAM chemical validation library on differentiation using marker gene expression in lmouse embryonic stem cells

EPA Science Inventory

The adherent cell differentiation and cytotoxicity (ACDC) assay was used to profile the effects of the ECVAM EST validation chemical library (19 compounds) on J1 mouse embryonic stem cells (mESC). PCR-based TaqMan Low Density Arrays (TLDA) provided a high-content assessment of al...

Toxicological effects of the different substances in tobacco smoke on human embryonic development by a systems chemo-biology approach.

PubMed

Feltes, Bruno César; de Faria Poloni, Joice; Notari, Daniel Luis; Bonatto, Diego

2013-01-01

The physiological and molecular effects of tobacco smoke in adult humans and the development of cancer have been well described. In contrast, how tobacco smoke affects embryonic development remains poorly understood. Morphological studies of the fetuses of smoking pregnant women have shown various physical deformities induced by constant fetal exposure to tobacco components, especially nicotine. In addition, nicotine exposure decreases fetal body weight and bone/cartilage growth in addition to decreasing cranial diameter and tibia length. Unfortunately, the molecular pathways leading to these morphological anomalies are not completely understood. In this study, we applied interactome data mining tools and small compound interaction networks to elucidate possible molecular pathways associated with the effects of tobacco smoke components during embryonic development in pregnant female smokers. Our analysis showed a relationship between nicotine and 50 additional harmful substances involved in a variety of biological process that can cause abnormal proliferation, impaired cell differentiation, and increased oxidative stress. We also describe how nicotine can negatively affect retinoic acid signaling and cell differentiation through inhibition of retinoic acid receptors. In addition, nicotine causes a stress reaction and/or a pro-inflammatory response that inhibits the agonistic action of retinoic acid. Moreover, we show that the effect of cigarette smoke on the developing fetus could represent systemic and aggressive impacts in the short term, causing malformations during certain stages of development. Our work provides the first approach describing how different tobacco constituents affect a broad range of biological process in human embryonic development.

Toxicological Effects of the Different Substances in Tobacco Smoke on Human Embryonic Development by a Systems Chemo-Biology Approach

PubMed Central

Feltes, Bruno César; Poloni, Joice de Faria; Notari, Daniel Luis; Bonatto, Diego

2013-01-01

The physiological and molecular effects of tobacco smoke in adult humans and the development of cancer have been well described. In contrast, how tobacco smoke affects embryonic development remains poorly understood. Morphological studies of the fetuses of smoking pregnant women have shown various physical deformities induced by constant fetal exposure to tobacco components, especially nicotine. In addition, nicotine exposure decreases fetal body weight and bone/cartilage growth in addition to decreasing cranial diameter and tibia length. Unfortunately, the molecular pathways leading to these morphological anomalies are not completely understood. In this study, we applied interactome data mining tools and small compound interaction networks to elucidate possible molecular pathways associated with the effects of tobacco smoke components during embryonic development in pregnant female smokers. Our analysis showed a relationship between nicotine and 50 additional harmful substances involved in a variety of biological process that can cause abnormal proliferation, impaired cell differentiation, and increased oxidative stress. We also describe how nicotine can negatively affect retinoic acid signaling and cell differentiation through inhibition of retinoic acid receptors. In addition, nicotine causes a stress reaction and/or a pro-inflammatory response that inhibits the agonistic action of retinoic acid. Moreover, we show that the effect of cigarette smoke on the developing fetus could represent systemic and aggressive impacts in the short term, causing malformations during certain stages of development. Our work provides the first approach describing how different tobacco constituents affect a broad range of biological process in human embryonic development. PMID:23637898

The embryogenesis of the tick Rhipicephalus (Boophilus) microplus: the establishment of a new chelicerate model system.

PubMed

Santos, Vitória Tobias; Ribeiro, Lupis; Fraga, Amanda; de Barros, Cíntia Monteiro; Campos, Eldo; Moraes, Jorge; Fontenele, Marcio Ribeiro; Araújo, Helena Marcolla; Feitosa, Natalia Martins; Logullo, Carlos; da Fonseca, Rodrigo Nunes

2013-12-01

Chelicerates, which include spiders, ticks, mites, scorpions, and horseshoe crabs, are members of the phylum Arthropoda. In recent years, several molecular experimental studies of chelicerates have examined the embryology of spiders; however, the embryology of other groups, such as ticks (Acari: Parasitiformes), has been largely neglected. Ticks and mites are believed to constitute a monophyletic group, the Acari. Due to their blood-sucking activities, ticks are also known to be vectors of several diseases. In this study, we analyzed the embryonic development of the cattle tick, Rhipicephalus (Boophilus) microplus (Acari: Ixodidae). First, we developed an embryonic staging system consisting of 14 embryonic stages. Second, histological analysis and antibody staining unexpectedly revealed the presence of a population of tick cells with similar characteristics to the spider cumulus. Cumulus cell populations also exist in other chelicerates; these cells are responsible for the breaking of radial symmetry through bone morphogenetic protein signaling. Third, it was determined that the posterior (opisthosomal) embryonic region of R. microplus is segmented. Finally, we identified the presence of a transient ventral midline furrow and the formation and regression of a fourth leg pair; these features may be regarded as hallmarks of late tick embryogenesis. Importantly, most of the aforementioned features are absent from mite embryos, suggesting that mites and ticks do not constitute a monophyletic group or that mites have lost these features. Taken together, our findings provide fundamental common ground for improving knowledge regarding tick embryonic development, thereby facilitating the establishment of a new chelicerate model system. Copyright © 2013 Wiley Periodicals, Inc.

Non-staining visualization of embryogenesis and energy metabolism in medaka fish eggs using near-infrared spectroscopy and imaging.

PubMed

Puangchit, Paralee; Ishigaki, Mika; Yasui, Yui; Kajita, Misato; Ritthiruangdej, Pitiporn; Ozaki, Yukihiro

2017-12-04

The energy metabolism and embryogenesis of fertilized Japanese medaka eggs were investigated in vivo at the molecular level using near-infrared (NIR) spectroscopy and imaging. Changes in chemical components, such as proteins and lipids, in yolk sphere and embryonic body were studied over the course of embryonic development. Metabolic changes that represent variations in the concentrations and molecular compositions of proteins and lipids in the yolk part, particularly on the 1 st day after fertilization and the day just before hatching, were successfully identified in the 4900-4000 cm -1 wavenumber region. The yolk components were shown to have specific functions at the very early and final stages of the embryonic development. Proteins with α-helix- or β-sheet-rich structures clearly showed the different variation patterns within the developing egg. Furthermore, the distribution of lipids could be selectively visualized using data from the higher wavenumber region. Detailed embryonic structures were clearly depicted in the NIR images using the data from the 6400-5500 cm -1 region in which the embryo parts had some characteristic peaks due to unsaturated fatty acids. It was made clear that yolk and embryo parts had different components especially lipid components. The present study provides new insights into material variations in the fertilized egg during its growth. NIR imaging proved to be valuable in investigating the embryogenesis in vivo at the molecular level in terms of changes in biomolecular concentrations and compositions, metabolic differentiation, and detailed information about embryonic structures without the need for staining.

Regulation of protein phosphatase 2A during embryonic diapause process in the silkworm, Bombyx mori.

PubMed

Gu, Shi-Hong; Hsieh, Hsiao-Yen; Lin, Pei-Ling

2017-11-01

Regulation of protein phosphorylation requires coordinated interactions between protein kinases and protein phosphatases. In the present study, we investigated regulation of protein phosphatase 2A (PP2A) during the embryonic diapause process of B. mori. An immunoblotting analysis showed that Bombyx eggs contained a catalytic C subunit, a major regulatory B subunit (B55/PR55 subunit), and a structural A subunit, with the A and B subunits undergoing differential changes between diapause and non-diapause eggs during embryonic process. In non-diapause eggs, eggs whose diapause initiation was prevented by HCl, and eggs in which diapause had been terminated by chilling of diapausing eggs at 5°C for 70days and then were transferred to 25°C, protein levels of the A and B subunits of PP2A gradually increased toward embryonic development. However, protein levels of the A and B subunits in diapause eggs remained at low levels during the first 8days after oviposition. The direct determination of PP2A enzymatic activity showed that the activity remained at low levels in diapause eggs during the first 8days after oviposition. However, in non-diapause eggs, eggs whose diapause initiation was prevented by HCl, and eggs in which diapause had been terminated by chilling, PP2A enzymatic activity sharply increased during the first several days, reached a peak during the middle embryonic development, and then greatly decreased 3 or 4days before hatching. Examination of temporal changes in mRNA expression levels of the catalytic β subunit and regulatory subunit of PP2A showed high levels in eggs whose diapause initiation was prevented by HCl compared to those in diapause eggs. These results demonstrate that the higher PP2A gene expression and PP2A A and B subunit protein levels and increased enzymatic activity are related to embryonic development of B. mori. Copyright © 2017 Elsevier Ltd. All rights reserved.

Graphic and movie illustrations of human prenatal development and their application to embryological education based on the human embryo specimens in the Kyoto collection.

PubMed

Yamada, Shigehito; Uwabe, Chigako; Nakatsu-Komatsu, Tomoko; Minekura, Yutaka; Iwakura, Masaji; Motoki, Tamaki; Nishimiya, Kazuhiko; Iiyama, Masaaki; Kakusho, Koh; Minoh, Michihiko; Mizuta, Shinobu; Matsuda, Tetsuya; Matsuda, Yoshimasa; Haishi, Tomoyuki; Kose, Katsumi; Fujii, Shingo; Shiota, Kohei

2006-02-01

Morphogenesis in the developing embryo takes place in three dimensions, and in addition, the dimension of time is another important factor in development. Therefore, the presentation of sequential morphological changes occurring in the embryo (4D visualization) is essential for understanding the complex morphogenetic events and the underlying mechanisms. Until recently, 3D visualization of embryonic structures was possible only by reconstruction from serial histological sections, which was tedious and time-consuming. During the past two decades, 3D imaging techniques have made significant advances thanks to the progress in imaging and computer technologies, computer graphics, and other related techniques. Such novel tools have enabled precise visualization of the 3D topology of embryonic structures and to demonstrate spatiotemporal 4D sequences of organogenesis. Here, we describe a project in which staged human embryos are imaged by the magnetic resonance (MR) microscope, and 3D images of embryos and their organs at each developmental stage were reconstructed based on the MR data, with the aid of computer graphics techniques. On the basis of the 3D models of staged human embryos, we constructed a data set of 3D images of human embryos and made movies to illustrate the sequential process of human morphogenesis. Furthermore, a computer-based self-learning program of human embryology is being developed for educational purposes, using the photographs, histological sections, MR images, and 3D models of staged human embryos. Copyright 2005 Wiley-Liss, Inc.

[Artificial propagation and embryonic development of Neolissochilus benasi].

PubMed

Pan, Xiao-Fu; Liu, Qian; Wang, Xiao-Ai; Yang, Jun-Xing; Chen, Xiao-Yong; Li, Zai-Yun; Li, Lie

2013-12-01

From 2009 to 2011, luteinizing hormone releasing hormone analogue (LHRH-A2) mixed with domperidon (DOM) was successfully applied during the artificial propagation of Neolissochilus benasi. Totally, 60 females and 100 males were injected with the hormone mixture, resulting in 47 (78.3%) females and 92 (92.0%) males being successfully spawned. A total of 1,986-5 854 eggs were spawned per female with an egg diameter varying between 2.2-2.8 mm, and an average nucleus deviation rate of 73.2%. Sperm density, vitality and life span were 16.32±2.89×10(9)/mL, 60.6±3.2% and 70.2±5.3 s, respectively. On the whole, the embryonic development of N. benasi was similar to that of zebra fish-albeit relatively slower-lasting approximately 120 hours. The development itself can be divided into six discrete stages: zygote, cleavage, blastula, gastrula, segmentation and hatching. Results showed that the average hatching rate was 32.4%, with 86.5% of larvae surviving 45 days after hatching. During embryonic development, deformities commonly occurred on the mouth, chest, ocular region, especially in the spinal column. To try to attempt improving future breeding efforts, we provided a survey of the embryonic developmental difficulties of N. benasi using LHRH-A2 followed by several potential solutions, including providing suitable breeding conditions and minimizing capture stresses.

Dimethadione embryotoxicity in the rat is neither correlated with maternal systemic drug concentrations nor embryonic tissue levels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozolinš, Terence R.S., E-mail: ozolinst@queensu.ca; Weston, Andrea D.; Perretta, Anthony

Pregnant rats treated with dimethadione (DMO), the N-demethylated metabolite of the anticonvulsant trimethadione, produce offspring having a 74% incidence of congenital heart defects (CHD); however, the incidence of CHD has high inter-litter variability (40–100%) that presents a challenge when studying the initiating events prior to the presentation of an abnormal phenotype. We hypothesized that the variability in CHD incidence was the result of differences in maternal systemic concentrations or embryonic tissue concentrations of DMO. To test this hypothesis, dams were administered 300 mg/kg DMO every 12 h from the evening of gestational day (GD) 8 until the morning of GDmore » 11 (six total doses). Maternal serum levels of DMO were assessed on GD 11, 12, 13, 14, 15, 18 and 21. Embryonic tissue concentrations of DMO were assessed on GD 11, 12, 13 and 14. In a separate cohort of GD 12 embryos, DMO concentrations and parameters of growth and development were assessed to determine if tissue levels of DMO were correlated with these endpoints. Embryos were exposed directly to different concentrations of DMO with whole embryo culture (WEC) and their growth and development assessed. Key findings were that neither maternal systemic concentrations nor tissue concentrations of DMO identified embryos that were sensitive or resistant to DMO in vivo. Direct exposure of embryos to DMO via WEC also failed to show correlations between embryonic concentrations of DMO with developmental outcomes in vitro. We conclude that neither maternal serum nor embryonic tissue concentrations of DMO predict embryonic outcome. - Highlights: • Dimethadione (DMO) induces septation defects (VSD) in rat offspring. • Despite high rate of VSD defects inter-litter variability is 40–100%. • Maternal and embryonic concentrations of DMO were assessed. • Neither serum nor tissue levels of DMO were correlated with embryotoxicity.« less

A Role for Caenorhabditis elegans Importin IMA-2 in Germ Line and Embryonic Mitosis

PubMed Central

Geles, Kenneth G.; Johnson, Jeffrey J.; Jong, Sena; Adam, Stephen A.

2002-01-01

The importin α family of nuclear-cytoplasmic transport factors mediates the nuclear localization of proteins containing classical nuclear localization signals. Metazoan animals express multiple importin α proteins, suggesting their possible roles in cell differentiation and development. Adult Caenorhabditis elegans hermaphrodites express three importin α proteins, IMA-1, IMA-2, and IMA-3, each with a distinct expression and localization pattern. IMA-2 was expressed exclusively in germ line cells from the early embryonic through adult stages. The protein has a dynamic pattern of localization dependent on the stage of the cell cycle. In interphase germ cells and embryonic cells, IMA-2 is cytoplasmic and nuclear envelope associated, whereas in developing oocytes, the protein is cytoplasmic and intranuclear. During mitosis in germ line cells and embryos, IMA-2 surrounded the condensed chromosomes but was not directly associated with the mitotic spindle. The timing of IMA-2 nuclear localization suggested that the protein surrounded the chromosomes after fenestration of the nuclear envelope in prometaphase. Depletion of IMA-2 by RNA-mediated gene interference (RNAi) resulted in embryonic lethality and a terminal aneuploid phenotype. ima-2(RNAi) embryos have severe defects in nuclear envelope formation, accumulating nucleoporins and lamin in the cytoplasm. We conclude that IMA-2 is required for proper chromosome dynamics in germ line and early embryonic mitosis and is involved in nuclear envelope assembly at the conclusion of mitosis. PMID:12221121

The king is dead, long live the king: entering a new era of stem cell research and clinical development.

PubMed

Ichim, Thomas; Riordan, Neil H; Stroncek, David F

2011-12-20

In mid November the biopharma industry was shocked by the announcement from Geron that they were ending work on embryonic stem cell research and therapy. For more than 10 years the public image of all stem cell research has been equated with embryonic stem cells. Unfortunately, a fundamentally important medical and financial fact was being ignored: embryonic stem cell therapy is extremely immature. In parallel to efforts in embryonic stem cell research and development, scientists and physicians in the field of adult stem cells realized that the natural role of adult stem cells in the body is to promote healing and to act like endogenous "repair cells" and, as a result, numerous companies have entered the field of adult stem cell therapy with the goal of expanding numbers of adult stem cells for administration to patients with various conditions. In contrast to embryonic stem cells, which are extremely expensive and potentially dangerous, adult cell cells are inexpensive and have an excellent safety record when used in humans. Many studies are now showing that adult stem cells are practical, patient-applicable, therapeutics that are very close to being available for incorporation into the practice of medicine. These events signal the entrance of the field of stem cells into a new era: an era where hype and misinformation no longer triumph over economic and medical realities.

Strand displacement amplification for ultrasensitive detection of human pluripotent stem cells.

PubMed

Wu, Wei; Mao, Yiping; Zhao, Shiming; Lu, Xuewen; Liang, Xingguo; Zeng, Lingwen

2015-06-30

Human pluripotent stem cells (hPSCs), such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), provide a powerful model system for studies of cellular identity and early mammalian development, which hold great promise for regenerative medicine. It is necessary to develop a convenient method to discriminate hPSCs from other cells in clinics and basic research. Herein, a simple and reliable biosensor for stem cell detection was established. In this biosensor system, stage-specific embryonic antigen-3 (SSEA-3) and stage-specific embryonic antigen-4 (SSEA-4) were used to mark human pluripotent stem cells (hPSCs). Antibody specific for SSEA-3 was coated onto magnetic beads for hPSCs enrichment, and antibody specific for SSEA-4 was conjugated with carboxyl-modified tDNA sequence which was used as template for strand displacement amplification (SDA). The amplified single strand DNA (ssDNA) was detected with a lateral flow biosensor (LFB). This biosensor is capable of detecting a minimum of 19 human embryonic stem cells by a strip reader and 100 human embryonic stem cells by the naked eye within 80min. This approach has also shown excellent specificity to distinguish hPSCs from other types of cells, showing that it is promising for specific and handy detection of human pluripotent stem cells. Copyright © 2015 Elsevier B.V. All rights reserved.

In vitro differentiation of embryonic stem cells into hepatocytes induced by fibroblast growth factors and bone morphological protein-4.

PubMed

Zhou, Qing-Jun; Huang, Yan-Dan; Xiang, Li-Xin; Shao, Jian-Zhong; Zhou, Guo-Shun; Yao, Hang; Dai, Li-Cheng; Lu, Yong-Liang

2007-01-01

The feasibility of transforming embryonic endoderm into different cell types is tightly controlled by mesodermal and septum transversumal signalings during early embryonic development. Here, an induction protocol tracing embryonic liver development was designed, in which, three growth factors, acid fibroblast growth factor, basic fibroblast growth factor and bone morphological protein-4 that secreted from pre-cardiac mesoderm and septum transversum mesenchyme, respectively, were employed to investigate their specific potency of modulating the mature hepatocyte proportion during the differentiation process. Results showed that hepatic differentiation took place spontaneously at a low level, however, supplements of the three growth factors gave rise to a significant up-regulation of mature hepatocytes. Bone morphological protein-4 highlighted the differentiation ratio to 40-55%, showing the most effective promotion, and also exhibited a synergistic effect with the other two fibroblast factors, whereas no similar phenomenon was observed between the other two factors, which was reported for the first time. Our study not only provides a high-performance system of embryonic stem cells differentiating into hepatocytes, which would supply a sufficient hepatic population for related studies, but also make it clear of the inductive effects of three important growth factors, which could support for further investigation on the mechanisms of mesodermal and septumal derived signalings that regulate hepatic differentiation.

Zebrafish as a Vertebrate Model System to Evaluate Effects of Environmental Toxicants on Cardiac Development and Function.

PubMed

Sarmah, Swapnalee; Marrs, James A

2016-12-16

Environmental pollution is a serious problem of the modern world that possesses a major threat to public health. Exposure to environmental pollutants during embryonic development is particularly risky. Although many pollutants have been verified as potential toxicants, there are new chemicals in the environment that need assessment. Heart development is an extremely sensitive process, which can be affected by environmentally toxic molecule exposure during embryonic development. Congenital heart defects are the most common life-threatening global health problems, and the etiology is mostly unknown. The zebrafish has emerged as an invaluable model to examine substance toxicity on vertebrate development, particularly on cardiac development. The zebrafish offers numerous advantages for toxicology research not found in other model systems. Many laboratories have used the zebrafish to study the effects of widespread chemicals in the environment on heart development, including pesticides, nanoparticles, and various organic pollutants. Here, we review the uses of the zebrafish in examining effects of exposure to external molecules during embryonic development in causing cardiac defects, including chemicals ubiquitous in the environment and illicit drugs. Known or potential mechanisms of toxicity and how zebrafish research can be used to provide mechanistic understanding of cardiac defects are discussed.

Maternal topoisomerase II alpha, not topoisomerase II beta, enables embryonic development of zebrafish top2a-/- mutants

PubMed Central

2011-01-01

Background Genetic alterations in human topoisomerase II alpha (TOP2A) are linked to cancer susceptibility. TOP2A decatenates chromosomes and thus is necessary for multiple aspects of cell division including DNA replication, chromosome condensation and segregation. Topoisomerase II alpha is also required for embryonic development in mammals, as mouse Top2a knockouts result in embryonic lethality as early as the 4-8 cell stage. The purpose of this study was to determine whether the extended developmental capability of zebrafish top2a mutants arises from maternal expression of top2a or compensation from its top2b paralogue. Results Here, we describe bloody minded (blm), a novel mutant of zebrafish top2a. In contrast to mouse Top2a nulls, zebrafish top2a mutants survive to larval stages (4-5 day post fertilization). Developmental analyses demonstrate abundant expression of maternal top2a but not top2b. Inhibition or poisoning of maternal topoisomerase II delays embryonic development by extending the cell cycle M-phase. Zygotic top2a and top2b are co-expressed in the zebrafish CNS, but endogenous or ectopic top2b RNA appear unable to prevent the blm phenotype. Conclusions We conclude that maternal top2a enables zebrafish development before the mid-zygotic transition (MZT) and that zebrafish top2a and top2b are not functionally redundant during development after activation of the zygotic genome. PMID:22111588

Cadherins in cerebellar development: translation of embryonic patterning into mature functional compartmentalization.

PubMed

Redies, Christoph; Neudert, Franziska; Lin, Juntang

2011-09-01

Cadherins are cell adhesion molecules with multiple morphogenic functions in brain development, for example, in neuroblast migration and aggregation, axon navigation, neural circuit formation, and synaptogenesis. More than 100 members of the cadherin superfamily are expressed in the developing and mature brain. Most of the cadherins investigated, in particular classic cadherins and δ-protocadherins, are expressed in the cerebellum. For several cadherin subtypes, expression begins at early embryonic stages and persists until mature stages of cerebellar development. At intermediate stages, distinct Purkinje cell clusters exhibit unique rostrocaudal and mediolateral expression profiles for each cadherin. In the chicken, mouse, and other species, the Purkinje cell clusters are separated by intervening raphes of migrating granule cells. This pattern of Purkinje cell clusters/raphes is, at least in part, continuous with the parasagittal striping pattern that is apparent in the mature cerebellar cortex, for example, for zebrin II/aldolase C. Moreover, subregions of the deep cerebellar nuclei, vestibular nuclei and the olivary complex also express cadherins differentially. Neuroanatomical evidence suggests that the nuclear subregions and cortical domains that express the same cadherin subtype are connected to each other, to form neural subcircuits of the cerebellar system. Cadherins thus provide a molecular code that specifies not only embryonic structures but also functional cerebellar compartmentalization. By following the implementation of this code, it can be revealed how mature functional architecture emerges from embryonic patterning during cerebellar development. Dysfunction of some cadherins is associated with psychiatric diseases and developmental impairments and may also affect cerebellar function.

Developmental cardiovascular physiology of the olive ridley sea turtle (Lepidochelys olivacea).

PubMed

Crossley, Dane Alan; Crossley, Janna Lee; Smith, Camilla; Harfush, Martha; Sánchez-Sánchez, Hermilo; Garduño-Paz, Mónica Vanessa; Méndez-Sánchez, José Fernando

2017-09-01

Our understanding of reptilian cardiovascular development and regulation has increased substantially for two species the American alligator (Alligator mississippiensis) and the common snapping turtle (Chelydra serpentina) during the past two decades. However, what we know about cardiovascular maturation in many other species remains poorly understood or unknown. Embryonic sea turtles have been studied to understand the maturation of metabolic function, but these studies have not addressed the cardiovascular system. Although prior studies have been pivotal in characterizing development, and factors that influence it, the development of cardiovascular function, which supplies metabolic function, is unknown in sea turtles. During our investigation we focused on quantifying how cardiovascular morphological and functional parameters change, to provide basic knowledge of development in the olive ridley sea turtle (Lepidochelys olivacea). Embryonic mass, as well as mass of the heart, lungs, liver, kidney, and brain increased during turtle embryo development. Although heart rate was constant during this developmental period, arterial pressure approximately doubled. Further, while embryonic olive ridley sea turtles lacked cholinergic tone on heart rate, there was a pronounced beta adrenergic tone on heart rate that decreased in strength at 90% of incubation. This beta adrenergic tone may be partially originating from the sympathetic nervous system at 90% of incubation, with the majority originating from circulating catecholamines. Data indicates that olive ridley sea turtles share traits of embryonic functional cardiovascular maturation with the American alligator (Alligator mississippiensis) but not the common snapping turtle (Chelydra serpentina). Copyright © 2017 Elsevier Inc. All rights reserved.

Uterine molecular changes for non-invasive embryonic attachment in the marsupials Macropus eugenii (Macropodidae) and Trichosurus vulpecula (Phalangeridae).

PubMed

Laird, Melanie K; Dargan, Jessica R; Paterson, Lillian; Murphy, Christopher R; McAllan, Bronwyn M; Shaw, Geoff; Renfree, Marilyn B; Thompson, Michael B

2017-10-01

Pregnancy in mammals requires remodeling of the uterus to become receptive to the implanting embryo. Remarkably similar morphological changes to the uterine epithelium occur in both eutherian and marsupial mammals, irrespective of placental type. Nevertheless, molecular differences in uterine remodeling indicate that the marsupial uterus employs maternal defences, including molecular reinforcement of the uterine epithelium, to regulate embryonic invasion. Non-invasive (epitheliochorial) embryonic attachment in marsupials likely evolved secondarily from invasive attachment, so uterine defences in these species may prevent embryonic invasion. We tested this hypothesis by identifying localization patterns of Talin, a key basal anchoring molecule, in the uterine epithelium during pregnancy in the tammar wallaby (Macropus eugenii; Macropodidae) and the brush tail possum (Trichosurus vulpecula; Phalangeridae). Embryonic attachment is non-invasive in both species, yet Talin undergoes a clear distributional change during pregnancy in M. eugenii, including recruitment to the base of the uterine epithelium just before attachment, that closely resembles that of invasive implantation in the marsupial species Sminthopsis crassicaudata. Basal localization occurs throughout pregnancy in T. vulpecula, although, as for M. eugenii, this pattern is most specific prior to attachment. Such molecular reinforcement of the uterine epithelium for non-invasive embryonic attachment in marsupials supports the hypothesis that less-invasive and non-invasive embryonic attachment in marsupials may have evolved via accrual of maternal defences. Recruitment of basal molecules, including Talin, to the uterine epithelium may have played a key role in this transition. © 2017 Wiley Periodicals, Inc.

The plurennial life cycles of the European Tettigoniidae (Insecta: Orthoptera) : 3. The effect of drought and the variable duration of the initial diapause.

PubMed

Ingrisch, Sigfrid

1986-11-01

The effect of drought on embryonic development and on hatching was studied in 13 European Tettigoniidae species. Drought can affect development in three different ways: (1) Embryonic development proceeds slower than if the eggs are in contact with water; (2) it stops (for final diapause) in an earlier embryonic stage; (3) it affects maintenance and termination of the initial embryonic diapause.In many Tettigoniidae species, the initial diapause is prolonged, and may last several years. Without draught stress, between 1 and 7 cold treatments in the laboratory, and with eggs of the Tettigonia-species between 1 and 6 winters in the field were necessary to enable all eggs to complete initial diapause. In Central European species, the number of eggs maintaining initial diapause significantly increased when the eggs had no contact with water at the time when they should recover from diapause. In contrast, termination of initial diapause in Tettigonia caudata from Greece, when the environment became favorable for growth again, was highest in that group of eggs that had lost most water in a preceding period of drought. The importance of the prolonged initial diapause for the survival of unpredictable adverse conditions is discussed.

Age-specific function of α5β1 integrin in microglial migration during early colonization of the developing mouse cortex.

PubMed

Smolders, Sophie Marie-Thérèse; Swinnen, Nina; Kessels, Sofie; Arnauts, Kaline; Smolders, Silke; Le Bras, Barbara; Rigo, Jean-Michel; Legendre, Pascal; Brône, Bert

2017-07-01

Microglia, the immune cells of the central nervous system, take part in brain development and homeostasis. They derive from primitive myeloid progenitors that originate in the yolk sac and colonize the brain mainly through intensive migration. During development, microglial migration speed declines which suggests that their interaction with the microenvironment changes. However, the matrix-cell interactions allowing dispersion within the parenchyma are unknown. Therefore, we aimed to better characterize the migration behavior and to assess the role of matrix-integrin interactions during microglial migration in the embryonic brain ex vivo. We focused on microglia-fibronectin interactions mediated through the fibronectin receptor α5β1 integrin because in vitro work indirectly suggested a role for this ligand-receptor pair. Using 2-photon time-lapse microscopy on acute ex vivo embryonic brain slices, we found that migration occurs in a saltatory pattern and is developmentally regulated. Most importantly, there is an age-specific function of the α5β1 integrin during microglial cortex colonization. At embryonic day (E) 13.5, α5β1 facilitates migration while from E15.5, it inhibits migration. These results indicate a developmentally regulated function of α5β1 integrin in microglial migration during colonization of the embryonic brain. © 2017 Wiley Periodicals, Inc.

Application of Carnegie stages of development to unify human and baboon ultrasound findings early in pregnancy.

PubMed

Santolaya-Forgas, Joaquin; De Leon-Luis, Juan; Friel, Lara A; Wolf, Roman

2007-09-01

The objective of this study was to determine if very early ultrasonographic measurements obtained from human and baboon are comparable. For this purpose, the gestational, amniotic and yolk sacs, embryonic crown rump length (CRL) and heart rate were measured ultrasonographically between 35 and 47 days from the mean day of a three-day mating period in baboons (n=18) and between 42 to 58 days from fertilization as calculated from the CRL measurements in human pregnancies (n=82). Ultrasonographic measurements from both species were then plotted in the same graph using Carnegie stages of embryonic development as the independent variable to allow for visual comparisons. Mean gestational age at ultrasonographic studies was significantly different for humans and baboons (50.4 vs. 41 days, respectively; p>0.01). Significant correlations (p>0.01) were noted between ultrasonographic measurements and Carnegie stages of development in both humans and baboons. Only the gestational and the yolk sacs were significantly smaller in baboons than in humans (p>0.05). The findings that embryonic CRL, extra-embryonic space and heart rate are very similar between the 17th and 23rd Carnegie developmental stages make the baboon a promising surrogate of human pregnancy for investigations using celocentesis.

Transcriptomic and anatomical complexity of primary, seminal, and crown roots highlight root type-specific functional diversity in maize (Zea mays L.)

PubMed Central

Tai, Huanhuan; Lu, Xin; Opitz, Nina; Marcon, Caroline; Paschold, Anja; Lithio, Andrew; Nettleton, Dan; Hochholdinger, Frank

2016-01-01

Maize develops a complex root system composed of embryonic and post-embryonic roots. Spatio-temporal differences in the formation of these root types imply specific functions during maize development. A comparative transcriptomic study of embryonic primary and seminal, and post-embryonic crown roots of the maize inbred line B73 by RNA sequencing along with anatomical studies were conducted early in development. Seminal roots displayed unique anatomical features, whereas the organization of primary and crown roots was similar. For instance, seminal roots displayed fewer cortical cell files and their stele contained more meta-xylem vessels. Global expression profiling revealed diverse patterns of gene activity across all root types and highlighted the unique transcriptome of seminal roots. While functions in cell remodeling and cell wall formation were prominent in primary and crown roots, stress-related genes and transcriptional regulators were over-represented in seminal roots, suggesting functional specialization of the different root types. Dynamic expression of lignin biosynthesis genes and histochemical staining suggested diversification of cell wall lignification among the three root types. Our findings highlight a cost-efficient anatomical structure and a unique expression profile of seminal roots of the maize inbred line B73 different from primary and crown roots. PMID:26628518

Asynchronous Replication and Autosome-Pair Non-Equivalence in Human Embryonic Stem Cells

PubMed Central

Dutta, Devkanya; Ensminger, Alexander W.; Zucker, Jacob P.; Chess, Andrew

2009-01-01

A number of mammalian genes exhibit the unusual properties of random monoallelic expression and random asynchronous replication. Such exceptional genes include genes subject to X inactivation and autosomal genes including odorant receptors, immunoglobulins, interleukins, pheromone receptors, and p120 catenin. In differentiated cells, random asynchronous replication of interspersed autosomal genes is coordinated at the whole chromosome level, indicative of chromosome-pair non-equivalence. Here we have investigated the replication pattern of the random asynchronously replicating genes in undifferentiated human embryonic stem cells, using fluorescence in situ hybridization based assay. We show that allele-specific replication of X-linked genes and random monoallelic autosomal genes occur in human embryonic stem cells. The direction of replication is coordinated at the whole chromosome level and can cross the centromere, indicating the existence of autosome-pair non-equivalence in human embryonic stem cells. These results suggest that epigenetic mechanism(s) that randomly distinguish between two parental alleles are emerging in the cells of the inner cell mass, the source of human embryonic stem cells. PMID:19325893

Transport of organic anions and cations in murine embryonic kidney development and in serially-reaggregated engineered kidneys

PubMed Central

Lawrence, Melanie L.; Chang, C-Hong; Davies, Jamie A.

2015-01-01

Recent advances in renal tissue engineering have shown that dissociated, early renogenic tissue from the developing embryo can self-assemble into morphologically accurate kidney-like organs arranged around a central collecting duct tree. In order for such self-assembled kidneys to be useful therapeutically or as models for drug screening, it is necessary to demonstrate that they are functional. One of the main functional characteristics of mature kidneys is transport of organic anions and cations into and out of the proximal tubule. Here, we show that the transport function of embryonic kidneys allowed to develop in culture follows a developmental time-course that is comparable to embryonic kidney development in vivo. We also demonstrate that serially-reaggregated engineered kidneys can transport organic anions and cations through specific uptake and efflux channels. These results support the physiological relevance of kidneys grown in culture, a commonly used model for kidney development and research, and suggest that serially-reaggregated kidneys self-assembled from separated cells have some functional characteristics of intact kidneys. PMID:25766625

Mechanical preconditioning enables electrophysiologic coupling of skeletal myoblast cells to myocardium

PubMed Central

Treskes, Philipp; Cowan, Douglas B.; Stamm, Christof; Rubach, Martin; Adelmann, Roland; Wittwer, Thorsten; Wahlers, Thorsten

2015-01-01

Objective The effect of mechanical preconditioning on skeletal myoblasts in engineered tissue constructs was investigated to resolve issues associated with conduction block between skeletal myoblast cells and cardiomyocytes. Methods Murine skeletal myoblasts were used to generate engineered tissue constructs with or without application of mechanical strain. After in vitro myotube formation, engineered tissue constructs were co-cultured for 6 days with viable embryonic heart slices. With the use of sharp electrodes, electrical coupling between engineered tissue constructs and embryonic heart slices was assessed in the presence or absence of pharmacologic agents. Results The isolation and expansion procedure for skeletal myoblasts resulted in high yields of homogeneously desmin-positive (97.1% ± 0.1%) cells. Mechanical strain was exerted on myotubes within engineered tissue constructs during gelation of the matrix, generating preconditioned engineered tissue constructs. Electrical coupling between preconditioned engineered tissue constructs and embryonic heart slices was observed; however, no coupling was apparent when engineered tissue constructs were not subjected to mechanical strain. Coupling of cells from engineered tissue constructs to cells in embryonic heart slices showed slower conduction velocities than myocardial cells with the embryonic heart slices (preconditioned engineered tissue constructs vs embryonic heart slices: 0.04 ± 0.02 ms vs 0.10 ± 0.05 ms, P = .011), lower stimulation frequencies (preconditioned engineered tissue constructs vs maximum embryonic heart slices: 4.82 ± 1.42 Hz vs 10.58 ± 1.56 Hz; P = .0009), and higher sensitivities to the gap junction inhibitor (preconditioned engineered tissue constructs vs embryonic heart slices: 0.22 ± 0.07 mmol/L vs 0.93 ± 0.15 mmol/L; P = .0004). Conclusions We have generated skeletal myoblast–based transplantable grafts that electrically couple to myocardium. PMID:22980065

The effects of an environmentally relevant 58-congener polychlorinated biphenyl (PCB) mixture on cardiac development in the chick embryo.

PubMed

Carro, Tiffany; Taneyhill, Lisa A; Ann Ottinger, Mary

2013-06-01

Chicken (Gallus domesticus) embryonic exposure in ovo to a 58-congener polychlorinated biphenyl (PCB) mixture resulted in teratogenic heart defects in chick embryos at critical heart developmental stages Hamburger-Hamilton (HH) stages 10, 16, and 20. The 58-congener mixture contained relative proportions of primary congeners measured in belted sandpiper (Megaceryle alcyon) and spotted sandpiper (Actitis macularia) eggs collected along the upper Hudson River, New York, USA, and chicken doses were well below observed environmental exposure levels. Embryos were injected with 0.08 µg PCBs/g egg weight and 0.50 µg PCBs/g egg weight (0.01 and 0.064 ng toxic equivalent/g, respectively) at embryonic day 0, prior to incubation. Mortality of exposed embryos was increased at all developmental stages, with a marked rise in cardiomyopathies at HH16 and HH20 (p < 0.05). Heart abnormalities occurred across all treatments, including abnormal elongation and expansion of the heart tube at HH10, improper looping and orientation, indentations in the emerging ventricular wall (HH16 and HH20), and irregularities in overall heart shape (HH10, HH16, and HH20). Histology was conducted on 2 cardiac proteins critical to embryonic heart development, ventricular myosin heavy chain and titin, to investigate potential mechanistic effects of PCBs on heart development, but no difference was observed in spatiotemporal expression. Similarly, cellular apoptosis in the developing heart was not affected by exposure to the PCB mixture. Conversely, cardiomyocyte proliferation rates dramatically declined (p < 0.01) at HH16 and HH20 as PCB exposure concentrations increased. Early embryonic cardiomyocyte proliferation contributes to proper formation of the morphology and overall thickness of the ventricular wall. Therefore, in ovo exposure to this 58-congener PCB mixture at critical stages adversely affects embryonic heart development. Copyright © 2013 SETAC.

Rhein Induces Oxidative Stress and Apoptosis in Mouse Blastocysts and Has Immunotoxic Effects during Embryonic Development.

PubMed

Huang, Chien-Hsun; Chan, Wen-Hsiung

2017-09-20

Rhein, a glucoside chemical compound found in a traditional Chinese medicine derived from the roots of rhubarb, induces cell apoptosis and is considered to have high potential as an antitumor drug. Several previous studies showed that rhein can inhibit cell proliferation and trigger mitochondria-related or endoplasmic reticulum (ER) stress-dependent apoptotic processes. However, the side effects of rhein on pre- and post-implantation embryonic development remain unclear. Here, we show that rhein has cytotoxic effects on blastocyst-stage mouse embryos and induces oxidative stress and immunotoxicity in mouse fetuses. Blastocysts incubated with 5-20 μM rhein showed significant cell apoptosis, as well as decreases in their inner cell mass cell numbers and total cell numbers. An in vitro development assay showed that rhein affected the developmental potentials of both pre- and post-implantation embryos. Incubation of blastocysts with 5-20 μM rhein was associated with increased resorption of post-implantation embryos and decreased fetal weight in an embryo transfer assay. Importantly, in an in vivo model, intravenous injection of dams with rhein (1, 3, and 5 mg/kg body weight/day) for four days resulted in apoptosis of blastocyst-stage embryos, early embryonic developmental injury, and decreased fetal weight. Intravenous injection of dams with 5 mg/kg body weight/day rhein significantly increased the total reactive oxygen species (ROS) content of fetuses and the transcription levels of antioxidant proteins in fetal livers. Additional work showed that rhein induced apoptosis through ROS generation, and that prevention of apoptotic processes effectively rescued the rhein-induced injury effects on embryonic development. Finally, the transcription levels of the innate-immunity related genes, CXCL1 , IL-1 β and IL-8 , were down-regulated in the fetuses of dams that received intravenous injections of rhein. These results collectively show that rhein has the potential to induce embryonic cytotoxicity and induce oxidative stress and immunotoxicity during the development of mouse embryos.

Defining the molecular pathologies in cloaca malformation: similarities between mouse and human

PubMed Central

Runck, Laura A.; Method, Anna; Bischoff, Andrea; Levitt, Marc; Peña, Alberto; Collins, Margaret H.; Gupta, Anita; Shanmukhappa, Shiva; Wells, James M.; Guasch, Géraldine

2014-01-01

Anorectal malformations are congenital anomalies that form a spectrum of disorders, from the most benign type with excellent functional prognosis, to very complex, such as cloaca malformation in females in which the rectum, vagina and urethra fail to develop separately and instead drain via a single common channel into the perineum. The severity of this phenotype suggests that the defect occurs in the early stages of embryonic development of the organs derived from the cloaca. Owing to the inability to directly investigate human embryonic cloaca development, current research has relied on the use of mouse models of anorectal malformations. However, even studies of mouse embryos lack analysis of the earliest stages of cloaca patterning and morphogenesis. Here we compared human and mouse cloaca development and retrospectively identified that early mis-patterning of the embryonic cloaca might underlie the most severe forms of anorectal malformation in humans. In mouse, we identified that defective sonic hedgehog (Shh) signaling results in early dorsal-ventral epithelial abnormalities prior to the reported defects in septation. This is manifested by the absence of Sox2 and aberrant expression of keratins in the embryonic cloaca of Shh knockout mice. Shh knockout embryos additionally develop a hypervascular stroma, which is defective in BMP signaling. These epithelial and stromal defects persist later, creating an indeterminate epithelium with molecular alterations in the common channel. We then used these animals to perform a broad comparison with patients with mild-to-severe forms of anorectal malformations including cloaca malformation. We found striking parallels with the Shh mouse model, including nearly identical defective molecular identity of the epithelium and surrounding stroma. Our work strongly suggests that early embryonic cloacal epithelial differentiation defects might be the underlying cause of severe forms of anorectal malformations in humans. Moreover, deranged Shh and BMP signaling is correlated with severe anorectal malformations in both mouse and humans. PMID:24524909

Microbial population analysis of broilers in different flock and embryonic ages from GI, yolk and egg wash via Illumina MiSeq and QIIME pipeline (abstract)

USDA-ARS?s Scientific Manuscript database

There are several food-safety issues related to broiler egg production, including the introduction/proliferation of zoonotic pathogens during embryonic gastrointestinal (GI) tract development. Little is known about the overall GI bacterial communities, how they change over time, or how their composi...

A MICROARRAY ANALYSIS OF GENE EXPRESSION IN THE EMBRYONIC FORELIMB OF THE C57BL/6J MOUSE REVEALS SIGNIFICANT ALTERATIONS METABOLIC AND DEVELOPMENTAL REGULATION FOLLOWING ETHANOL EXPOSURE.

EPA Science Inventory

The observation of transcriptional changes following embryonic ethanol exposure may provide significant insights into the biological response to ethanol exposure. In this study, we used microarray analysis to examine the transcriptional response of the developing limb to a dose ...

Dynamic Proteomic Analysis of Pancreatic Mesenchyme Reveals Novel Factors That Enhance Human Embryonic Stem Cell to Pancreatic Cell Differentiation.

PubMed

Russ, Holger A; Landsman, Limor; Moss, Christopher L; Higdon, Roger; Greer, Renee L; Kaihara, Kelly; Salamon, Randy; Kolker, Eugene; Hebrok, Matthias

2016-01-01

Current approaches in human embryonic stem cell (hESC) to pancreatic beta cell differentiation have largely been based on knowledge gained from developmental studies of the epithelial pancreas, while the potential roles of other supporting tissue compartments have not been fully explored. One such tissue is the pancreatic mesenchyme that supports epithelial organogenesis throughout embryogenesis. We hypothesized that detailed characterization of the pancreatic mesenchyme might result in the identification of novel factors not used in current differentiation protocols. Supplementing existing hESC differentiation conditions with such factors might create a more comprehensive simulation of normal development in cell culture. To validate our hypothesis, we took advantage of a novel transgenic mouse model to isolate the pancreatic mesenchyme at distinct embryonic and postnatal stages for subsequent proteomic analysis. Refined sample preparation and analysis conditions across four embryonic and prenatal time points resulted in the identification of 21,498 peptides with high-confidence mapping to 1,502 proteins. Expression analysis of pancreata confirmed the presence of three potentially important factors in cell differentiation: Galectin-1 (LGALS1), Neuroplastin (NPTN), and the Laminin α-2 subunit (LAMA2). Two of the three factors (LGALS1 and LAMA2) increased expression of pancreatic progenitor transcript levels in a published hESC to beta cell differentiation protocol. In addition, LAMA2 partially blocks cell culture induced beta cell dedifferentiation. Summarily, we provide evidence that proteomic analysis of supporting tissues such as the pancreatic mesenchyme allows for the identification of potentially important factors guiding hESC to pancreas differentiation.

Dynamic Proteomic Analysis of Pancreatic Mesenchyme Reveals Novel Factors That Enhance Human Embryonic Stem Cell to Pancreatic Cell Differentiation

PubMed Central

Russ, Holger A.; Landsman, Limor; Moss, Christopher L.; Higdon, Roger; Greer, Renee L.; Kaihara, Kelly; Salamon, Randy; Kolker, Eugene; Hebrok, Matthias

2016-01-01

Current approaches in human embryonic stem cell (hESC) to pancreatic beta cell differentiation have largely been based on knowledge gained from developmental studies of the epithelial pancreas, while the potential roles of other supporting tissue compartments have not been fully explored. One such tissue is the pancreatic mesenchyme that supports epithelial organogenesis throughout embryogenesis. We hypothesized that detailed characterization of the pancreatic mesenchyme might result in the identification of novel factors not used in current differentiation protocols. Supplementing existing hESC differentiation conditions with such factors might create a more comprehensive simulation of normal development in cell culture. To validate our hypothesis, we took advantage of a novel transgenic mouse model to isolate the pancreatic mesenchyme at distinct embryonic and postnatal stages for subsequent proteomic analysis. Refined sample preparation and analysis conditions across four embryonic and prenatal time points resulted in the identification of 21,498 peptides with high-confidence mapping to 1,502 proteins. Expression analysis of pancreata confirmed the presence of three potentially important factors in cell differentiation: Galectin-1 (LGALS1), Neuroplastin (NPTN), and the Laminin α-2 subunit (LAMA2). Two of the three factors (LGALS1 and LAMA2) increased expression of pancreatic progenitor transcript levels in a published hESC to beta cell differentiation protocol. In addition, LAMA2 partially blocks cell culture induced beta cell dedifferentiation. Summarily, we provide evidence that proteomic analysis of supporting tissues such as the pancreatic mesenchyme allows for the identification of potentially important factors guiding hESC to pancreas differentiation. PMID:26681951

Organizational Changes to Thyroid Regulation in Alligator mississippiensis: Evidence for Predictive Adaptive Responses

PubMed Central

Boggs, Ashley S. P.; Lowers, Russell H.; Cloy-McCoy, Jessica A.; Guillette, Louis J.

2013-01-01

During embryonic development, organisms are sensitive to changes in thyroid hormone signaling which can reset the hypothalamic-pituitary-thyroid axis. It has been hypothesized that this developmental programming is a ‘predictive adaptive response’, a physiological adjustment in accordance with the embryonic environment that will best aid an individual's survival in a similar postnatal environment. When the embryonic environment is a poor predictor of the external environment, the developmental changes are no longer adaptive and can result in disease states. We predicted that endocrine disrupting chemicals (EDCs) and environmentally-based iodide imbalance could lead to developmental changes to the thyroid axis. To explore whether iodide or EDCs could alter developmental programming, we collected American alligator eggs from an estuarine environment with high iodide availability and elevated thyroid-specific EDCs, a freshwater environment contaminated with elevated agriculturally derived EDCs, and a reference freshwater environment. We then incubated them under identical conditions. We examined plasma thyroxine and triiodothyronine concentrations, thyroid gland histology, plasma inorganic iodide, and somatic growth at one week (before external nutrition) and ten months after hatching (on identical diets). Neonates from the estuarine environment were thyrotoxic, expressing follicular cell hyperplasia (p = 0.01) and elevated plasma triiodothyronine concentrations (p = 0.0006) closely tied to plasma iodide concentrations (p = 0.003). Neonates from the freshwater contaminated site were hypothyroid, expressing thyroid follicular cell hyperplasia (p = 0.01) and depressed plasma thyroxine concentrations (p = 0.008). Following a ten month growth period under identical conditions, thyroid histology (hyperplasia p = 0.04; colloid depletion p = 0.01) and somatic growth (body mass p<0.0001; length p = 0.02) remained altered among the contaminated sites. This work supports the hypothesis that embryonic EDC exposure or iodide imbalance could induce adult metabolic disease states, thereby stressing the need to consider the multiple environmental variables present during development. PMID:23383213

Parthenogenesis in non-rodent species: developmental competence and differentiation plasticity.

PubMed

Brevini, T A L; Pennarossa, G; Vanelli, A; Maffei, S; Gandolfi, F

2012-03-01

An oocyte can activate its developmental process without the intervention of the male counterpart. This form of reproduction, known as parthenogenesis, occurs spontaneously in a variety of lower organisms, but not in mammals. However, it must be noted that mammalian oocytes can be activated in vitro, mimicking the intracellular calcium wave induced by the spermatozoon at fertilization, which triggers cleavage divisions and embryonic development. The resultant parthenotes are not capable of developing to term and arrest their growth at different stages, depending on the species. It is believed that this arrest is due to genomic imprinting, which causes the repression of genes normally expressed by the paternal allele. Human parthenogenetic embryos have recently been proposed as an alternative, less controversial source of embryonic stem cell lines, based on their inherent inability to form a new individual. However many aspects related to the biology of parthenogenetic embryos and parthenogenetically derived cell lines still need to be elucidated. Limited information is available in particular on the consequences of the lack of centrioles and on the parthenote's ability to assemble a new embryonic centrosome in the absence of the sperm centriole. Indeed, in lower species, successful parthenogenesis largely depends upon the oocyte's ability to regenerate complete and functional centrosomes in the absence of the material supplied by a male gamete, while the control of this event appears to be less stringent in mammalian cells. In an attempt to better elucidate some of these aspects, parthenogenetic cell lines, recently derived in our laboratory, have been characterized for their pluripotency. In vitro and in vivo differentiation plasticity have been assessed, demonstrating the ability of these cells to differentiate into cell types derived from the three germ layers. These results confirmed common features between uni- and bi-parental embryonic stem cells. However data obtained with parthenogenetic cells indicate the presence of an intrinsic deregulation of the mechanisms controlling proliferation vs. differentiation and suggest their uni-parental origin as a possible cause. Copyright © 2012 Elsevier Inc. All rights reserved.

When Is Embryonic Arrest Broken in Turtle Eggs?

PubMed

Williamson, Sean A; Evans, Roger G; Reina, Richard D

Turtle embryos enter a state of arrested development in the oviduct, allowing the mother greater flexibility in her reproductive schedule. Development recommences once eggs transition from the hypoxic oviduct to the normoxic nest. Significant mortality can occur if turtle eggs are moved between 12 h and 20 d after oviposition, and this is linked to the recommencement of embryonic development. To better understand the timing of developmental arrest and to determine how movement-induced mortality might be avoided, we determined the latency (i.e., time elapsed since oviposition) to recommencement of development following oviposition by exposing the eggs of green turtles (Chelonia mydas) to hypoxia (oxygen tension <8 mmHg) for 3 d, commencing 30 min to 48 h after oviposition. Embryonic development-including development of the characteristic opaque white spot on the eggshell-was halted by hypoxic incubation. When the delay before hypoxic incubation was 12 h or less, hatching success did not differ from a control group. If the hypoxic treatment began after 16 h or more in normoxia, then all embryos died. Thus, by returning eggs to a hypoxic environment before they have broken from arrest (i.e., within 12 h of oviposition), it is possible to extend embryonic arrest for at least 3 d, with no apparent detriment to hatching success. Therefore, hypoxic incubation may provide a new approach for avoidance of movement-induced mortality when conservation or research efforts require the relocation of eggs. Our findings also suggest that movement-induced mortality may have constrained the evolution of viviparity in turtles.

Expression analysis of the insulin-like growth factors I and II during embryonic and early larval development of turbot ( Scophthalmus maximus)

NASA Astrophysics Data System (ADS)

Wen, Haishen; Qi, Qian; Hu, Jian; Si, Yufeng; He, Feng; Li, Jifang

2015-04-01

The insulin-like growth factors I and II (IGF-I and IGF-II) are important proteins involved in fish growth and development. Here, we report the isolation of IGF-II and expression analysis of IGFs in turbot Scophthalmus maximus, aiming to clarify their function in embryonic and larval development of fish. The deduced IGF-II gene is 808 bp in full length, which encodes a protein of 219 amino acids and is 93% similar with that of Paralichthys olicaceus in amino acid sequence. The tissue abundance and the expression pattern of IGFs in a turbot at early development stages were investigated via reverse transcription-polymer chain reaction. Result showed that the IGF-I and IGF-II genes were widely expressed in tissues of S. maximus. IGF-I was detected in all tissues except intestines with the highest level in liver, while IGF-II transcript presented in all tissues except muscle. At the stages of embryonic and larval development, the mRNA levels of IGFs sharply increased from the stage of unfertilized egg to post larva, followed by a decrease with larval development. However, there was an increase in IGF-I at the embryonic stage and IGF-II at the gastrula stage, respectively. These results suggested that IGFs play important roles in cell growth and division of the turbot. Our study provides reference data for further investigation of growth regulation in turbot, which can guarantee better understanding of the physiological role that IGFs play in fish.

Postnatal growth rates covary weakly with embryonic development rates and do not explain adult mortality probability among songbirds on four continents.

PubMed

Martin, Thomas E; Oteyza, Juan C; Mitchell, Adam E; Potticary, Ahva L; Lloyd, Penn

2015-03-01

Growth and development rates may result from genetic programming of intrinsic processes that yield correlated rates between life stages. These intrinsic rates are thought to affect adult mortality probability and longevity. However, if proximate extrinsic factors (e.g., temperature, food) influence development rates differently between stages and yield low covariance between stages, then development rates may not explain adult mortality probability. We examined these issues based on study of 90 songbird species on four continents to capture the diverse life-history strategies observed across geographic space. The length of the embryonic period explained little variation (ca. 13%) in nestling periods and growth rates among species. This low covariance suggests that the relative importance of intrinsic and extrinsic influences on growth and development rates differs between stages. Consequently, nestling period durations and nestling growth rates were not related to annual adult mortality probability among diverse songbird species within or among sites. The absence of a clear effect of faster growth on adult mortality when examined in an evolutionary framework across species may indicate that species that evolve faster growth also evolve physiological mechanisms for ameliorating costs on adult mortality. Instead, adult mortality rates of species in the wild may be determined more strongly by extrinsic environmental causes.

Postnatal growth rates covary weakly with embryonic development rates and do not explain adult mortality probability among songbirds on four continents

USGS Publications Warehouse

Martin, Thomas E.; Oteyza, Juan C.; Mitchell, Adam E.; Potticary, Ahva L.; Lloyd, P.

2016-01-01

Growth and development rates may result from genetic programming of intrinsic processes that yield correlated rates between life stages. These intrinsic rates are thought to affect adult mortality probability and longevity. However, if proximate extrinsic factors (e.g., temperature, food) influence development rates differently between stages and yield low covariance between stages, then development rates may not explain adult mortality probability. We examined these issues based on study of 90 songbird species on four continents to capture the diverse life-history strategies observed across geographic space. The length of the embryonic period explained little variation (ca. 13%) in nestling periods and growth rates among species. This low covariance suggests that the relative importance of intrinsic and extrinsic influences on growth and development rates differs between stages. Consequently, nestling period durations and nestling growth rates were not related to annual adult mortality probability among diverse songbird species within or among sites. The absence of a clear effect of faster growth on adult mortality when examined in an evolutionary framework across species may indicate that species that evolve faster growth also evolve physiological mechanisms for ameliorating costs on adult mortality. Instead, adult mortality rates of species in the wild may be determined more strongly by extrinsic environmental causes.

Let-7 represses Nr6a1 and a mid-gestation developmental program in adult fibroblasts

PubMed Central

Gurtan, Allan M.; Ravi, Arvind; Rahl, Peter B.; Bosson, Andrew D.; JnBaptiste, Courtney K.; Bhutkar, Arjun; Whittaker, Charles A.; Young, Richard A.; Sharp, Phillip A.

2013-01-01

MicroRNAs (miRNAs) are critical to proliferation, differentiation, and development. Here, we characterize gene expression in murine Dicer-null adult mesenchymal stem cell lines, a fibroblast cell type. Loss of Dicer leads to derepression of let-7 targets at levels that exceed 10-fold to 100-fold with increases in transcription. Direct and indirect targets of this miRNA belong to a mid-gestation embryonic program that encompasses known oncofetal genes as well as oncogenes not previously associated with an embryonic state. Surprisingly, this mid-gestation program represents a distinct period that occurs between the pluripotent state of the inner cell mass at embryonic day 3.5 (E3.5) and the induction of let-7 upon differentiation at E10.5. Within this mid-gestation program, we characterize the let-7 target Nr6a1, an embryonic transcriptional repressor that regulates gene expression in adult fibroblasts following miRNA loss. In total, let-7 is required for the continual suppression of embryonic gene expression in adult cells, a mechanism that may underlie its tumor-suppressive function. PMID:23630078

Parthenogenesis-derived Multipotent Stem Cells Adapted for Tissue Engineering Applications

PubMed Central

Koh, Chester J.; Delo, Dawn M.; Lee, Jang Won; Siddiqui, M. Minhaj; Lanza, Robert P.; Soker, Shay; Yoo, James J.; Atala, Anthony

2009-01-01

Embryonic stem cells are envisioned as a viable source of pluripotent cells for use in regenerative medicine applications when donor tissue is not available. However, most current harvest techniques for embryonic stem cells require the destruction of embryos, which has led to significant political and ethical limitations on their usage. Parthenogenesis, the process by which an egg can develop into an embryo in the absence of sperm, may be a potential source of embryonic stem cells that may avoid some of the political and ethical concerns surrounding embryonic stem cells. Here we provide the technical aspects of embryonic stem cell isolation and expansion from the parthenogenetic activation of oocytes. These cells were characterized for their stem-cell properties. In addition, these cells were induced to differentiate to the myogenic, osteogenic, adipogenic, and endothelial lineages, and were able to form muscle-like and bony-like tissue in vivo. Furthermore, parthenogenetic stem cells were able to integrate into injured muscle tissue. Together, these results demonstrate that parthenogenetic stem cells can be successfully isolated and utilized for various tissue engineering applications. PMID:18799133

Development of the endolymphatic sac in chick embryos, with reference to the degradation of otoconia

NASA Technical Reports Server (NTRS)

Yoshihara, T.; Kaname, H.; Narita, N.; Ishii, T.; Igarashi, M.; Fermin, C. D.

1992-01-01

The endolymphatic sac of chick embryos (from embryonic day 7 to 1-day-old chicks) was studied light- and electron-microscopically. At stage 30-31 (embryonic day 7-7.5), the epithelial cells of the endolymphatic sac were cuboidal to columnar in shape. Microvilli were relatively well developed. The intercellular space was wide. In the endolymphatic space of the endolymphatic sac, varying shapes and sizes of otoconia-like bodies were often observed. Intracytoplasmic phagosomes containing these bodies were rarely found. After stage 37 (embryonic day 11), otoconia-like bodies in the endolymphatic sac decreased in number and size. They were almost the same as the otoconia in the macular organs, ultrastructurally. These findings indicate that the endolymphatic sac of the chick embryos may possess the function of otoconial degradation and removal of calcium from otoconia.

Comparison of NMDA and AMPA Channel Expression and Function between Embryonic and Adult Neurons Utilizing Microelectrode Array Systems.

PubMed

Edwards, Darin; Sommerhage, Frank; Berry, Bonnie; Nummer, Hanna; Raquet, Martina; Clymer, Brad; Stancescu, Maria; Hickman, James J

2017-12-11

Microelectrode arrays (MEAs) are innovative tools used to perform electrophysiological experiments for the study of electrical activity and connectivity in populations of neurons from dissociated cultures. Reliance upon neurons derived from embryonic tissue is a common limitation of neuronal/MEA hybrid systems and perhaps of neuroscience research in general, and the use of adult neurons could model fully functional in vivo parameters more closely. Spontaneous network activity was concurrently recorded from both embryonic and adult rat neurons cultured on MEAs for up to 10 weeks in vitro to characterize the synaptic connections between cell types. The cultures were exposed to synaptic transmission antagonists against NMDA and AMPA channels, which revealed significantly different receptor profiles of adult and embryonic networks in vitro. In addition, both embryonic and adult neurons were evaluated for NMDA and AMPA channel subunit expression over five weeks in vitro. The results established that neurons derived from embryonic tissue did not express mature synaptic channels for several weeks in vitro under defined conditions. Consequently, the embryonic response to synaptic antagonists was significantly different than that of neurons derived from adult tissue sources. These results are especially significant because most studies reported with embryonic hippocampal neurons do not begin at two to four weeks in culture. In addition, the utilization of MEAs in lieu of patch-clamp electrophysiology avoided a large-scale, labor-intensive study. These results establish the utility of this unique hybrid system derived from adult hippocampal tissue in combination with MEAs and offer a more appropriate representation of in vivo function for drug discovery. It has application for neuronal development and regeneration as well as for investigations into neurodegenerative disease, traumatic brain injury, and stroke.

Droplet barcoding for single cell transcriptomics applied to embryonic stem cells

PubMed Central

Klein, Allon M; Mazutis, Linas; Akartuna, Ilke; Tallapragada, Naren; Veres, Adrian; Li, Victor; Peshkin, Leonid; Weitz, David A; Kirschner, Marc W

2015-01-01

Summary It has long been the dream of biologists to map gene expression at the single cell level. With such data one might track heterogeneous cell sub-populations, and infer regulatory relationships between genes and pathways. Recently, RNA sequencing has achieved single cell resolution. What is limiting is an effective way to routinely isolate and process large numbers of individual cells for quantitative in-depth sequencing. We have developed a high-throughput droplet-microfluidic approach for barcoding the RNA from thousands of individual cells for subsequent analysis by next-generation sequencing. The method shows a surprisingly low noise profile and is readily adaptable to other sequencing-based assays. We analyzed mouse embryonic stem cells, revealing in detail the population structure and the heterogeneous onset of differentiation after LIF withdrawal. The reproducibility of these high-throughput single cell data allowed us to deconstruct cell populations and infer gene expression relationships. PMID:26000487