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Sample records for early chick embryo

  1. Method of Electroporation for the Early Chick Embryo

    NASA Astrophysics Data System (ADS)

    Hatakeyama, Jun; Shimamura, Kenji

    Chick embryos have long been one of the favored model systems in the field of embryology and developmental biology. Recent advances in the gene manipulation technologies (Muramatsu et al., 1997; Nakamura et al., 2004) make this model system even more attractive for the developmental biologists (see review by Stern, 2005). Thanks to its two dimensional geometry, easiness in accessibility and observation, and well-established fate maps (e.g. Couly and Le Douarin, 1988; Garcia-Martinez et al., 1993; Hatada and Stern, 1994; Psychoyos and Stern, 1996; Sawada and Aoyama, 1999; Cobos et al., 2001; Lopez-Sanchez et al., 2001; Redkar et al., 2001; Fernandez-Garre et al., 2002; Kimura et al., 2006; Matsushita et al., 2008), it has great advantages especially for studies at the early embryonic stages, such as the processes of gastrulation, neural induction, left-right patterning, etc. For such purposes, a whole embryo culture system, originally invented by Dennis A. T. New (New, 1955), and its derivatives (Flamme, 1987; Sundin and Eichele, 1992; Stern, 1993; Chapman et al., 2001) have been widely used.

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

    PubMed

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

    2016-06-15

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

  3. γ-BUTYROBETAINE AS A SPECIFIC ANTAGONIST FOR CARNITINE IN THE DEVELOPMENT OF THE EARLY CHICK EMBRYO

    PubMed Central

    Ito, Toshio; Fraenkel, G.

    1957-01-01

    The effect of γ-butyrobetaine alone and with the addition of carnitine on the development of the early excised chick embryo has been studied. γ-Butyrobetaine in appropriate amounts exerts an inhibitory effect which can be relieved or annulled by the inclusion of appropriate amounts of carnitine. This has been interpreted as a metabolite-antimetabolite relationship, in which the normal metabolite, carnitine, is antagonized by the structurally closely related γ-butyrobetaine, and is regarded as evidence of an important role of carnitine in the metabolism of the developing chick embryo. PMID:13475691

  4. The effects of levetiracetam on neural tube development in the early stage of chick embryos.

    PubMed

    Guvenc, Yahya; Dalgic, Ali; Billur, Deniz; Karaoglu, Derya; Aydin, Sevim; Daglioglu, Ergun; Ozdol, Cagatay; Nacar, Osman Arikan; Yildirim, Ali Erdem; Belen, Deniz

    2013-01-01

    This study aimed to investigate the effects of a new generation antiepileptic agent, levetiracetam, on the neural tube development in a chick embryo model that corresponds to the first month of vertebral development in mammals. Forty-five Atabey® breed fertilized chicken eggs with no specific pathogens were randomly divided into 5 groups. All of the eggs were incubated at 37.8±2°C and 60±5 % relative humidity in an incubator. Group A was control group. The other eggs were applied physiological saline and drugs at a volume of 10 μL by the in ovo method at the 28th hour of the incubation period. Group B was given distilled water; Group C, physiological saline; Group D, Levetiracetam (L8668) at a dose equivalent to the treatment dose for humans (10 mg/ kg), and Group E, Levetiracetam (L8668) at a dose of 10 times the treatment dose. The embryos in all of the groups were removed from the shells at the 48th hour and morphologically and histologically evaluated. Of the 45 embryos incubated, neural tubes of 41 were closed and the embryos displayed normal development. Levetiracetam, at a dose equivalent to human treatment dose and 10 times the treatment dose, was shown not to cause neural tube defects in chick embryos.

  5. The impact of high-salt exposure on cardiovascular development in the early chick embryo.

    PubMed

    Wang, Guang; Zhang, Nuan; Wei, Yi-Fan; Jin, Yi-Mei; Zhang, Shi-Yao; Cheng, Xin; Ma, Zheng-Lai; Zhao, Shu-Zhu; Chen, You-Peng; Chuai, Manli; Hocher, Berthold; Yang, Xuesong

    2015-11-01

    In this study, we show that high-salt exposure dramatically increases chick mortality during embryo development. As embryonic mortality at early stages mainly results from defects in cardiovascular development, we focused on heart formation and angiogenesis. We found that high-salt exposure enhanced the risk of abnormal heart tube looping and blood congestion in the heart chamber. In the presence of high salt, both ventricular cell proliferation and apoptosis increased. The high osmolarity induced by high salt in the ventricular cardiomyocytes resulted in incomplete differentiation, which might be due to reduced expression of Nkx2.5 and GATA4. Blood vessel density and diameter were suppressed by exposure to high salt in both the yolk sac membrane (YSM) and chorioallantoic membrane models. In addition, high-salt-induced suppression of angiogenesis occurred even at the vasculogenesis stage, as blood island formation was also inhibited by high-salt exposure. At the same time, cell proliferation was repressed and cell apoptosis was enhanced by high-salt exposure in YSM tissue. Moreover, the reduction in expression of HIF2 and FGF2 genes might cause high-salt-suppressed angiogenesis. Interestingly, we show that high-salt exposure causes excess generation of reactive oxygen species (ROS) in the heart and YSM tissues, which could be partially rescued through the addition of antioxidants. In total, our study suggests that excess generation of ROS might play an important role in high-salt-induced defects in heart and angiogenesis. © 2015. Published by The Company of Biologists Ltd.

  6. Optimization of CRISPR/Cas9 genome editing for loss-of-function in the early chick embryo.

    PubMed

    Gandhi, Shashank; Piacentino, Michael L; Vieceli, Felipe M; Bronner, Marianne E

    2017-12-01

    The advent of CRISPR/Cas9 has made genome editing possible in virtually any organism, including those not previously amenable to genetic manipulations. Here, we present an optimization of CRISPR/Cas9 for application to early avian embryos with improved efficiency via a three-fold strategy. First, we employed Cas9 protein flanked with two nuclear localization signal sequences for improved nuclear localization. Second, we used a modified guide RNA (gRNA) scaffold that obviates premature termination of transcription and unstable Cas9-gRNA interactions. Third, we used a chick-specific U6 promoter that yields 4-fold higher gRNA expression than the previously utilized human U6. For rapid screening of gRNAs for in vivo applications, we also generated a chicken fibroblast cell line that constitutively expresses Cas9. As proof of principle, we performed electroporation-based loss-of-function studies in the early chick embryo to knock out Pax7 and Sox10, key transcription factors with known functions in neural crest development. The results show that CRISPR/Cas9-mediated deletion causes loss of their respective proteins and transcripts, as well as predicted downstream targets. Taken together, the results reveal the utility of this optimized CRISPR/Cas9 method for targeted gene knockout in chicken embryos in a manner that is reproducible, robust and specific. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Expression of Wise in chick embryos.

    PubMed

    Shigetani, Y; Itasaki, N

    2007-08-01

    We have performed in situ hybridization to study the expression of Wise in early chick embryos. Wise expression is first detectable in the ectoderm at posterior levels of late neurula. As development proceeds, Wise expression is seen in specific patterns in the ectoderm of the trunk region, pharyngeal arches, limb buds, and feather buds. In addition to these areas, particular cartilages such as the ones in the maxillary process and limbs start to express Wise at the late pharyngula stage, and the expression in these cartilages becomes stronger than that in epidermal components at later stages. Importantly, Wise is expressed in regions where other signaling molecules such as Wnt, Bmp, and Shh are known to function in morphogenesis and differentiation. Direct comparisons of the expression of Wise and these genes are also demonstrated. (c) 2007 Wiley-Liss, Inc.

  8. Culturing Chick Embryos--A Simplification of New's Method.

    ERIC Educational Resources Information Center

    Downie, J. R.

    1979-01-01

    Describes a simplified version of New's method for culturing early chick embryos. The technique allows continuous observation of the critical first three days of development and the conditions for setting up successful cultures are also presented to help both teachers and students. (HM)

  9. [Calcium in the developing skeletal muscles of the chick embryo].

    PubMed

    Samosudova, N V; Enenko, S O; Larin, Iu S; Shungskaia, V E

    1982-07-01

    The osmium-pyroantimonate technique was used for the ultrastructural study of Ca2+-localization in two types of chick embryo skeletal muscles: m. pectoralis and m. soleus. In 8- and 12-day old embryos the pyroantimonate precipitate was found on plasmalemma, condensed chromatine and ribosomes and in N-lines of I-band. During myogenesis (15-, 21-day old embryos) the calcium precipitate is redistributed from the above mentioned sites to terminal cisternae and N-line of I-band. It is proposed that calcium of N-lines may be involved in the glycogenolysis, its association with the muscle contraction occurring particularly at early developmental stages.

  10. Short periods of incubation, egg turning during storage and broiler breeder hens age for early development of embryos, hatching results, chicks quality and juvenile growth.

    PubMed

    Damaziak, K; Paweska, M; Gozdowski, D; Niemiec, J

    2018-05-14

    An effect of modification of storage conditions of the eggs of broiler breeder flocks at the age of 49-, 52- and 70-, 73-wks of life on an early embryonic development, hatching time and synchronization, hatchability rates, chicks quality and broiler growth was investigated. The eggs were divided into 4 experimental groups: COI = eggs storage 5 d, at turning every 12 h; NSP = eggs storage 12 d, at turning every 12 h; SPIDES = were treated with 4 h pre-incubation at 30°C and 50-55% air humidity, delivered at 5 and 10 d over of 12 d of storage, and turning every 12 h; NCOI = eggs storage 12 d, no turning and no pre-incubation. Eggs from older hens were characterized by poorer hatchability and poorer chicks quality. The use of 2 × 4 h pre-incubation in 12 d of eggs storage could have an effect on the initial acceleration of embryonic development in eggs of young hens, contributing to the alignment of embryos development in eggs from young and older hens to 72 h of incubation. Pre-incubation had no effect on the length of incubation period, hatching window, but it increased the hatchability of the set and apparently fertilized eggs and decreased the number of eggs not hatched, and also improved chicks quality. Eggs turning by 90° every 12 h during the storage positively affected the embryonic development, shortening the incubation time and the quality of chicks, but had no effect on hatchability rates and body weight in 42 d of life. Based on the obtained results, it can be concluded that the applied modifications can be effective in counteracting the negative effects of storage of hatching eggs from both young and older birds.

  11. Antioxidant activities of chick embryo egg hydrolysates

    PubMed Central

    Sun, Hao; Ye, Ting; Wang, Yuntao; Wang, Ling; Chen, Yijie; Li, Bin

    2014-01-01

    Chick embryo egg hydrolysates (CEEH) were obtained by enzymatic hydrolysis of chick embryo egg in vitro-simulated gastrointestinal digestion. The antioxidant activities of CEEH were investigated by employing three in vitro assays, including the 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonate)/1,1-diphenyl-2-picrylhydrazyl (ABTS/DPPH)/hydroxyl radical-scavenging assays. The radical-scavenging effect of CEEH (1.0 mg/mL) was in a dose-dependent manner, with the highest trolox equivalent antioxidant capacity for ABTS, DPPH, and that of hydroxyl radicals found to be 569, 2097, and 259.6 μmol/L, respectively; whereas the trolox equivalent antioxidant capacity of unhatched egg for ABTS, DPPH, and that of hydroxyl radicals were found to be 199, 993, and 226.5 μmol/L, respectively. CEEH showed stronger scavenging activity than the hydrolysates of unhatched egg against free radicals such as ABTS, DPPH, and hydroxyl radicals. The antioxidant amino acid analysis indicated that the 14-day CEEH possess more antioxidant amino acids than that of the unhatched egg. In addition, essential amino acids analysis showed that the 14-day CEEH have the highest nutritional value. Combined with the results of the amino acid profiles, CEEH were believed to have higher nutritive value in addition to antioxidant activities than the unhatched egg. PMID:24804065

  12. Microwave effects on isolated chick embryo hearts

    SciT

    Caddemi, A.; Tamburello, C.C.; Zanforlin, L.

    1986-01-01

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

  13. Excess Imidacloprid Exposure Causes the Heart Tube Malformation of Chick Embryos.

    PubMed

    Gao, Lin-Rui; Li, Shuai; Zhang, Jing; Liang, Chang; Chen, En-Ni; Zhang, Shi-Yao; Chuai, Manli; Bao, Yong-Ping; Wang, Guang; Yang, Xuesong

    2016-11-30

    As a neonicotinoid pesticide, imidacloprid is widely used to control sucking insects on agricultural planting and fleas on domestic animals. However, the extent to which imidacloprid exposure has an influence on cardiogensis in early embryogenesis is still poorly understood. In vertebrates, the heart is the first organ to be formed. In this study, to address whether imidacloprid exposure affects early heart development, the early chick embryo has been used as an experimental model because of its accessibility at its early developmental stage. The results demonstrate that exposure of the early chick embryo to imidacloprid caused malformation of heart tube. Furthermore, the data reveal that down-regulation of GATA4, NKX2.5, and BMP4 and up-regulation of Wnt3a led to aberrant cardiomyocyte differentiation. In addition, imidacloprid exposure interfered with basement membrane breakdown, E-cadherin/laminin expression, and mesoderm formation during the epithelial-mesenchymal transition (EMT) in gastrula chick embryos. Finally, the DiI-labeled cell migration trajectory indicated that imidacloprid restricted the cell migration of cardiac progenitors to primary heart field in gastrula chick embryos. A similar observation was also obtained from the cell migration assay of scratch wounds in vitro. Additionally, imidacloprid exposure negatively affected the cytoskeleton structure and expression of corresponding adhesion molecules. Taken together, these results reveal that the improper EMT, cardiac progenitor migration, and differentiation are responsible for imidacloprid exposure-induced malformation of heart tube during chick embryo development.

  14. Use of chick embryo in screening for teratogenicity.

    PubMed

    Kotwani, A

    1998-04-01

    A teratology screening system would detect agents hazardous to the conceptus before they can perturb embryonic development in humans. The back log of untested chemicals and the rate at which new substances enter the market exceed the developmental effects testing by standard in vivo method. Thus, cheaper, quicker in vitro systems afford a unique opportunity for investigating the direct interaction of substances with developing morphogenetic system (MGSs), since maternal influences are excluded. As a carrier of a complete set of MGSs, the chick embryo in ovo manifests an advantage over those in vitro systems that employ isolated embryos or embryonic tissues that have only limited survival. Under controlled experimental conditions including standardization of subjects, administration technique and mode of evaluation, according to the basic principles of teratology, the chick embryo test is demonstrated to be reliable and to afford quantifiable end points for evaluation. Individual compounds, mixtures of compounds and against and antagonist can easily be administered and tested. The chick embryo possesses its own basic enzyme-catalyzed drug-transformation capacity and moreover, it can be used for screening specific human metabolites. Different newer techniques e.g. chick embryotoxicity screening test (CHEST), Chick embryo blastoderm model etc are described in detail. Chick embryo fulfills all the criteria which a test should have at a lower level of tier system in teratological studies i.e. modest laboratory equipment, moderate skill, minimal expenditure of time and money, ease of accessibility of embryo, known embryological development, possibility of experimenting on a large scale for statistically valid results and whole animals are also not required.

  15. Ex Ovo Model for Directly Visualizing Chick Embryo Development

    ERIC Educational Resources Information Center

    Dorrell, Michael I.; Marcacci, Michael; Bravo, Stephen; Kurz, Troy; Tremblay, Jacob; Rusing, Jack C.

    2012-01-01

    We describe a technique for removing and growing chick embryos in culture that utilizes relatively inexpensive materials and requires little space. It can be readily performed in class by university, high school, or junior high students, and teachers of any grade level should be able to set it up for their students. Students will be able to…

  16. Effect of coniine on the developing chick embryo.

    PubMed

    Forsyth, C S; Frank, A A; Watrous, B J; Bohn, A A

    1994-04-01

    Coniine, an alkaloid from Conium maculatum (poison hemlock), has been shown to be teratogenic in livestock. The major teratogenic outcome is arthrogryposis, presumably due to nicotinic receptor blockade. However, coniine has failed to produce arthrogryposis in rats or mice and is only weakly teratogenic in rabbits. The purpose of this study was to evaluate and compare the effects of coniine and nicotine in the developing chick. Concentrations of coniine and nicotine sulfate were 0.015%, 0.03%, 0.075%, 0.15%, 0.75%, 1.5%, 3%, and 6% and 1%, 5%, and 10%, respectively. Both compounds caused deformations and lethality in a dose-dependent manner. All concentrations of nicotine sulfate caused some lethality but a no effect level for coniine lethality was 0.75%. The deformations caused by both coniine and nicotine sulfate were excessive flexion or extension of one or more toes. No histopathological alterations or differences in bone formation were seen in the limbs or toes of any chicks from any group; however, extensive cranial hemorrhage occurred in all nicotine sulfate-treated chicks. There was a statistically significant (P < or = 0.01) decrease in movement in coniine and nicotine sulfate treated chicks as determined by ultrasound. Control chicks were in motion an average of 33.67% of the time, while coniine-treated chicks were only moving 8.95% of a 5-min interval, and no movement was observed for nicotine sulfate treated chicks. In summary, the chick embryo provides a reliable and simple experimental animal model of coniine-induced arthrogryposis. Data from this model support a mechanism involving nicotinic receptor blockade with subsequent decreased fetal movement.

  17. Development of respiratory rhythms in perinatal chick embryos.

    PubMed

    Chiba, Y; Khandoker, A H; Nobuta, M; Moriya, K; Akiyama, R; Tazawa, H

    2002-04-01

    In chick embryos, gas exchange takes place via the chorioallantoic membrane (CAM) and the lungs at approximately 1 day prior to hatching. The present study was designed to elucidate the development of respiratory rhythms in the chick embryo during the whole pipping (perinatal) period with a condenser-microphone measuring system. The microphone was hermetically attached on the eggshell over the air cell on day 18 of incubation. It first detected a cardiogenic signal (i.e. acoustocardiogram), and then beak clapping and breathing signals (acoustorespirogram, ARG). The first signals of lung ventilation appeared intermittently and irregularly approximately once per 5 s among the clapping signals after the embryo penetrated its beak into the air cell (internal pipping, IP). The respiratory rhythm then developed irregularly, with a subsequent more regular rate. The envelope pattern of breathing from the onset of IP through external pipping (EP) to hatching was constructed by a specially devised procedure, which eliminated external and internal noises. The envelope patterns indicated that the IP, EP and whole perinatal periods of 10 embryos were 14.1+/-6.4 (S.D.), 13.6+/-4.0 and 27.6+/-5.4 h, respectively. In addition, they also indicated the period of embryonic hatching activity (i.e. climax) which was 48+/-19 min. The development of respiratory rhythm was also shown by the instantaneous respiratory rate (IRR) which was designated as an inverse value of two adjacent ARG waves.

  18. Bilateral neuro-retinitis following chick embryo cell anti-rabies vaccination – a case report

    PubMed Central

    Saxena, Rohit; Sethi, Harinder Singh; Rai, Harminder Kumar; Menon, Vimla

    2005-01-01

    Background The Optic nerve is rarely involved after sheep brain anti-rabies vaccination in the form of retrobulbar neuritis or papillitis. Bilateral neuroretinitis after chick embryo cell antirabies vaccination has not been reported. Case presentation We report the case of a 56 year old male who developed bilateral neuro-retinitis following three injections of antirabies vaccine prepared from the chick embryo. Conclusion The chick embryo cell antirabies vaccine can cause bilateral neuroretinits which has not been reported previously. PMID:16105182

  19. Effects of choline on sodium arsenite-induced neural tube defects in chick embryos.

    PubMed

    Song, Ge; Cui, Yi; Han, Zhong-Ji; Xia, Hong-Fei; Ma, Xu

    2012-12-01

    Arsenic passes through the placenta and accumulates in the neuroepithelium of embryo, whereby inducing congenital malformations such as neural tube defects (NTDs) in animals. Choline (CHO), a methyl-rich nutrient, functions as a methyl donor to participate in methyl group metabolism. Arsenic methylation has been regarded as a detoxification process and choline (CHO) is the major source of methyl-groups. However, whether CHO intake reverses the abnormal embryo development induced by sodium arsenite (SA) and the relationship between CHO intake and arsenite-induced NTDs are still unclear. In this study, we used chick embryos as animal model to investigate the effects of SA and CHO supplementation on the early development of nervous system. Our results showed that the administration of SA led to reduction in embryo viability, embryo body weight and extraembryonic vascular area, accompanied by a significantly increased incidence of the failed closure of the caudal end of the neural tube. CHO, at low dose (25 μg/μL), reversed the decrease in embryo viability and the increase in the failed closure of the caudal end of the neural tube, which were induced by SA. In addition, CHO (25 μg/μL) inhibited not only the SA-induced cell apoptosis by up-regulating Bcl-2 level, but also the global DNA methylation by increasing the expressions of DNMT1 and DNMT3a. However, less significant difference was found between the embryos co-treated with SA and CHO (50 μg/μL) and the ones treated with SA alone. Taken together, these findings suggest that low dose CHO could protect chick embryos from arsenite-induced NTDs by a possible mechanism related to the methyl metabolism. Copyright © 2012 Elsevier Ltd. All rights reserved.

  20. Sulphonated phthalocyanine induced caudal malformative syndrome in the chick embryo.

    PubMed

    Sandor, S; Prelipceanu, O; Checiu, I

    1985-01-01

    Sulphonated phthalocyanine (Pht.) has been tested for its possible noxious effect on the developing chick embryo. When injected into the subembryonic cavity of 40-45 hours incubated chick embryos (mainly 10-20 somite pairs), Pht. induces a highly reproducible caudal malformative syndrome (trunk and taillessness, various anomalies of the limbs). The main effect is--in about 15% of the malformed specimens--associated with unilateral microphthalmy and, less frequently, with coelosomy. Microscopically developmental disturbances of the caudal axial organs, of the mesonephros and of the limbs are observed. The initial pathological changes, at microscopic level, are necrosis and hemorrhages in the caudal axial and paraxial area. The allantois is poorly developed or even absent. Skeletal changes involve anomalies of the ribs and of the vertebral column and total or partial absence of the pelvic girdle bones. The high mortality, mainly during the first week, is due--first of all--to the developmental disturbances including the poor development or absence of the allantois. Control experiments with CuCl2 suggest the ethiological role of Cu. Pathogenetic aspects are discussed.

  1. The generation of vertebral segmental patterning in the chick embryo

    PubMed Central

    Senthinathan, Biruntha; Sousa, Cátia; Tannahill, David; Keynes, Roger

    2012-01-01

    We have carried out a series of experimental manipulations in the chick embryo to assess whether the notochord, neural tube and spinal nerves influence segmental patterning of the vertebral column. Using Pax1 expression in the somite-derived sclerotomes as a marker for segmentation of the developing intervertebral disc, our results exclude such an influence. In contrast to certain teleost species, where the notochord has been shown to generate segmentation of the vertebral bodies (chordacentra), these experiments indicate that segmental patterning of the avian vertebral column arises autonomously in the somite mesoderm. We suggest that in amniotes, the subdivision of each sclerotome into non-miscible anterior and posterior halves plays a critical role in establishing vertebral segmentation, and in maintaining left/right alignment of the developing vertebral elements at the body midline. PMID:22458512

  2. The chick embryo: a leading model in somitogenesis studies.

    PubMed

    Pourquié, Olivier

    2004-09-01

    The vertebrate body is built on a metameric organization which consists of a repetition of functionally equivalent units, each comprising a vertebra, its associated muscles, peripheral nerves and blood vessels. This periodic pattern is established during embryogenesis by the somitogenesis process. Somites are generated in a rhythmic fashion from the presomitic mesoderm and they subsequently differentiate to give rise to the vertebrae and skeletal muscles of the body. Somitogenesis has been very actively studied in the chick embryo since the 19th century and many of the landmark experiments that led to our current understanding of the vertebrate segmentation process have been performed in this organism. Somite formation involves an oscillator, the segmentation clock whose periodic signal is converted into the periodic array of somite boundaries by a spacing mechanism relying on a traveling threshold of FGF signaling regressing in concert with body axis extension.

  3. The generation of vertebral segmental patterning in the chick embryo.

    PubMed

    Senthinathan, Biruntha; Sousa, Cátia; Tannahill, David; Keynes, Roger

    2012-06-01

    We have carried out a series of experimental manipulations in the chick embryo to assess whether the notochord, neural tube and spinal nerves influence segmental patterning of the vertebral column. Using Pax1 expression in the somite-derived sclerotomes as a marker for segmentation of the developing intervertebral disc, our results exclude such an influence. In contrast to certain teleost species, where the notochord has been shown to generate segmentation of the vertebral bodies (chordacentra), these experiments indicate that segmental patterning of the avian vertebral column arises autonomously in the somite mesoderm. We suggest that in amniotes, the subdivision of each sclerotome into non-miscible anterior and posterior halves plays a critical role in establishing vertebral segmentation, and in maintaining left/right alignment of the developing vertebral elements at the body midline. © 2012 The Authors. Journal of Anatomy © 2012 Anatomical Society.

  4. Morphogenesis and calcification of the statoconia in the chick (Gallus domesticus) embryo - Implications for future studies

    NASA Technical Reports Server (NTRS)

    Fermin, C. D.; Igarashi, M.

    1985-01-01

    The morphogenesis of the statoconia in the chick, Gallus domesticus, injected with a carbon anhydrase inhibitor is studied. The preparation of the embryo specimens for analysis is described. The early, middle, and late stages of embryonic development are examined. The data reveal that acetozolamide inhibits statoconia formation in the middle stage of development and the calcification process follows statoconia formation. The spatial relationship between the development of type 1 and type 2 hair cells and the appearance and maturation of the statoconia is investigated.

  5. Embryonic toxico-pathological effects of meglumine antimoniate using a chick embryo model.

    PubMed

    Khosravi, Ahmad; Sharifi, Iraj; Tavakkoli, Hadi; Derakhshanfar, Amin; Keyhani, Ali Reza; Salari, Zohreh; Mosallanejad, Seyedeh Saedeh; Bamorovat, Mehdi

    2018-01-01

    Leishmaniasis is one of the diverse and neglected tropical diseases. Embryo-toxicity of drugs has always been a major concern. Chick embryo is a preclinical model relevant in the assessment of adverse effects of drugs. The current study aimed to assess embryonic histopathological disorders and amniotic fluid biochemical changes following meglumine antimoniate treatment. The alteration of vascular branching pattern in the chick's extra-embryonic membrane and exploration of molecular cues to early embryonic vasculogenesis and angiogenesis were also quantified. Embryonated chicken eggs were treated with 75 or 150 mg/kg of meglumine antimoniate. Embryo malformations, growth retardation and haemorrhages on the external body surfaces were accompanied by histopathological lesions in the brain, kidney, liver and heart in a dose-dependent manner. Significant rise occurred in the biochemical indices of alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase and amylase in the amniotic fluid. Quantification of the extra-embryonic membrane vasculature showed that the anti-angiogenic and anti-vasculogenic effects of the drug were revealed by a significant decrease in fractal dimension value and mean capillary area. The relative expression levels of vascular endothelial growth factor A and vascular endothelial growth factor receptor 2 mRNA also significantly reduced. Concerns of a probable teratogenicity of meglumine antimoniate were established by data presented in this study. It is concluded that tissue lesions, amniotic fluid disturbance, altered early extra-embryonic vascular development and gene expression as well as the consecutive cascade of events, might eventually lead to developmental defects in embryo following meglumine antimoniate treatment. Therefore, the use of meglumine antimoniate during pregnancy should be considered as potentially embryo-toxic. Hence, physicians should be aware of such teratogenic effects and limit the use of this drug

  6. Patterning in time and space: HoxB cluster gene expression in the developing chick embryo.

    PubMed

    Gouveia, Analuce; Marcelino, Hugo M; Gonçalves, Lisa; Palmeirim, Isabel; Andrade, Raquel P

    2015-01-01

    The developing embryo is a paradigmatic model to study molecular mechanisms of time control in Biology. Hox genes are key players in the specification of tissue identity during embryo development and their expression is under strict temporal regulation. However, the molecular mechanisms underlying timely Hox activation in the early embryo remain unknown. This is hindered by the lack of a rigorous temporal framework of sequential Hox expression within a single cluster. Herein, a thorough characterization of HoxB cluster gene expression was performed over time and space in the early chick embryo. Clear temporal collinearity of HoxB cluster gene expression activation was observed. Spatial collinearity of HoxB expression was evidenced in different stages of development and in multiple tissues. Using embryo explant cultures we showed that HoxB2 is cyclically expressed in the rostral presomitic mesoderm with the same periodicity as somite formation, suggesting a link between timely tissue specification and somite formation. We foresee that the molecular framework herein provided will facilitate experimental approaches aimed at identifying the regulatory mechanisms underlying Hox expression in Time and Space.

  7. Patterning in time and space: HoxB cluster gene expression in the developing chick embryo

    PubMed Central

    Gouveia, Analuce; Marcelino, Hugo M; Gonçalves, Lisa; Palmeirim, Isabel; Andrade, Raquel P

    2015-01-01

    The developing embryo is a paradigmatic model to study molecular mechanisms of time control in Biology. Hox genes are key players in the specification of tissue identity during embryo development and their expression is under strict temporal regulation. However, the molecular mechanisms underlying timely Hox activation in the early embryo remain unknown. This is hindered by the lack of a rigorous temporal framework of sequential Hox expression within a single cluster. Herein, a thorough characterization of HoxB cluster gene expression was performed over time and space in the early chick embryo. Clear temporal collinearity of HoxB cluster gene expression activation was observed. Spatial collinearity of HoxB expression was evidenced in different stages of development and in multiple tissues. Using embryo explant cultures we showed that HoxB2 is cyclically expressed in the rostral presomitic mesoderm with the same periodicity as somite formation, suggesting a link between timely tissue specification and somite formation. We foresee that the molecular framework herein provided will facilitate experimental approaches aimed at identifying the regulatory mechanisms underlying Hox expression in Time and Space. PMID:25602523

  8. Respiratory gas exchange of high altitude adapted chick embryos

    NASA Technical Reports Server (NTRS)

    Wangensteen, O. D.; Rahn, H.; Burton, R. R.; Smith, A. H.

    1974-01-01

    Study of gas exchange by embryos from chickens acclimatized to an altitude of 3800 m. The oxygen partial pressure and carbon dioxide partial pressure differences across the egg shell were measured and found to be less than the values previously reported for sea-level eggs by about a factor of two. Further measurements of embryonic oxygen consumption and shell conductivity to oxygen indicated that, compared to eggs at sea level, oxygen consumption was reduced by a factor of 0.58 while conductivity to oxygen was increased only by a factor of 1.07 in the high-altitude eggs. These independent measurements predict the change in oxygen partial pressure across the egg shell of the high-altitude eggs to be only 0.54 times that of sea-level eggs; the directly measured factor was 0.53. The authors conclude that at high altitude, a major adaptation of the chick embryo is a reduced metabolism which decreases the change in oxygen partial pressure across the egg shell since its gas conductivity remains essentially unchanged.

  9. Development of a green fluorescent protein metastatic-cancer chick-embryo drug-screen model.

    PubMed

    Bobek, Vladimir; Plachy, Jiri; Pinterova, Daniela; Kolostova, Katarina; Boubelik, Michael; Jiang, Ping; Yang, Meng; Hoffman, Robert M

    2004-01-01

    The chick-embryo model has been an important tool to study tumor growth, metastasis, and angiogenesis. However, an imageable model with a genetic fluorescent tag in the growing and spreading cancer cells that is stable over time has not been developed. We report here the development of such an imageable fluorescent chick-embryo metastatic cancer model with the use of green fluorescent protein (GFP). Lewis lung carcinoma cells, stably expressing GFP, were injected on the 12th day of incubation in the chick embryo. GFP-Lewis lung carcinoma metastases were visualized by fluorescence, after seven days additional incubation, in the brain, heart, and sternum of the developing chick embryo, with the most frequent site being the brain. The combination of streptokinase and gemcitabine was evaluated in this GFP metastatic model. Twelve-day-old chick embryos were injected intravenously with GFP-Lewis lung cancer cells, along with these two agents either alone or in combination. The streptokinase-gemcitabine combination inhibited metastases at all sites. The effective dose of gemcitabine was found to be 10 mg/kg and streptokinase 2000 IU per embryo. The data in this report suggest that this new stably fluorescent imageable metastatic-cancer chick-embryo model will enable rapid screening of new antimetastatic agents.

  10. Effects of antibodies to EG-VEGF on angiogenesis in the chick embryo chorioallantoic membrane.

    PubMed

    Feflea, Stefana; Cimpean, Anca Maria; Ceausu, Raluca Amalia; Gaje, Pusa; Raica, Marius

    2012-01-01

    Endocrine gland-related vascular endothelial growth factor (EG-VEGF), is an angiogenic factor specifically targeting endothelial cells derived from endocrine tissues. The inhibition of the EG-VEGF/prokineticin receptor pathway could represent a selective antiangiogenic and anticancer strategy. to evaluate the impact of an antibody to EG-VEGF on the rapidly growing capillary plexus of the chick embryo chorioallantoic membrane (CAM). The in ovo CAM assay was performed for the humanized EG-VEGF antibody. Hemorrhagic damage was induced in the capillaries, which led to early death of the embryos. Upon morphological staining, there was evidence of vascular disruption and extravasation of red blood cells in the chorion. Signs of vacuolization of the covering epithelium were also observed. Blocking endogenous EG-VEGF might represent a valuable approach of impairing or inhibiting angiogenesis in steroidogenic-derived embryonic tissues.

  11. Transformation of primary chick embryo fibroblasts by Marek's disease virus.

    PubMed

    Buranathai, C; Rodriguez, J; Grose, C

    1997-12-08

    Marek's disease virus (MDV) is an alphaherpesvirus, which can mediate the malignant transformation of lymphocytes to form lymphomas in chickens. In this study, we demonstrate that MDV can transform primary chick embryo fibroblasts (CEF). The cell line derived from primary CEF infected with the GA strain of MDV was called CEM(MDV). The fibroblast nature of CEM(MDV) was verified by absence of cytokeratin type II. The CEM(MDV) phenotype differed from either primary CEF or MDV-infected CEF. CEM(MDV) were extensively vacuolated, with unusual multilamellar structures in the cytoplasm, The nuclei were considerably larger than those in primary CEF and were uniformly positive for proliferating cell nuclear antigen. The cell line was subcultured for more than 10 generations; however, CEM(MDV) did not support a fully productive MDV infection, because complete nucleocapsids were not detected and infectivity assays showed that cell line produced no infectious virus. PCR analyses demonstrated that this cell line carried both polypeptide 38 (pp38) and Meq DNA, MDV-specific genes associated with transformation. In addition, examination by laser scanning confocal microscopy revealed that CEM(MDV) constitutively produced MDV MEQ protein in nuclei and pp38 as well as glycoprotein B in the cytoplasm and on the plasma membrane. Growth in soft agar assay demonstrated that CEM(MDV) formed colonies, similar to HeLa and human melanoma cells. Retroviral insertion was not detected in DNA from the CEM(MDV) line.

  12. Measuring the electric activity of chick embryos heart through 16 bit audio card monitored by the Goldwavetm software

    NASA Astrophysics Data System (ADS)

    Silva, Dilson; Cortez, Celia Martins

    2015-12-01

    In the present work we used a high-resolution, low-cost apparatus capable of detecting waves fit inside the sound bandwidth, and the software package GoldwaveTM for graphical display, processing and monitoring the signals, to study aspects of the electric heart activity of early avian embryos, specifically at the 18th Hamburger & Hamilton stage of the embryo development. The species used was the domestic chick (Gallus gallus), and we carried out 23 experiments in which cardiographic spectra of QRS complex waves representing the propagation of depolarization waves through ventricles was recorded using microprobes and reference electrodes directly on the embryos. The results show that technique using 16 bit audio card monitored by the GoldwaveTM software was efficient to study signal aspects of heart electric activity of early avian embryos.

  13. Ethical euthanasia and short-term anesthesia of the chick embryo.

    PubMed

    Aleksandrowicz, Ewa; Herr, Ingrid

    2015-01-01

    Fertilized chicken eggs are suggested as an alternative to mammalian models. The chorioallantoic membrane (CAM) of the chick embryo is widely used for examination of angiogenesis, xenotransplants and for virus production. Unfortunately, it is mostly not taken into account, that the chick embryo's ability to experience pain starts to develop at day 7 of breeding. In our view, this model is only in accordance with the 3 R principles, if an appropriate anesthesia of the chick embryo in potentially painful procedures is provided. Although many experimental approaches are performed on the none-innervated CAM, the euthanasia of the embryo strongly requires a more human technique than the usually used freezing at -20°C, decapitation or in ovo fixation with paraformaldehyde without prior anesthesia. However, protocols regarding feasible and ethical methods for anesthesia and euthanasia of avian embryos are currently not available. Therefore, we established an easy and reliable method for the euthanasia and short-term anesthesia of the chick embryo.

  14. Atrial and ventricular septal changes in ethanol vapour exposed chick embryos.

    PubMed

    Kamran, Kiran; Khan, Muhammad Yunus; Minhas, Liaqat Ali

    2015-03-01

    To study the effects of ethanol vapour exposure on development of atrial and ventricular septa of chick embryo. The experimental study was conducted at the College of Physicians and Surgeons, Islamabad, from 2006 to 2007. The experimental and control groups were further divided into three subgroups based on the day of sacrifice. The experimental group was exposed to ethanol vapours produced in a specially-designed vapour chamber and then compared with age-matched controls. There were 90 eggs in each of the two groups. The development of inter-ventricular septum completed at day 7 of development in chick embryo. Ethanol vapour exposure produced a small discontinuity at day 10 of development in a chick embryo which may be labelled as ventricular septal defect since ventricular development is completed by day 7. Interatrial septum formed till day 7 with small perforations which persisted till hatching. Ethanol vapour exposure may lead to ventricular septal defect.

  15. Development of teeth in chick embryos after mouse neural crest transplantations.

    PubMed

    Mitsiadis, Thimios A; Chéraud, Yvonnick; Sharpe, Paul; Fontaine-Pérus, Josiane

    2003-05-27

    Teeth were lost in birds 70-80 million years ago. Current thinking holds that it is the avian cranial neural crest-derived mesenchyme that has lost odontogenic capacity, whereas the oral epithelium retains the signaling properties required to induce odontogenesis. To investigate the odontogenic capacity of ectomesenchyme, we have used neural tube transplantations from mice to chick embryos to replace the chick neural crest cell populations with mouse neural crest cells. The mouse/chick chimeras obtained show evidence of tooth formation showing that avian oral epithelium is able to induce a nonavian developmental program in mouse neural crest-derived mesenchymal cells.

  16. Atg7-Mediated Autophagy Is Involved in the Neural Crest Cell Generation in Chick Embryo.

    PubMed

    Wang, Guang; Chen, En-Ni; Liang, Chang; Liang, Jianxin; Gao, Lin-Rui; Chuai, Manli; Münsterberg, Andrea; Bao, Yongping; Cao, Liu; Yang, Xuesong

    2018-04-01

    Autophagy plays a very important role in numerous physiological and pathological events. However, it still remains unclear whether Atg7-induced autophagy is involved in the regulation of neural crest cell production. In this study, we found the co-location of Atg7 and Pax7 + neural crest cells in early chick embryo development. Upregulation of Atg7 with unilateral transfection of full-length Atg7 increased Pax7 + and HNK-1 + cephalic and trunk neural crest cell numbers compared to either Control-GFP transfection or opposite neural tubes, suggesting that Atg7 over-expression in neural tubes could enhance the production of neural crest cells. BMP4 in situ hybridization and p-Smad1/5/8 immunofluorescent staining demonstrated that upregulation of Atg7 in neural tubes suppressed the BMP4/Smad signaling, which is considered to promote the delamination of neural crest cells. Interestingly, upregulation of Atg7 in neural tubes could significantly accelerate cell progression into the S phase, implying that Atg7 modulates cell cycle progression. However, β-catenin expression was not significantly altered. Finally, we demonstrated that upregulation of the Atg7 gene could activate autophagy as did Atg8. We have also observed that similar phenotypes, such as more HNK-1 + neural crest cells in the unilateral Atg8 transfection side of neural tubes, and the transfection with full-length Atg8-GFP certainly promote the numbers of BrdU + neural crest cells in comparison to the GFP control. Taken together, we reveal that Atg7-induced autophagy is involved in regulating the production of neural crest cells in early chick embryos through the modification of the cell cycle.

  17. Eye and heart morphogenesis are dependent on melatonin signaling in chick embryos.

    PubMed

    Nogueira, Renato C; Sampaio, Lucia de Fatima S

    2017-10-15

    Calmodulin is vital for chick embryos morphogenesis in the incubation time 48-66 h when the rudimentary C-shaped heart attains an S-shaped pattern and the optic vesicles develop into optic cups. Melatonin is in the extraembryonic yolk sac of the avian egg; melatonin binds calmodulin. The aim of this study was to investigate the function of melatonin in the formation of the chick embryo optic cups and S-shaped heart, by pharmacological methods and immunoassays. Mel1a melatonin receptor immunofluorescence was distributed in the optic cups and rudimentary hearts. We separated embryonated chicken eggs at 48 h of incubation into basal, control and drug-treated groups, with treatment applied in the egg air sac. At 66 h of incubation, embryos were excised from the eggs and analyzed. Embryos from the basal, control (distilled water), melatonin and 6-chloromelatonin (melatonin receptor agonist) groups had regular optic cups and an S-shaped heart, while those from the calmidazolium (calmodulin inhibitor) group did not. Embryos from the luzindole (melatonin receptor antagonist) and prazosin (Mel1c melatonin receptor antagonist) groups did not have regular optic cups. Embryos from the 4-P-PDOT (Mel1b melatonin receptor antagonist) group did not have an S-shaped heart. Previous application of the melatonin, 6-chloromelatonin or forskolin (adenylate cyclase enhancer) prevented the abnormal appearance of chick embryos from the calmidazolium, luzindole, prazosin and 4-P-PDOT groups. However, 6-chloromelatonin and forskolin only partially prevented the development of defective eye cups in embryos from the calmidazolium group. The results suggested that melatonin modulates chick embryo morphogenesis via calmodulin and membrane receptors. © 2017. Published by The Company of Biologists Ltd.

  18. High glucose environment inhibits cranial neural crest survival by activating excessive autophagy in the chick embryo

    PubMed Central

    Wang, Xiao-Yu; Li, Shuai; Wang, Guang; Ma, Zheng-Lai; Chuai, Manli; Cao, Liu; Yang, Xuesong

    2015-01-01

    High glucose levels induced by maternal diabetes could lead to defects in neural crest development during embryogenesis, but the cellular mechanism is still not understood. In this study, we observed a defect in chick cranial skeleton, especially parietal bone development in the presence of high glucose levels, which is derived from cranial neural crest cells (CNCC). In early chick embryo, we found that inducing high glucose levels could inhibit the development of CNCC, however, cell proliferation was not significantly involved. Nevertheless, apoptotic CNCC increased in the presence of high levels of glucose. In addition, the expression of apoptosis and autophagy relevant genes were elevated by high glucose treatment. Next, the application of beads soaked in either an autophagy stimulator (Tunicamycin) or inhibitor (Hydroxychloroquine) functionally proved that autophagy was involved in regulating the production of CNCC in the presence of high glucose levels. Our observations suggest that the ERK pathway, rather than the mTOR pathway, most likely participates in mediating the autophagy induced by high glucose. Taken together, our observations indicated that exposure to high levels of glucose could inhibit the survival of CNCC by affecting cell apoptosis, which might result from the dysregulation of the autophagic process. PMID:26671447

  19. 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.

  20. Neurologic illness following post-exposure prophylaxis with purifiled chick embryo cell antirabies vaccine.

    PubMed

    Chakravarty, A

    2001-09-01

    Clinical details of a neurologic illness simultating Guillain Barre syndrome developing in a patient after post-exposure prophylaxis with purified chick embryo cell (PCEC) anti-rabies vaccine is reported. Neurologic complication following PCEC vaccination has not been reported earlier.

  1. Impaired cytoskeletal arrangements and failure of ventral body wall closure in chick embryos treated with rock inhibitor (Y-27632).

    PubMed

    Duess, Johannes W; Puri, Prem; Thompson, Jennifer

    2016-01-01

    Rho-associated kinase (ROCK) signaling regulates numerous fundamental developmental processes during embryogenesis, primarily by controlling actin-cytoskeleton assembly and cell contractility. ROCK knockout mice exhibit a ventral body wall defect (VBWD) phenotype due to disorganization of actin filaments at the umbilical ring. However, the exact molecular mechanisms leading to VBWD still remain unclear. Improper somitogenesis has been hypothesized to contribute to failure of VBW closure. We designed this study to investigate the hypothesis that administration of ROCK inhibitor (Y-27632) disrupts cytoskeletal arrangements in morphology during early chick embryogenesis, which may contribute to the development of VBWD. At 60 h incubation, chick embryos were explanted into shell-less culture and treated with 50 µL of vehicle for controls (n = 33) or 50 µL of 500 µM of Y-27632 for the experimental group (Y-27, n = 56). At 8 h post-treatment, RT-PCR was performed to evaluate mRNA levels of N-cadherin, E-cadherin and connexin43. Immunofluorescence confocal microscopy was performed to analyze the expression and distribution of actin, vinculin and microtubules in the neural tube and somites. A further cohort of embryos was treated in ovo by dropping 50 µL of vehicle or 50 µL of different concentrations of Y-27632 onto the embryo and allowing development to 12 and 14 days for further assessment. Gene expression levels of N-cadherin, E-cadherin and connexin43 were significantly decreased in treated embryos compared with controls (p < 0.05). Thickened actin filament bundles were recorded in the neural tube of Y-27 embryos. In somites, cells were dissociated with reduced actin distribution in affected embryos. Clumping of vinculin expression was found in the neural tube and somites, whereas reduced expression of microtubules was observed in Y-27 embryos compared with controls. At 12 and 14 days of development, affected embryos presented with an enlarged

  2. Mechanical Effects of the Surface Ectoderm on Optic Vesicle Morphogenesis in the Chick Embryo

    PubMed Central

    Hosseini, Hadi S.; Beebe, David C.; Taber, Larry A.

    2014-01-01

    Precise shaping of the eye is crucial for proper vision. Here, we use experiments on chick embryos along with computational models to examine the mechanical factors involved in the formation of the optic vesicles (OVs), which grow outward from the forebrain of the early embryo. First, mechanical dissections were used to remove the surface ectoderm (SE), a membrane that contacts the outer surfaces of the OVs. Principal components analysis of OV shapes suggests that the SE exerts asymmetric loads that cause the OVs to flatten and shear caudally during the earliest stages of eye development and later to bend in the caudal and dorsal directions. These deformations cause the initially spherical OVs to become pear-shaped. Exposure to the myosin II inhibitor blebbistatin reduced these effects, suggesting that cytoskeletal contraction controls OV shape by regulating tension in the SE. To test the physical plausibility of these interpretations, we developed 2-D finite-element models for frontal and transverse cross-sections of the forebrain, including frictionless contact between the SE and OVs. With geometric data used to specify differential growth in the OVs, these models were used to simulate each experiment (control, SE removed, no contraction). For each case, the predicted shape of the OV agrees reasonably well with experiments. The results of this study indicate that differential growth in the OV and external pressure exerted by the SE are suffcient to cause the global changes in OV shape observed during the earliest stages of eye development. PMID:25458577

  3. A homeobox gene involved in node, notochord and neural plate formation of chick embryos.

    PubMed

    Stein, S; Kessel, M

    1995-01-01

    We have isolated a chicken cDNA clone, Cnot, resembling in sequence and expression pattern the Xenopus homeobox gene Xnot. The major, early transcription domains of Cnot are the node, the notochord and prenodal and postnodal neural plate caudal from the prospective hindbrain level. All these cell populations appear to be descendants of the Cnot-expressing cells of the node, suggesting a cell lineage relationship. After the onset of somitogenesis, a second, independent expression domain appears in the neural folds at the prospective mid- and forebrain levels, and further transcripts are found in the epiphysis, the ventral diencephalon, the preoral gut and the limb buds. Transplantation of nodes from extended streak embryos leads to the formation of ectopic notochords, which express Cnot in the typical, cranially decreasing gradient. Transplantation of young nodes to young hosts has previously been described to induce secondary embryos. We observed that secondary chick embryos express Cnot in node derived, notochord-like structures and in the anterior neural plate, similar to the domains seen in primary embryos. However, expression was absent from the posterior neural plate, which in the induction experiments is excluded from the node lineage. This finding corroborates our initial conclusion about a cell lineage relationship between node, notochord, and neural plate defined by Cnot expression. The midline mesoderm of vertebrate embryos consists of two tissues, the prechordal mesoderm and the notochord. The anterior notochord, the head process, may represent an intermediate form. The transition from prechordal to chordal mesoderm can be followed by the expression of the two marker homeobox genes goosecoid and Cnot, first in the primitive streak, and then in the head process. We suggest that expression of goosecoid or Cnot is involved in the specification of a prechordal or notochordal identity, respectively. A transition from goosecoid to Cnot expression may proceed

  4. AUTORADIOGRAPHIC STUDIES OF THE UTILIZATION OF S35-SULFATE BY THE CHICK EMBRYO

    PubMed Central

    Johnston, Perry M.; Comar, Cyril L.

    1957-01-01

    From studies of autoradiograms of various developmental stages of the chick embryo containing S35 given us sulfate it was determined that as early as Stages 3+ and 4 there is a selective utilization or accumulation of sulfate by the various parts. The earliest accumulation site is the axial portion of the primitive streak and the floor of the groove. Later S35 was found in the head process, Hensen's node, notochord, amniocardiac vesicle, wall of the omphalomesenteric vein, endocardium, subendocardial jelly, mesenchyme destined to become cartilage, basement membrane area of the gut, and a mucopolysaccharide layer formed on the free surface of the stomach. The early notochordal localizations of S35 coincide with the region in which a thin ring of chondroitin sulfate is subsequently laid down. However, it is apparent that there is an intracellular accumulation of inorganic sulfate by the chondroitin-forming cells prior to the time they produce sufficient chondroitin sulfate to be demonstrable histochemically. It was interesting to note that the endocardium appears to concentrate sulfate that later apparently finds its way into the subendocardial jelly. The fact that those mesenchymal cells which later form chondroblasts begin to utilize sulfate selectively before histological differentiation is apparent was determined. In addition, the presence of sulfate-containing substances in the forming basement membrane of the gut would seem to indicate that sulfate is important in the histological differentiation of this membrane. PMID:13438906

  5. THE SUSCEPTIBILITY OF CHICK EMBRYO SKIN ORGAN CULTURES TO INFLUENZA VIRUS FOLLOWING EXCESS VITAMIN A

    PubMed Central

    Huang, J. S.; Bang, F. B.

    1964-01-01

    The conversion of chick embryonic epidermis to mucous epithelium by excess vitamin A in organ culture as reported by Fell and Mellanby (5) was shown to be accompanied by a corresponding change of susceptibility to influenza and vaccinia viruses. Untreated epidermis of 10- to 12-day chick embryos supported the growth of influenza (PR8) virus in organ cultures and a maximum infectivity (EID50) titer was reached 2 to 3 days after infection. At the same time) the epidermis showed squamous keratinization, beginning about the 4th day of cultivation. Addition of excess vitamin A (40 µg per ml) to the skin organ culture induced the following changes: (a) mucous metaplasia of the epidermis which was usually first evident after 4 to 5 days in the vitamin A medium, (b) increase in the daily and maximum yield of influenza virus, if the epidermis had been grown for 4 or more days in the vitamin A medium before infection took place, and (c) decrease in the production of vaccinia virus under similar conditions. The maximum yield of both viruses remained unchanged, however, if excess vitamin A was introduced to the organ culture at the time of virus inoculation. The magnitude of increase in the yield of influenza virus in this organ culture system was found to be proportionally related to the concentration of vitamin A added 4 or more days before inoculation of this virus. Increasing doses of vitamin A however, had no effect on the short-term growth of influenza virus in tissue cultures of chorio-allantoic membrane. Observation on the early period (2 to 12 hours) of influenza virus growth initiated in the 4-day organ cultures of chick embryonic skin showed no significant difference in virus production between the normal and the vitamin A medium groups. The change of virus specificity apparently is not due to the presence of excess vitamin A per se, but appears to be related to the change of differentiation produced in the organ culture system. PMID:14206436

  6. Simultaneous Growth of Rabies and Canine Distemper Viruses in Chick Embryos

    PubMed Central

    Chang, James C. C.

    1965-01-01

    Rabies virus and canine distemper virus were grown simultaneously, and possibly symbiotically, in the same chick embryos. There seemed to be no adverse effect on either virus when cultured in such manner. Bivalent vaccines for rabies and canine distemper were produced. The potencies and the virus titers of such vaccines were comparable to those of rabies vaccine and canine distemper vaccine produced separately. PMID:14290942

  7. Induction of metallothionein in chick embryos as a mechanism of tolerance to platinum group metal exposure.

    PubMed

    Gagnon, Zofia E; Patel, Amit

    2007-02-15

    Recent data show that platinum group metals (PGMs), primarily platinum (Pt), palladium (Pd) and rhodium (Rd), from automobile catalytic converters are being deposited in the environment. We investigated the PGM neurotoxicity and tolerance mechanism by induction of metallothionein (MT) in developing chick embryos. Chick embryos were injected on the 7th and 14th days of incubation with different concentrations of Pt and mixture of Pt, Pd and Rh (PGM mix) solutions. It is documented that induction of MT by zinc (Zn+2) protects against metal and non-metal hepatotoxicity. In this study the MT induction was examined through pretreatment of the two highest Pt(IV) exposure levels with exogenous Zn2+ on the 4th and 11th days of incubation. SDS-PAGE assay and digital image system were used to identify and quantify MT in homogenized brain and liver tissues. Quantitative analysis revealed an increase of MT in the 5 ppm Pt exposure as compared to controls. The 10 ppm Pt treatment was a lethal dose for exposed embryos. There was increased mortality at the 1.0 PGM mix level. The interaction of Pt, Pd and Rh in the mixture seems to favor metal accumulation and MT induction in the liver but not the brain. Pretreatment with exogenous Zn2+ increased chick survival. These results indicate that induction of MT plays a protective role against PGM toxicity. Metal analysis using atomic absorption spectrometer in graphite furnace mode (GFAAS) revealed PGM accumulation in chick embryo liver and brain tissues proportional to exposure concentration. Our results may imply that MT has an important role as a tolerance mechanism against PGM toxicity. The presence of Pt(IV) in brain tissue suggests that the undeveloped blood-brain barrier is permeable to PGMs. This raises concerns regarding the implication of these metals on neural injury.

  8. Systematic assessment of blood circulation time of functionalized upconversion nanoparticles in the chick embryo

    NASA Astrophysics Data System (ADS)

    Nadort, Annemarie; Liang, Liuen; Grebenik, Ekaterina; Guller, Anna; Lu, Yiqing; Qian, Yi; Goldys, Ewa; Zvyagin, Andrei

    2015-12-01

    Nanoparticle-based delivery of drugs and contrast agents holds great promise in cancer research, because of the increased delivery efficiency compared to `free' drugs and dyes. A versatile platform to investigate nanotechnology is the chick embryo chorioallantoic membrane tumour model, due to its availability (easy, cheap) and accessibility (interventions, imaging). In our group, we developed this model using several tumour cell lines (e.g. breast cancer, colon cancer). In addition, we have synthesized in-house silica coated photoluminescent upconversion nanoparticles with several functional groups (COOH, NH2, PEG). In this work we will present the systematic assessment of their in vivo blood circulation times. To this end, we injected chick embryos grown ex ovo with the functionalized UCNPs and obtained a small amount of blood at several time points after injection to create blood smears The UCNP signal from the blood smears was quantified using a modified inverted microscope imaging set-up. The results of this systematic study are valuable to optimize biochemistry protocols and guide nanomedicine advancement in the versatile chick embryo tumour model.

  9. Growth Retardation Of Chick Embryo Exposed To A Low Dose Of Electromagnetic Waves.

    PubMed

    Siddiqi, Najam; John C, Muthusami; Norrish, Mark; Heming, Thomas

    2016-01-01

    The objectives of this study were to explore the effects of low dose of the nonionizing (REW) emitted by a mobile phone on the development of chick embryo. one hundred and twenty chick fertilized eggs were equally divided into a control and an exposed group. Sixty fertilized eggs were placed in an egg incubator with a mobile phone (SAR US: 1.10W/kg (head) 0.47 W/kg body) in silent mode having vibration disable mode. Mobile was called for a total of 20 minutes in 24 hours. Twenty embryos each were sacrificed at day 5, 10 and 15, mortality, wet body weight, head to rump length, eye diameter and morphological changes were noted. The control group, 60 eggs were incubated in the same conditions, having removed the phone. No mortality was noted. The experimental group exposed to REW showed subcutaneous haemorrhagic areas and significant growth retardation at day 10 as evidence by smaller eye diameter, wet weight and CR length than the control group. There were no significant growth differences at either day 5 or at day 15. Electromagnetic waves emitted from mobile phones even though for a very short duration of 20 minutes per day have affected the growth of the chick embryo at day 10 of incubation, Hence exposure of these waves are not 100% safe.

  10. [A new method for quantitative measurement of the cadmium absorbed by chick embryos].

    PubMed

    Gottofrey, J

    1984-01-01

    We attempted to determine the quantity of cadmium incorporated in hens eggs after immersion in cadmium solutions, and the cadmium concentration measured in embryos. We discussed equipment allowing simultaneous treatment of up to 42 samples, and called it " digestor ". It consisted of two gas-heated sand baths, two stands for cooling down solutions and an evacuation system for toxic vapours. Our method was based on wet mineralisation. It consisted of desintegrating experimental chick embryos in a HNO3/H2O2 mixed solution. After heating and evaporating, the quantity of cadmium in the remnant was determined by atomic absorption spectrophotometry. The reliability of such a technique was tested by studying as controls controls 17 days-old chick embryos injected with a known quantity of Cd(NO3). It showed no loss of cadmium. We also compared our procedure with a dry ashing method. The latter showed unacceptable losses and insufficient precision for the problems we wanted to investigate. Our method gave us much more precise results. The equipment we developed has functioned wholly satisfactorily and allowed us to investigate for instance cadmium distribution and concentration in embryonic organs of 17 days-old chicks. It could also be useful for researches concerning other biological samples analyzed for different heavy metals.

  11. Mechanical origins of rightward torsion in early chick brain development

    NASA Astrophysics Data System (ADS)

    Chen, Zi; Guo, Qiaohang; Dai, Eric; Taber, Larry

    2015-03-01

    During early development, the neural tube of the chick embryo undergoes a combination of progressive ventral bending and rightward torsion. This torsional deformation is one of the major organ-level left-right asymmetry events in development. Previous studies suggested that bending is mainly due to differential growth, however, the mechanism for torsion remains poorly understood. Since the heart almost always loops rightwards that the brain twists, researchers have speculated that heart looping affects the direction of brain torsion. However, direct evidence is lacking, nor is the mechanical origin of such torsion understood. In our study, experimental perturbations show that the bending and torsional deformations in the brain are coupled and that the vitelline membrane applies an external load necessary for torsion to occur. Moreover, the asymmetry of the looping heart gives rise to the chirality of the twisted brain. A computational model and a 3D printed physical model are employed to help interpret these findings. Our work clarifies the mechanical origins of brain torsion and the associated left-right asymmetry, and further reveals that the asymmetric development in one organ can induce the asymmetry of another developing organ through mechanics, reminiscent of D'Arcy Thompson's view of biological form as ``diagram of forces''. Z.C. is supported by the Society in Science - Branco Weiss fellowship, administered by ETH Zurich. L.A.T acknowledges the support from NIH Grants R01 GM075200 and R01 NS070918.

  12. The effect of ethanol vapour exposure on atrial and ventricular walls of chick embryos.

    PubMed

    Kamran, Kiran; Khan, Muhammad Yunus; Minhas, Liaqat Ali

    2016-10-01

    To study the effects of ethanol vapour exposure on atrial and ventricular walls of heart in chick embryo. The study design was experimental, conducted at Islamabad Centre of College of Physicians and Surgeons, Pakistan. One hundred and eighty chicken eggs were divided into two groups, experimental and control, of 90 eggs each. Each group was subdivided into three subgroups of 30 eggs each based on the day of sacrifice. Experimental group was exposed to ethanol vapours and then compared with age matched controls. The thickness of atrial and ventricular walls along with lengths of valvular cusps increased in hearts of day 7 and day 10 chick embryos in experimental group. There was thinning of walls and decreased length of valvular cusps in hearts of experimental chicks on hatching as compared to age matched controls. Ethanol vapour exposure during development causes cardiac and septal wall thickening during initial days of development followed by cardiac and septal wall thinning which is a classical picture of alcohol induced cardiomyopathies.

  13. The effects of thermal manipulations during embryogenesis of broiler chicks on growth of embryo and skeletal traits

    NASA Astrophysics Data System (ADS)

    Aygün, Ali; Narinç, Doǧan

    2016-04-01

    Incubation temperature is one of the important environmental factors that can induce epigenetic thermal adaptation of different physiological control systems. Thus, post hatch thermo tolerance ability of birds may be gained using these manipulations during different incubation periods. The current study was carried out to reveal the effects of temperature manipulations during early and late embryogenesis on weight of embryo and size of skeletal bilateral traits (face, wings, metatarsus, tibia, and femur) in broiler chicken embryos. One thousand commercial broiler eggs from 46 week old breeder flock were used in study. Treatments consisted of eggs incubated at 37.8°C and 55% relative humidity throughout (control; DG1), heated to 36.9°C and supplied 60% relative humidity for 6 hours daily from day 0 to 8 (DG2), heated to 36.9°C and supplied 60% relative humidity for 6 hours daily from day 10 to 18 (DG3), heated to 41°C and supplied 65% relative humidity for 3 hours daily from day 8 to 10 (DG4), and heated to 41°C and supplied 65% relative humidity for 3 hours daily from day 16 to 18 (DG5). Measurements of embryo weight and bilateral traits were obtained at 20 day of incubation and at hatch (at day 21). It was determined that the live weights of embryo and chick were affected significantly by treatment; DG3 group has shown higher mean values than the other treatment groups (P<0.05). There were differences in lengths of femur, tibia and metatarsus among treatment groups at hatch. Particularly, the high incubator temperatures at the second half of incubation accelerated growth of body and bone in embryos. These consequences of the treatments performed at different temperatures and times indicate that the different metabolic shifts realized by the embryos.

  14. The effects of thermal manipulations during embryogenesis of broiler chicks on growth of embryo and skeletal traits

    SciT

    Aygün, Ali, E-mail: aaygun@selcuk.edu.tr; Narinç, Doğan, E-mail: narincd@gmail.com

    Incubation temperature is one of the important environmental factors that can induce epigenetic thermal adaptation of different physiological control systems. Thus, post hatch thermo tolerance ability of birds may be gained using these manipulations during different incubation periods. The current study was carried out to reveal the effects of temperature manipulations during early and late embryogenesis on weight of embryo and size of skeletal bilateral traits (face, wings, metatarsus, tibia, and femur) in broiler chicken embryos. One thousand commercial broiler eggs from 46 week old breeder flock were used in study. Treatments consisted of eggs incubated at 37.8°C and 55%more » relative humidity throughout (control; DG1), heated to 36.9°C and supplied 60% relative humidity for 6 hours daily from day 0 to 8 (DG2), heated to 36.9°C and supplied 60% relative humidity for 6 hours daily from day 10 to 18 (DG3), heated to 41°C and supplied 65% relative humidity for 3 hours daily from day 8 to 10 (DG4), and heated to 41°C and supplied 65% relative humidity for 3 hours daily from day 16 to 18 (DG5). Measurements of embryo weight and bilateral traits were obtained at 20 day of incubation and at hatch (at day 21). It was determined that the live weights of embryo and chick were affected significantly by treatment; DG3 group has shown higher mean values than the other treatment groups (P<0.05). There were differences in lengths of femur, tibia and metatarsus among treatment groups at hatch. Particularly, the high incubator temperatures at the second half of incubation accelerated growth of body and bone in embryos. These consequences of the treatments performed at different temperatures and times indicate that the different metabolic shifts realized by the embryos.« less

  15. Chick embryogenesis: a unique platform to study the effects of environmental factors on embryo development.

    PubMed

    Yahav, S; Brake, J

    2014-01-01

    . Manipulating environmental temperature during the period of egg storage, during the intermediate pre-incubation period, and incubation period per se has been found to significantly affect embryo development, hatching progress, chick quality at hatching, and chick development post-hatching. These temperature manipulations have also been shown to affect the acquisition of thermotolerance to subsequent post-hatching thermal challenge. This chapter will focus on: a. "maternal effects" on embryo and post-hatching development; b. environmental effects during the post-ovipositional period of egg storage, the intermediate pre-incubation period, and incubation period per se on chick embryogenesis and subsequent post-hatching growth and development; and c. effects of temperature manipulations during the pre-incubation and incubation periods on acquisition of thermotolerance and development of secondary sexual characteristics in broiler chickens.

  16. Dose related shifts in the developmental progress of chick embryos exposed to mobile phone induced electromagnetic fields.

    PubMed

    Zareen, Nusrat; Khan, Muhammad Yunus; Minhas, Liaqat Ali

    2009-01-01

    The possible adverse effects of Electromagnetic Fields (EMFs) emitted from mobile phones present a major public concern today. Some studies indicate EMFs effects on genes, free radical production, immunological and carcinogenic effects. On the other hand there are studies which do not support the hypothesis of any biological impacts of EMFs. This study was designed to observe the effects of mobile phone induced EMFs on survival and general growth and development of chick embryo, investigating dose-response relationship if any. This was an experimental study in which developing chick embryos were exposed to different doses of mobile phone induced EMFs. For this purpose a mobile phone was placed in the incubator in the centre of fertilised eggs in silent ringing mode and was 'rung' upon from any other line or cell phone. After incubation for 10 or 15 days the eggs were opened and the developmental mile-stones of the surviving embryos were compared with the non exposed subgroup. EMFs exposure significantly decreased the survivability of the chick embryos. The lower doses of EMFs caused growth retardation. However, this effect of growth retardation reallocated to partial growth enhancement on increasing the dose of EMFs and shifted over to definite growth enhancement on further raising the dose. There is an adverse effect of EMFs exposure on embryo survivability. Chick embryos developmental process is influenced by EMFs. However, these effects are variable depending upon the dose of EMFs exposure.

  17. In vivo marking of single cells in chick embryos using photoactivation of GFP.

    PubMed

    Stark, D A; Kulesa, P M

    2005-10-01

    Selective marking of a single cell within a living embryo is often difficult due to the inaccuracy and invasiveness of standard techniques. This unit describes a minimally invasive optical protocol that uses 405-nm laser light to photoactivate a variant of green fluorescent protein (PAGFP). This method takes advantage of the accessibility of the chick embryo to inject PAGFP into a region of interest and uses electroporation to deliver the construct into cells. This unit describes in detail how single and small groups of cells (n<10) that express PAGFP can be made visually distinguishable from the host population using the photoactivation process. Included is a means to maximize the fluorescence increase due to photoactivated GFP signal and to reduce photobleaching. Briefly outlined are previously developed chick culture and time-lapse imaging techniques to allow for the subsequent monitoring of photoactivated cell migratory behaviors. The technique has the potential to be a less-invasive, accurate tool for in vivo studies that involve following cell lineage and cell migration.

  18. Effects of posture on the respiratory mechanics of the chick embryo.

    PubMed

    Menna, Tara M; Mortola, Jacopo P

    2002-10-01

    In the chicken embryo, pulmonary ventilation and pulmonary gas exchange begin approximately one day before the completion of hatching. We asked to what extent the posture inside the egg, and the presence of the eggshell and membranes, may alter the mechanical behaviour of the respiratory system. The passive mechanical properties of the respiratory system were studied in chicken embryos during the internal pipping phase (rupture of the air cell) or the external pipping phase (hole in the eggshell). Tracheal pressure and changes in lung volume were recorded during mechanical ventilation, first, with the embryo curled up inside the egg, then again after exteriorization from the eggshell. In the internal pippers, respiratory system compliance increased and expiratory resistance decreased after exteriorization, whereas the mean inspiratory impedance did not change. In the external pippers, exteriorization had no significant effects on respiratory compliance, resistance, or impedance, and the values were similar to those of newly hatched chicks. We conclude that, in the chicken embryo, at a time when pulmonary ventilation becomes an important mechanism for gas exchange, the curled up posture inside the egg does not provide any significant mechanical constraint to breathing.

  19. Effect of antithyroid drug on chick embryos during the last week of development: delayed hatching and decreased cerebellar acetylcholinesterase activity.

    PubMed

    Haba, Gen; Nishigori, Hidekazu; Tezuka, Yu; Kagami, Keisuke; Sugiyama, Toru; Nishigori, Hideo

    2011-11-01

    Hypothyroid state during embryogenesis disturbs normal growth and brain development, influencing later life. To evaluate the harmful consequences of the state during embryogenesis using an animal model, we inhibited thyroid hormone biosynthesis in chick embryos by using methimazole (MMI). Typically, embryos were treated with MMI (20 µmol/egg) on day 14, and examined on specific days.  Of the control embryos, 94% hatched on day 21, whereas 0% and 60% of MMI-treated embryos hatched on days 21 and 24, respectively. MMI retarded the rates of bodyweight gain as well as liver and heart development, and delayed hatching. However, the external differences in appearance and differences in the weights of the newly hatched control chicks on day 21 and the MMI-treated chicks on day 24 were less obvious. Embryos treated with MMI exhibited increased mass in their brain parts on day 24. Most notably, the treatment resulted in a 1.35-fold increase in cerebellum weight compared to that of the untreated animals. Acetylcholinesterase activity in the cerebellum on the day of hatching decreased significantly to 0.85-fold that of the untreated controls. Thyroid hormone receptor β mRNA was detected from day 12 and dramatically expressed from day 19 to the day of hatching. The 'fertilized hen's egg-chick embryo-chick system' is an appropriate animal model for investigating the hypothyroid state during embryogenesis. Decreased cerebellar acetylcholinesterase activity after MMI treatment was assumed to relate to a mechanism of motor and cognitive deficits in congenital hypothyroidism. © 2011 The Authors. Journal of Obstetrics and Gynaecology Research © 2011 Japan Society of Obstetrics and Gynecology.

  20. Direct visualization of the Wntless-induced redistribution of WNT1 in developing chick embryos.

    PubMed

    Galli, Lisa M; Santana, Frederick; Apollon, Chantilly; Szabo, Linda A; Ngo, Keri; Burrus, Laura W

    2018-04-30

    Paracrine Wnt signals are critical regulators of cell proliferation, specification, and differentiation during embryogenesis. Consistent with the discovery that Wnt ligands are post-translationally modified with palmitoleate (a 16 carbon mono-unsaturated fatty acid), our studies show that the vast majority of bioavailable chick WNT1 (cWNT1) produced in stably transfected L cells is cell-associated. Thus, it seems unlikely that the WNT1 signal is propagated by diffusion alone. Unfortunately, the production and transport of vertebrate Wnt proteins has been exceedingly difficult to study as few antibodies are able to detect endogenous Wnt proteins and fixation is known to disrupt the architecture of cells and tissues. Furthermore, vertebrate Wnts have been extraordinarily refractory to tagging. To help overcome these obstacles, we have generated a number of tools that permit the detection of WNT1 in palmitoylation assays and the visualization of chick and zebrafish WNT1 in live cells and tissues. Consistent with previous studies in fixed cells, live imaging of cells and tissues with overexpressed cWNT1-moxGFP shows predominant localization of the protein to a reticulated network that is likely to be the endoplasmic reticulum. As PORCN and WLS are important upstream regulators of Wnt gradient formation, we also undertook the generation of mCherry-tagged variants of both proteins. While co-expression of PORCN-mCherry had no discernible effect on the localization of WNT1-moxGFP, co-expression of WLS-mCherry caused a marked redistribution of WNT1-moxGFP to the cell surface and cellular projections in cultured cells as well as in neural crest and surface ectoderm cells in developing chick embryos. Our studies further establish that the levels of WLS, and not PORCN, are rate limiting with respect to WNT1 trafficking. Copyright © 2018. Published by Elsevier Inc.

  1. Retinoic acid, hemin and hexamethylen bisacetamide interference with "in vitro" differentiation of chick embryo chondrocytes.

    PubMed

    Manduca, P; Abelmoschi, M L

    1992-01-01

    We have investigated the effect of all-trans Retinoic acid, and of substances (Hemine and Hexamethylene bisacetamide) which interfere with "in vitro" differentiation of mesenchyme derived cell lineages on the expression of specific markers of hyperthrophy in "in vitro" differentiating chick embryo chondrocytes. (Castagnola P., et al., 1986). Continuous treatment of chondrogenic cells in conditions allowing differentiation "in vitro" with Retinoic acid resulted in persistence of type I collagen synthesis and in lack of type X collagen and Ch 21 protein expression. Hemin treated cells secreted a reduced amount of type X collagen. HMBA treatment inhibited type X collagen expression and caused reduction of the ratio between type II collagen and Ch 21 synthesized. The data indicate an independent regulation of these markers during chondrocyte differentiation.

  2. The Chick Embryo Chorioallantoic Membrane as an In Vivo Assay to Study Antiangiogenesis

    PubMed Central

    Ribatti, Domenico

    2010-01-01

    Antiangiogenesis, e.g., inhibition of blood vessel growth, is being investigated as a way to prevent the growth of tumors and other angiogenesis-dependent diseases. Pharmacological inhibition interferes with the angiogenic cascade or the immature neovasculature with synthetic or semi-synthetic substances, endogenous inhibitors or biological antagonists. The chick embryo chorioallantoic membrane (CAM) is an extraembryonic membrane, which serves as a gas exchange surface and its function is supported by a dense capillary network. Because its extensive vascularization and easy accessibility, CAM has been used to study morphofunctional aspects of the angiogenesis process in vivo and to study the efficacy and mechanism of action of pro- and anti-angiogenic molecules. The fields of application of CAM in the study of antiangiogenesis, including our personal experience, are illustrated in this review article. PMID:27713265

  3. Experimental tumor growth of canine osteosarcoma cell line on chick embryo chorioallantoic membrane (in vivo studies).

    PubMed

    Walewska, Magdalena; Dolka, Izabella; Małek, Anna; Wojtalewicz, Anna; Wojtkowska, Agata; Żbikowski, Artur; Lechowski, Roman; Zabielska-Koczywąs, Katarzyna

    2017-05-12

    The chick embryo chorioallantoic membrane (CAM) model is extensively used in human medicine in preclinical oncological studies. The CAM model has several advantages: low cost, simple experimental approach, time saving and following "3R principles". Research has shown that the human osteosarcoma cell lines U2OS, MMNG-HOS, and SAOS can form tumors on the CAM. In veterinary medicine, this has been described only for feline fibrosarcomas, feline mammary carcinomas and canine osteosarcomas. However, in case of canine osteosarcomas, it has been shown that only non-adherent osteosarcoma stem cells isolated from KTOSA5 and CSKOS cell lines have the ability to form microtumors on the CAM after an incubation period of 5 days, in contrast to adherent KTOSA5 and CSKOS cells. In the presented study, we have proven that the commercial adherent canine osteosarcoma cell line (D-17) can form vascularized tumors on the CAM after the incubation period of 10 days.

  4. NMR Based Cerebrum Metabonomic Analysis Reveals Simultaneous Interconnected Changes during Chick Embryo Incubation.

    PubMed

    Feng, Yue; Zhu, Hang; Zhang, Xu; Wang, Xuxia; Xu, Fuqiang; Tang, Huiru; Ye, Chaohui; Liu, Maili

    2015-01-01

    To find out if content changes of the major functional cerebrum metabolites are interconnected and formed a network during the brain development, we obtained high-resolution magic-angle-spinning (HR-MAS) 1H NMR spectra of cerebrum tissues of chick embryo aged from incubation day 10 to 20, and postnatal day 1, and analyzed the data with principal component analysis (PCA). Within the examined time window, 26 biological important molecules were identified and 12 of them changed their relative concentration significantly in a time-dependent manner. These metabolites are generally belonged to three categories, neurotransmitters, nutrition sources, and neuronal or glial markers. The relative concentration changes of the metabolites were interconnected among/between the categories, and, more interestingly, associated with the number and size of Nissl-positive neurons. These results provided valuable biochemical and neurochemical information to understand the development of the embryonic brain.

  5. NMR Based Cerebrum Metabonomic Analysis Reveals Simultaneous Interconnected Changes during Chick Embryo Incubation

    PubMed Central

    Feng, Yue; Zhu, Hang; Zhang, Xu; Wang, Xuxia; Xu, Fuqiang; Tang, Huiru; Ye, Chaohui; Liu, Maili

    2015-01-01

    To find out if content changes of the major functional cerebrum metabolites are interconnected and formed a network during the brain development, we obtained high-resolution magic-angle-spinning (HR-MAS) 1H NMR spectra of cerebrum tissues of chick embryo aged from incubation day 10 to 20, and postnatal day 1, and analyzed the data with principal component analysis (PCA). Within the examined time window, 26 biological important molecules were identified and 12 of them changed their relative concentration significantly in a time-dependent manner. These metabolites are generally belonged to three categories, neurotransmitters, nutrition sources, and neuronal or glial markers. The relative concentration changes of the metabolites were interconnected among/between the categories, and, more interestingly, associated with the number and size of Nissl-positive neurons. These results provided valuable biochemical and neurochemical information to understand the development of the embryonic brain. PMID:26485040

  6. Glycoconjugate sugar residues in the chick embryo developing lung: a lectin histochemical study.

    PubMed

    Gheri, G; Sgambati, E; Bryk, S G

    2000-03-01

    A lectin histochemical study was performed to investigate the distribution and changes of the oligosaccharidic component of the glycoconjugates in the lung of chick embryos, of 1-day-old chick, and of the adult animal. For this purpose, a battery of seven horseradish peroxidase-conjugated lectins (PNA, SBA, DBA, WGA, Con A, LTA, and UEA I) were employed. During the first phase of parabronchi and atria formation, D-galactose-(beta1-->3)-N-acetyl-D-galactosamine, beta-N-acetyl-D-galactosamine, D-glucosamine, alpha-D-mannose, and sialic acid, present at the level of the surface and of cytoplasmic granules of the lining epithelial cells, seem to play a role in regulating morphogenetic phenomena. In the subsequent phases, the parabronchial lumen and the atrial cavities were characterized by the presence of lectin-reactive material rich in terminal D-galactose-(beta1-->3)-N-acetyl-D-galactosamine, beta-N-acetyl-D-galactosamine, D-glucosamine and alpha-D-mannose. From day 18 onwards and immediately after hatching, the free border of the cells lining the air capillaries was characterized by the presence of beta-N-acetyl-D-galactosamine and alpha-D-mannose. The appearance of these sugar residues was concomitant with the beginning of respiratory activity. Copyright 2000 Wiley-Liss, Inc.

  7. 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.

  8. Teratogenic effects of the neonicotinoid thiacloprid on chick embryos (Gallus gallus domesticus).

    PubMed

    Salvaggio, Antonio; Antoci, Francesco; Messina, Antonino; Ferrante, Margherita; Copat, Chiara; Ruberto, Claudia; Scalisi, Elena Maria; Pecoraro, Roberta; Brundo, Maria Violetta

    2018-06-19

    Thiacloprid is an insecticide belonging to the family of neonicotinoids, substances initially underestimated for their potential adverse effects, that they may manifest in the long term leading to an extensive use. The objective of this study was to evaluate the effect at increasing concentrations of thiacloprid on chick embryos development. The research was carried out on 75 fertile eggs of Gallus gallus domesticus. The eggs were opened after 10, 15 and 20 days of incubation and in treated embryos were observed developmental alterations, growth retardation, limbs defects and ectopia viscerale. The histological analysis showed hepatic steatosis and haemorrhages both in the liver and in the lungs. Moreover, the immunohistochemical analysis performed on the liver sections showed a strong positivity only for the erythrocytes to the anti-CYP1A antibody. Thiacloprid exposure increases the risks of teratogenic effects especially at the higher doses tested, therefore its use should be more controlled and limited. Since the literature on the topic is lacking, then the human health impacts resulting from neonicotinoids exposure is not yet fully understood, and, our data will be helpful to allow the assessment of an oral reference dose and health risk characterization. Copyright © 2018. Published by Elsevier Ltd.

  9. Lectin histochemistry of goblet cell sugar residues in the gut of the chick embryo and of the newborn.

    PubMed

    Bryk, S G; Sgambati, E; Gheri Bryk, G

    1999-04-01

    The anlage of duodenum, ileum and colon were removed from chick embryos of day 8-21 of incubation and from 1-day-old chicks. A battery of seven different horseradish peroxidase-conjugated lectins (PNA, SBA, DBA, Con A, WGA, LTA and UEAI) was used to study the carbohydrate residues of the glycoconjugates in the goblet cells of the three parts of the intestine. The main results can be summarized as follows: differences in lectin binding were absent in the proximal and distal parts of the duodenum, ileum and colon. Lectin histochemistry showed differences among the three intestinal segments for the time of appearance of the oligosaccharides in the goblet mucus. In the colonic goblet cells of 1-day-old chicks, LTA and UEAI lectins showed two different types of linkage of alpha-L-fucose. This is the first demonstration of UEAI reactive sites in Gallus domesticus.

  10. Microgravity in the STS-29 space shuttle discovery affected the vestibular system of chick embryos

    NASA Technical Reports Server (NTRS)

    Fermin, C. D.; Martin, D.; Jones, T.; Vellinger, J.; Deuser, M.; Hester, P.; Hullinger, R.

    1996-01-01

    Out of 32 embryos flown (16 @ E2 + 16 @ E9) for 5 days, 16 survived. All sixteen E2 were dead at landing. Eight were opened and eight were incubated at 1.0G. Autopsy showed that 4 E2 survived over 24 hours in space. Eight E14 hatched without anatomical malformations, and 8 E14 were fixed. The height of the macular epithelia was 31 mu m (mean) in control and 26 mu m in flight chicks. The cross-sectional area of macular nuclei of control was 17 mu m(2) for hair cells and 14 mu m(2) in supporting cells. In flight, cross-sectional area was 17 mu m(2) in hair cells and 15 mu m(2) in supporting cells (n=250). The shape factor of cartilage cells (1.0 = perfect circle) between control (mean = 0.70) and flight (mean = 0.72), and the area of cartilaginous cells between controls (mean = 9 mu m(2)) and flight (mean = 9 mu m(2)) did not differ (n=300). The nuclei of support cells were closer to the basement membrane in flight than in control chicks. The immunoreactivity of otoconia with anti keratan, fibronectin or chrondroitin sulfate was not different between flight and control ears. There were more afferent fibers inside the macular epithelia of flight (p<0.05) than control. Three of 8 flight animals had elevated vestibular thresholds (VT), with normal mean response amplitudes and latencies. Modified afferent innervation patterns requiring weeks to compensate are sufficient to elevate VT, and should be investigated further. Other reversible (sublethal) microgravity effects on sensory epithelia (vacuoles, swelling, etc) require quantification.

  11. Secretion pathway of liver IGF-1 via JAK2/STAT3 in chick embryo under the monochromatic light.

    PubMed

    Wang, Tuanjie; Dong, Yulan; Wang, Zixu; Cao, Jing; Chen, Yaoxing

    2016-02-01

    This study reveals mechanism of monochromatic light on the IGF-1 secretion of chick embryo liver. The chick embryos were incubated and exposed to continuous red, green, blue light or a dark environment. Compared to other light-treated groups, green light increased IGF-1 and melatonin concentrations both in plasma and liver, and Mel1a, Mel1b and Mel1c receptors expressions in liver but decreased p-JAK2, p-STAT3 and ROS in liver. IGF-1 had a positive correlation with melatonin, but a negative relevance with p-JAK2 and p-STAT3. In vitro, the IGF-1 level in the hepatocyte supernatant was enhanced by melatonin with lower p-JAK2/p-STAT3 and ROS levels, which was suppressed by Mel1c antagonist but not Mel1a/Mel1b or Mel1b antagonists. AG490 (JAK/STAT inhibitor) promoted role of melatonin-Mel1c modulated IGF-1 secretion. These results suggest the antioxidant effect of melatonin mediated the green light-enhanced IGF-1 secretion of chick embryo liver through Mel1c receptor to inhibit the JAK2/STAT3 pathway.

  12. Extracting cardiac shapes and motion of the chick embryo heart outflow tract from four-dimensional optical coherence tomography images

    NASA Astrophysics Data System (ADS)

    Yin, Xin; Liu, Aiping; Thornburg, Kent L.; Wang, Ruikang K.; Rugonyi, Sandra

    2012-09-01

    Recent advances in optical coherence tomography (OCT), and the development of image reconstruction algorithms, enabled four-dimensional (4-D) (three-dimensional imaging over time) imaging of the embryonic heart. To further analyze and quantify the dynamics of cardiac beating, segmentation procedures that can extract the shape of the heart and its motion are needed. Most previous studies analyzed cardiac image sequences using manually extracted shapes and measurements. However, this is time consuming and subject to inter-operator variability. Automated or semi-automated analyses of 4-D cardiac OCT images, although very desirable, are also extremely challenging. This work proposes a robust algorithm to semi automatically detect and track cardiac tissue layers from 4-D OCT images of early (tubular) embryonic hearts. Our algorithm uses a two-dimensional (2-D) deformable double-line model (DLM) to detect target cardiac tissues. The detection algorithm uses a maximum-likelihood estimator and was successfully applied to 4-D in vivo OCT images of the heart outflow tract of day three chicken embryos. The extracted shapes captured the dynamics of the chick embryonic heart outflow tract wall, enabling further analysis of cardiac motion.

  13. Glassfrog embryos hatch early after parental desertion.

    PubMed

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

    2014-06-22

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

  14. Glassfrog embryos hatch early after parental desertion

    PubMed Central

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

    2014-01-01

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

  15. Apoptosis generates mechanical forces that close the lens vesicle in the chick embryo

    NASA Astrophysics Data System (ADS)

    Oltean, Alina; Taber, Larry A.

    2018-03-01

    During the initial stages of eye development, optic vesicles grow laterally outward from both sides of the forebrain and come into contact with the surrounding surface ectoderm (SE). Within the region of contact, these layers then thicken locally to create placodes and invaginate to form the optic cup (primitive retina) and lens vesicle (LV), respectively. This paper examines the biophysical mechanisms involved in LV formation, which consists of three phases: (1) lens placode formation; (2) invagination to create the lens pit (LP); and (3) closure to form a complete ellipsoidally shaped LV. Previous studies have suggested that extracellular matrix deposited between the SE and optic vesicle causes the lens placode to form by locally constraining expansion of the SE as it grows, while actomyosin contraction causes this structure to invaginate. Here, using computational modeling and experiments on chick embryos, we confirm that these mechanisms for Phases 1 and 2 are physically plausible. Our results also suggest, however, that they are not sufficient to close the LP during Phase 3. We postulate that apoptosis provides an additional mechanism by removing cells near the LP opening, thereby decreasing its circumference and generating tension that closes the LP. This hypothesis is supported by staining that shows a ring of cell death located around the LP opening during closure. Inhibiting apoptosis in cultured embryos using caspase inhibitors significantly reduced LP closure, and results from a finite-element model indicate that closure driven by cell death is plausible. Taken together, our results suggest an important mechanical role for apoptosis in lens development.

  16. Neural retina of chick embryo in organ culture: effects of blockade of growth factors by suramin.

    PubMed

    Cirillo, A; Chifflet, S; Villar, B

    2001-06-01

    The neural retina is a highly organized organ whose final histoarchitecture depends on the presence of diverse growth factors and on their interactions with extracellular matrix components. However, the role of growth factors on retinal development is not fully understood. Suramin has been shown to produce diverse cellular effects via the simultaneous block of the action of several growth factors. We have therefore studied the effects of suramin on organotypic culture of chick embryo neural retina in order to gain further insights into the participation of growth factors in neural retinal development. Neural retina was incubated for 24 h with suramin at 50-200 microM and then processed to determine cell proliferation, nuclear morphology, and actin distribution. Suramin provoked extensive morphological changes revealed by a decrease in BrdU incorporation, alterations in cellular organization, and disruption of the outer limiting membrane, with the emergence of cellular elements through it. All of these effects were dose-dependent and markedly attenuated by the simultaneous presence of suramin and fibroblast growth factor 2 (FGF-2) in the culture medium. These findings indicate that suramin induces pleiotropic effects on the histoarchitecture of the chicken neural retina in organ culture and suggest that FGF-2 is one of the biological modulators involved in the maintenance of the structural organization of the chicken neural retina.

  17. Effects of chromium picolinate on the viability of chick embryo fibroblast.

    PubMed

    Bai, Y; Zhao, X; Qi, C; Wang, L; Cheng, Z; Liu, M; Liu, J; Yang, D; Wang, S; Chai, T

    2014-04-01

    Chromium picolinate (CrPic), which is used as a nutritional supplement and to treat type 2 diabetes, has gained much attention because of its cytotoxicity. This study evaluated the effects of CrPic on the viability of the chick embryo fibroblast (CEF) using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, morphological detection, and flow cytometry. The results show that lower concentrations of CrPic (8 and 16 μM) did not damage CEF viability (p > 0.05). However, higher CrPic concentrations (400 and 600 μM) indicated a highly significant effect on the production of intracellular reactive oxygen species, alteration of mitochondrial membrane potential, intracellular calcium ion concentration, and the apoptosis rate (p < 0.01), contrary to lower CrPic concentrations (8 and 16 μM) and control group. Moreover, apoptotic morphological changes induced by these processes in CEF were confirmed using Hoechst 33258 staining. Cell death induced by higher concentrations of CrPic was caused by an apoptotic and a necrotic mechanism, whereas the main mechanism of oxidative stress-induced mitochondrial dysfunction was apoptotic death.

  18. Tissue morphodynamics shaping the early mouse embryo.

    PubMed

    Sutherland, Ann E

    2016-07-01

    Generation of the elongated vertebrate body plan from the initially radially symmetrical embryo requires comprehensive changes to tissue form. These shape changes are generated by specific underlying cell behaviors, coordinated in time and space. Major principles and also specifics are emerging, from studies in many model systems, of the cell and physical biology of how region-specific cell behaviors produce regional tissue morphogenesis, and how these, in turn, are integrated at the level of the embryo. New technical approaches have made it possible more recently, to examine the morphogenesis of the mouse embryo in depth, and to elucidate the underlying cellular mechanisms. This review focuses on recent advances in understanding the cellular basis for the early fundamental events that establish the basic form of the embryo. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Bidirectional Fusion of the Heart-forming Fields in the Developing Chick Embryo

    PubMed Central

    Moreno-Rodriguez, R.A.; Krug, E.L.; Reyes, L.; Villavicencio, L.; Mjaatvedt, C.H.; Markwald, R.R.

    2007-01-01

    It is generally thought that the early pre-tubular chick heart is formed by fusion of the anterior or cephalic limits of the paired cardiogenic fields. However, this study shows that the heart fields initially fuse at their midpoint to form a transitory “butterfly”-shaped, cardiogenic structure. Fusion then progresses bi-directionally along the longitudinal axis in both cranial and caudal directions. Using in vivo labeling, we demonstrate that cells along the ventral fusion line are highly motile, crossing future primitive segments. We found that mesoderm cells migrated cephalically from the unfused tips of the anterior/cephalic wings into the head mesenchyme in the region that has been called the secondary heart field. Perturbing the anterior/cranial fusion results in formation of a biconal heart. A theoretical role of the ventral fusion line acting as a “heart organizer” and its role in cardia bifida is discussed. PMID:16252277

  20. Mechanisms of reoxygenation-induced calcium overload in cultured chick embryo heart cells

    SciT

    Murphy, J.G.; Smith, T.W.; Marsh, J.D.

    1988-06-01

    We examined mechanisms by which Ca enters cultured myocardial cells during posthypoxic reoxygenation. Monolayer cultures of chick embryo ventricular cells were prepared from hearts 10 days in ovo. Cells were exposed to hypoxic conditions (PO/sub 2/ less than 1.5 Torr), and /sup 45/Ca uptake during subsequent reoxygenation was then examined in the absence and presence of modulators of Ca channel-dependent Ca entry and Na-Ca exchange. Modulation of Ca entry by free radical-scavenging enzymes was also examined. Hypoxia for 120 min followed by reoxygenation increased Ca content from 1.9 to 6.1 nmol/mg protein (P less than 0.05) at 30 min. Verapamilmore » (10(-5) M) added before reoxygenation reduced Ca overload to 3.1 +/- 0.2 nmol/mg protein (P less than 0.05), but both verapamil and BAY K 8644 were without effect on modulating Ca entry if added 30 min after reoxygenation. /sup 24/Na content of cells increased from 70 nmol/mg protein in control cells to 157 nmol/mg protein (P less than 0.05) after hypoxia and reoxygenation, favoring Ca entry via Na-Ca exchange. Dichlorobenzamil significantly ameliorated reoxygenation-induced Ca overload, as did catalase and superoxide dismutase. We conclude that reoxygenation-induced Ca overload is unlikely to occur via the Ca channel. It occurs in part via Na-Ca exchange and is substantially ameliorated by enzymatic O/sub 2/ free radical scavengers.« less

  1. Lectin histochemistry of enterocytes sugar residues in the gut of the chick embryo and of the newborn.

    PubMed

    Bryk, S Gheri; Gheri, G

    2002-01-01

    The glycoconjugates sugar residues content, distribution and changes in the enterocytes of different tracts of the developing intestine of the chick embryo and of 1-day-old chick were investigated, using a battery of seven HRP-conjugated lectins (DBA, SBA, PNA, WGA, ConA, LTA and UEAI). The results of the present research have shown the presence of a large amount of glycoconjugates sugar residues in the enterocytes of duodenal, ileal and colonic anlage, starting from the beginning of the second week of incubation. Differences were detected among the three investigated intestinal segments, as to the time of appearance of the glycoconjugates sugar residues in the enterocytes. The duodenal enterocytes showed the most precocious appearance of lectin-reactive material, followed by the ileal enterocytes and afterwards by colonic enterocytes. The duodenal enterocytes were characterised by the presence of SBA binding sites, which were not detectable in the duodenal enterocytes of the adult animal.

  2. Establishment of a chick embryo shell-less culture system and its use to observe change in behavior caused by nicotine and substances from cigarette smoke.

    PubMed

    Hamamichi, S; Nishigori, H

    2001-02-28

    Early detection and removal of harmful factors are essential to the proper physical and psychological development of the fetus, presumably showing the effects during the prenatal period and after birth. As one procedure to aid in understanding such factors, we have established a shell-less culture system for video monitoring to observe change in behavior of 7-day-old chick embryos. Nicotine and aqueous cigarette smoke extract (ACSE) were selected for the present experiments, and the results showed a complete stoppage of swing-like movements by administrations of 10 microg nicotine and 1xACSE, possibly displaying paralytic symptoms. Quantitative analysis of nicotine in 1xACSE indicated that more than 10 microg of nicotine were contained in 100 microl of the extract. The present system, although in initial stage of development, may be a useful preliminary screening procedure for perhaps supervision and warning about the environment surrounding pregnant women.

  3. Early development of the circumferential axonal pathway in mouse and chick spinal cord.

    PubMed

    Holley, J A

    1982-03-10

    The early development of the circumferential axonal pathway in the brachial and lumbar spinal cord of mouse and chick embryos was studied by scanning and transmission electron microscopy. The cellular processes which comprise this pathway grow in the transverse plane and along the lateral margin of the marginal zone (i.e., circumferentially oriented), as typified by the early embryonic commissural axons. The first formative event observed was in the ventrolateral margin of the primitive spinal cord ventricular zone. Cellular processes were found near the external limiting membrane that appeared to grow a variable distance either dorsally or ventrally. Later in development, presumptive motor column neurons migrated into the ventrolateral region, distal to these early circumferentially oriented processes. Concurrently, other circumferentially oriented perikarya and processes appeared along the dorsolateral margin. Due to their aligned sites of origin and parallel growth, the circumferential processes formed a more or less continuous line or pathway, which in about 10% of the scanned specimens could be followed along the entire lateral margin of the embryonic spinal cord. Several specimens later in development had two sets of aligned circumferential processes in the ventral region. Large numbers of circumferential axons were then found to follow the preformed pathway by fasciculation, after the primitive motor column had become established. Since the earliest circumferential processes appeared to differentiate into axons and were found nearly 24 hours prior to growth of most circumferential axons, their role in guidance as pioneering axons was suggested.

  4. Culture of domestic cat ovarian tissue in vitro and in the chick embryo chorioallantoic membrane.

    PubMed

    Vilela, J M V; Leonel, E C R; D'Oliveira, L; Paiva, R E G; Miranda-Vilela, A L; Amorim, C A; Pic-Taylor, A; Lucci, C M

    2016-10-15

    In vitro culture and transplantation procedures are essential protocols employed in the evaluation of ovarian follicle survival and development. Culture in the chorioallantoic membrane (CAM) of chick embryos is an intermediate method that provides important follicle development information and has not been tested for cat ovaries to date. The aim of this study was to investigate if in vitro and CAM culture could be used as short-term systems to study cat ovarian tissue development. The ovaries of eight cats were dissected into 3-mm(3) cubes, cultured in vitro and in CAM for up to 5 days, and stained with hematoxylin-eosin and Gomori trichrome. Cell proliferation was analyzed using anti-Ki67. Possible differences among groups were investigated by analysis of variance or the Kruskal-Wallis test followed by Bonferroni correction. The T-test or Wilcoxon test was used to verify differences between the CAM and IVC. Results revealed that 87.5% of all follicles were primordial during culture. The percentage of primordial follicles in the morphologically normal follicles (MNF) pool was always higher than 80%, with the exception of Day 3 of CAM culture, but the number of MNF reduced significantly from Day 0 (600 out of 777 follicles) to Day 5 in the CAM (91 out of 171) and IVC (296 out of 686). The number of primordial follicles in 1 mm(3) in Days 2, 3, and 5 in the CAM was significantly lower than that in the control (Day 0). No cellular proliferation was observed in culture. Vascularization occurred in the CAM culture, but with no association to follicular viability. In addition, both methods showed an increase in connective tissue during culture. Although no significant differences were observed in the percentage of MNF, there was a reduction in the total number of follicles, both for IVC and CAM-cultured ovarian tissue. Furthermore, anti-Ki67 did not stain any follicle after Day 0 in IVC or in CAM culture. Neither system was capable of promoting follicle growth and

  5. The culture of chick embryo mesoderm cells in hydrated collagen gels.

    PubMed

    Sanders, E J; Prasad, S

    1983-04-01

    Chick embryo mesoderm cells are various stages of differentiation were cultured in three-dimensional matrices of hydrated collagen. The tissues used were: stage 5 mesoderm from regions adjacent to the primitive streak; stage 12 mesoderm, comprising somitic, unsegmented (segmental plate) and lateral plate mesoderm; and stage 18 sclerotome. Explants were examined by phase contrast microscopy, including time-lapse, and scanning and transmission electron microscopy. The cells showed an increased ability to adhere to, and move in, the collagen gel with advancing stage. Of the stage 12 tissues, the unsegmented mesoderm was initially the slowest to grow out of the explant. Sclerotome cells showed by far the greatest ability to move within the gel. Where the collagen fibrils were randomly oriented, the cell morphology was polypodial and advancing lamellipodia showed clear undulations at their leading edges. A distinction was drawn between these undulations and the classical major ruffles which are seen in two-dimensional culture to uplift and pass back along the cell surface. The latter were not seen in the collagen matrix and were presumably suppressed by the three-dimensional culture configuration while the leading edge undulations were not. Ultrastructural examination showed that the cells possessed patches of amorphous material on their surface, which was sometimes interposed between the plasma membrane and collagen fibrils. Addition of hyaluronic acid (2 mg/ml) had an effect only the segmented mesoderm, where outgrowth was enhanced. Although the addition of plasma fibronectin (50 micrograms/ml) to the cultures did not affect any of the tissues, the removal of this substance, by antifibronectin antiserum or by the use of fibronectin depleted serum, inhibited outgrowth in most cases. The only tissue not reproducibly inhibited in this way was sclerotome. Alignment of the collagen fibres by the explants was observed, accompanied by an elongation of the outgrowing cells

  6. Chick embryo chorioallantoic membrane (CAM): an alternative predictive model in acute toxicological studies for anti-cancer drugs.

    PubMed

    Kue, Chin Siang; Tan, Kae Yi; Lam, May Lynn; Lee, Hong Boon

    2015-01-01

    The chick embryo chorioallantoic membrane (CAM) is a preclinical model widely used for vascular and anti-vascular effects of therapeutic agents in vivo. In this study, we examine the suitability of CAM as a predictive model for acute toxicology studies of drugs by comparing it to conventional mouse and rat models for 10 FDA-approved anticancer drugs (paclitaxel, carmustine, camptothecin, cyclophosphamide, vincristine, cisplatin, aloin, mitomycin C, actinomycin-D, melphalan). Suitable formulations for intravenous administration were determined before the average of median lethal dose (LD50) and median survival dose (SD(50)) in the CAM were measured and calculated for these drugs. The resultant ideal LD(50) values were correlated to those reported in the literature using Pearson's correlation test for both intravenous and intraperitoneal routes of injection in rodents. Our results showed moderate correlations (r(2)=0.42 - 0.68, P<0.005-0.05) between the ideal LD(50) values obtained using the CAM model with LD(50) values from mice and rats models for both intravenous and intraperitoneal administrations, suggesting that the chick embryo may be a suitable alternative model for acute drug toxicity screening before embarking on full toxicological investigations in rodents in development of anticancer drugs.

  7. Chick embryo chorioallantoic membrane (CAM): an alternative predictive model in acute toxicological studies for anti-cancer drugs

    PubMed Central

    KUE, Chin Siang; TAN, Kae Yi; LAM, May Lynn; LEE, Hong Boon

    2015-01-01

    The chick embryo chorioallantoic membrane (CAM) is a preclinical model widely used for vascular and anti-vascular effects of therapeutic agents in vivo. In this study, we examine the suitability of CAM as a predictive model for acute toxicology studies of drugs by comparing it to conventional mouse and rat models for 10 FDA-approved anticancer drugs (paclitaxel, carmustine, camptothecin, cyclophosphamide, vincristine, cisplatin, aloin, mitomycin C, actinomycin-D, melphalan). Suitable formulations for intravenous administration were determined before the average of median lethal dose (LD50) and median survival dose (SD50) in the CAM were measured and calculated for these drugs. The resultant ideal LD50 values were correlated to those reported in the literature using Pearson’s correlation test for both intravenous and intraperitoneal routes of injection in rodents. Our results showed moderate correlations (r2=0.42 − 0.68, P<0.005–0.05) between the ideal LD50 values obtained using the CAM model with LD50 values from mice and rats models for both intravenous and intraperitoneal administrations, suggesting that the chick embryo may be a suitable alternative model for acute drug toxicity screening before embarking on full toxicological investigations in rodents in development of anticancer drugs. PMID:25736707

  8. The effect of temporary reductions in incubation temperature on growth characteristics and lipid utilisation in the chick embryo

    PubMed Central

    FEAST, M.; NOBLE, R. C.; SPEAKE, B. K.; FERGUSON, M. W. J.

    1998-01-01

    The effect(s) of finite exposure(s) to 22.0 °C on embryo weight gain, 3rd tarsus length, transfer of lipid from the yolk and uptake of specific lipid components by the liver, were measured in the chick embryo. Embryo growth was completely suspended over the period of temperature reduction, but resumed unimpaired when the optimum temperature of 37.5 °C was restored. Two successive periods of embryonic exposure to 22.0 °C (36 h at d 3 followed by 24 at d 10) were additive in their effect on growth retardation. Embryos subjected to temperature reductions on d 3 of incubation displayed significant changes in lipid concentrations and polyunsaturated fatty acid profiles within the liver when measured at d 18 of incubation. Yolk lipid uptake was significantly reduced but not suspended by embryonic exposure to 22.0 °C. It is suggested that the observed suspension of embryonic development was the result of a temperature dependent reduction in metabolism according to a Q10∼2, which is typical of an ectothermic organism. PMID:9877293

  9. The expression dynamics of mechanosensitive genes in extra-embryonic vasculature after heart starts to beat in chick embryo.

    PubMed

    Rajendran, Saranya; Sundaresan, Lakshmikirupa; Rajendran, Krithika; Selvaraj, Monica; Gupta, Ravi; Chatterjee, Suvro

    2016-02-11

    Fluid flow plays an important role in vascular development. However, the detailed mechanisms, particularly the link between flow and modulation of gene expression during vascular development, remain unexplored. In chick embryo, the key events of vascular development from initiation of heart beat to establishment of effective blood flow occur between the stages HH10 and HH13. Therefore, we propose a novel in vivo model to study the flow experienced by developing endothelium. Using this model, we aimed to capture the transcriptome dynamics of the pre- and post-flow conditions. RNA was isolated from extra embryonic area vasculosa (EE-AV) pooled from three chick embryos between HH10-HH13 and RNA sequencing was performed. The whole transcriptome sequencing of chick identified up-regulation of some of the previously well-known mechanosensitive genes including NFR2, HAND1, CTGF and KDR. GO analyses of the up-regulated genes revealed enrichment of several biological processes including heart development, extracellular matrix organization, cell-matrix adhesion, cell migration, blood vessel development, patterning of blood vessels, collagen fibril organization. Genes encoding for gap junctions proteins which are involved in vascular remodeling and arterial-venous differentiation, and genes involved in cell-cell adhesion, and ECM interactions were significantly up-regulated. Validation of selected genes through semi quantitative PCR was performed. The study indicates that shear stress plays a major role in development. Through appropriate validation, this platform can serve as an in vivo model to study conditions of disturbed flow in pathology as well as normal flow during development.

  10. Method for Dissecting the Auditory Epithelium (Basilar Papilla) in Developing Chick Embryos.

    PubMed

    Levic, Snezana; Yamoah, Ebenezer N

    2016-01-01

    Chickens are an invaluable model for exploring auditory physiology. Similar to humans, the chicken inner ear is morphologically and functionally close to maturity at the time of hatching. In contrast, chicks can regenerate hearing, an ability lost in all mammals, including humans. The extensive morphological, physiological, behavioral, and pharmacological data available, regarding normal development in the chicken auditory system, has driven the progress of the field. The basilar papilla is an attractive model system to study the developmental mechanisms of hearing. Here, we describe the dissection technique for isolating the basilar papilla in developing chick inner ear. We also provide detailed examples of physiological (patch clamping) experiments using this preparation.

  11. Significance of chick quality score in broiler production.

    PubMed

    van de Ven, L J F; van Wagenberg, A V; Uitdehaag, K A; Groot Koerkamp, P W G; Kemp, B; van den Brand, H

    2012-10-01

    The quality of day old chicks is crucial for profitable broiler production, but a difficult trait to define. In research, both qualitative and quantitative measures are used with variable predictive value for subsequent performance. In hatchery practice, chick quality is judged on a binomial scale, as chicks are divided into first grade (Q1-saleable) and second grade (Q2) chicks right after hatch. Incidences and reasons for classifying chicks as Q2, and potential of these chicks for survival and post-hatch performance have hardly been investigated, but may provide information for flock performance. We conducted an experiment to investigate (1) the quality of a broiler flock and the relation with post-hatch flock performance based on a qualitative score (Pasgar©score) of Q1 chicks and based on the incidence of Q2 chicks and (2) the reasons for classifying chicks as Q2, and the potential of these chicks for survival and post-hatch growth. The performance was followed of Q1 and Q2 chicks obtained from two breeder flocks that hatched in two different hatching systems (a traditional hatcher or a combined hatching and brooding system, named Patio). Eggs were incubated until embryo day 18, when they were transferred to one of the two hatching systems. At embryo day 21/post-hatch day 0, all chicks from the hatcher (including Q2 chicks) were brought to Patio, where the hatchery manager marked the Q2 chicks from both flocks and hatching systems and registered apparent reasons for classifying these chicks as Q2. Chick quality was assessed of 100 Q1 chicks from each flock and hatching system. Weights of all chicks were determined at days 0, 7, 21 and 42. There were no correlations between mean Pasgar©score and post-hatch growth or mortality, and suboptimal navel quality was the only quality trait associated with lower post-hatch growth. Growth was clearly affected by breeder flock and hatching system, which could not be linked to mean Pasgar©score or incidence of Q2 chicks

  12. Fostering efficacy and toxicity evaluation of traditional Chinese medicine and natural products: Chick embryo as a high throughput model bridging in vitro and in vivo studies.

    PubMed

    Wu, Tong; Yu, Gui-Yuan; Xiao, Jia; Yan, Chang; Kurihara, Hiroshi; Li, Yi-Fang; So, Kwok-Fai; He, Rong-Rong

    2018-04-19

    Efficacy and safety assessments are essential thresholds for drug candidates from preclinical to clinical research. Conventional mammalian in vivo models cannot offer rapid pharmacological and toxicological screening, whereas cell-based or cell-free in vitro systems often lead to inaccurate results because of the lack of physiological environment. Within the avian species, gallus gallus is the first bird to have its genome sequencing. Meantime, chick embryo is an easily operating, relatively transparent and extensively accessible model, whose physiological and pathological alterations can be visualized by egg candler, staining and image technologies. These features facilitate chick embryo as a high-throughput screening platform bridging in vivo and in vitro gaps in the pharmaceutical research. Due to the complicated ingredients and multiple-targets natures of traditional Chinese medicine (TCM), testing the efficacy and safety of TCM by in vitro methods are laborious and inaccurate, while testing in mammalian models consume massive cost and time. As such, the productive living organism chick embryo serves as an ideal biological system for pharmacodynamics studies of TCM. Herein, we comprehensively update recent progresses on the specialty of chick embryo in evaluation of efficacy and toxicity of drugs, with special concerns of TCM. Copyright © 2018 Elsevier Ltd. All rights reserved.

  13. Dual Labeling of Neural Crest Cells and Blood Vessels Within Chicken Embryos Using ChickGFP Neural Tube Grafting and Carbocyanine Dye DiI Injection

    PubMed Central

    Delalande, Jean-Marie; Thapar, Nikhil; Burns, Alan J.

    2015-01-01

    All developing organs need to be connected to both the nervous system (for sensory and motor control) as well as the vascular system (for gas exchange, fluid and nutrient supply). Consequently both the nervous and vascular systems develop alongside each other and share striking similarities in their branching architecture. Here we report embryonic manipulations that allow us to study the simultaneous development of neural crest-derived nervous tissue (in this case the enteric nervous system), and the vascular system. This is achieved by generating chicken chimeras via transplantation of discrete segments of the neural tube, and associated neural crest, combined with vascular DiI injection in the same embryo. Our method uses transgenic chickGFP embryos for intraspecies grafting, making the transplant technique more powerful than the classical quail-chick interspecies grafting protocol used with great effect since the 1970s. ChickGFP-chick intraspecies grafting facilitates imaging of transplanted cells and their projections in intact tissues, and eliminates any potential bias in cell development linked to species differences. This method takes full advantage of the ease of access of the avian embryo (compared with other vertebrate embryos) to study the co-development of the enteric nervous system and the vascular system. PMID:26065540

  14. Dual labeling of neural crest cells and blood vessels within chicken embryos using Chick(GFP) neural tube grafting and carbocyanine dye DiI injection.

    PubMed

    Delalande, Jean-Marie; Thapar, Nikhil; Burns, Alan J

    2015-05-28

    All developing organs need to be connected to both the nervous system (for sensory and motor control) as well as the vascular system (for gas exchange, fluid and nutrient supply). Consequently both the nervous and vascular systems develop alongside each other and share striking similarities in their branching architecture. Here we report embryonic manipulations that allow us to study the simultaneous development of neural crest-derived nervous tissue (in this case the enteric nervous system), and the vascular system. This is achieved by generating chicken chimeras via transplantation of discrete segments of the neural tube, and associated neural crest, combined with vascular DiI injection in the same embryo. Our method uses transgenic chick(GFP) embryos for intraspecies grafting, making the transplant technique more powerful than the classical quail-chick interspecies grafting protocol used with great effect since the 1970s. Chick(GFP)-chick intraspecies grafting facilitates imaging of transplanted cells and their projections in intact tissues, and eliminates any potential bias in cell development linked to species differences. This method takes full advantage of the ease of access of the avian embryo (compared with other vertebrate embryos) to study the co-development of the enteric nervous system and the vascular system.

  15. The gga-let-7 family post-transcriptionally regulates TGFBR1 and LIN28B during the differentiation process in early chick development.

    PubMed

    Lee, Sang In; Jeon, Mi-Hyang; Kim, Jeom Sun; Jeon, Ik-Soo; Byun, Sung June

    2015-12-01

    Early chick embryogenesis is governed by a complex mechanism involving transcriptional and post-transcriptional regulation, although how post-transcriptional processes influence the balance between pluripotency and differentiation during early chick development have not been previously investigated. Here, we characterized the microRNA (miRNA) signature associated with differentiation in the chick embryo, and found that as expression of the gga-let-7 family increases through early development, expression of their direct targets, TGFBR1 and LIN28B, decreases; indeed, gga-let-7a-5p and gga-let-7b miRNAs directly bind to TGFBR1 and LIN28B transcripts. Our data further indicate that TGFBR1 and LIN28B maintain pluripotency by regulating POUV, NANOG, and CRIPTO. Therefore, gga-let-7 miRNAs act as post-transcriptional regulators of differentiation in blastodermal cells by repressing the expression of the TGFBR1 and LIN28B, which intrinsically controls blastodermal cell differentiation in early chick development. © 2015 Wiley Periodicals, Inc.

  16. Simultaneous real-time quantification of blood flow and vascular growth in the chick embryo using optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Kowalski, William J.; Teslovich, Nikola C.; Chen, Chia-Yuan; Keller, Bradley B.; Pekkan, Kerem

    2014-03-01

    Experimental and clinical data indicate that hemodynamic forces within the embryo provide critical biomechanical cues for cardiovascular morphogenesis, growth, and remodeling and that perturbed flow is a major etiology of congenital heart disease. However, embryonic flow-growth relationships are largely qualitative and poorly defined. In this work, we provide a quantitative analysis of in vivo flow and growth trends in the chick embryo using optical coherence tomography (OCT) to acquire simultaneous velocity and structural data of the right vitelline artery continuously over a ten hour period beginning at stage 16 (hour 54). We obtained 3D vessel volumes (15 μm lateral, 4.3 μm axial resolutions, 6 μm slice spacing) at 60 minute intervals, taking a B-scan time series totaling one cardiac cycle at each slice. Embryos were maintained at a constant 37°C and 60% humidity during the entire acquisition period through an inhouse built chamber. The 3D vessel lumen geometries were reconstructed manually to assess growth. Blood flow velocity was computed from the central B-scan using red blood cell particle image velocimetry. The use of extended OCT imaging as a non-invasive method for continuous and simultaneous flow and structural data can enhance our understanding of the biomechanical regulation of critical events in morphogenesis. Data acquired will be useful to validate predictive finite-element 3D growth models.

  17. Evidence against a direct role for oxidative stress in cadmium-induced axial malformation in the chick embryo

    SciT

    Thompson, Jennifer, E-mail: jennifer.thompson@ucd.i; Doi, Takashi; The Children's Research Centre, Our Lady's Children's Hospital, Dublin 12

    Cadmium (Cd) is a powerful inducer of oxidative stress. It also causes ventral body wall defects in chick embryos treated at Hamburger-Hamilton stages 16-17. By measuring malondialdehyde levels (TBARS method) and cotreating with antioxidants (tempol, ascorbate, and N-acetylcysteine), we sought to determine if oxidative stress were directly related to teratogenesis. We also investigated the expression of mRNAs for antioxidant enzymes superoxide dismutase (SOD) -1 and -2, catalase (CAT), and glutathione peroxidase (GPx). RT-PCR showed reductions in SOD-1, SOD-2, and CAT 1 hour after treatment with Cd. MDA levels increased 4 hours after Cd, and remained elevated 24 hours after treatment.more » Of the antioxidants, only N-acetylcysteine reduced MDA levels to control values. Nonetheless, no antioxidant could reduce embryo lethality or malformation rates. Furthermore, MDA levels 24 hours after treatment were identical in malformed and normal embryos exposed to Cd. Hence, we conclude that oxidative stress may not have a direct role in Cd teratogenesis.« less

  18. Neurotoxicological effects of nicotine on the embryonic development of cerebellar cortex of chick embryo during various stages of incubation.

    PubMed

    El-Beltagy, Abd El-Fattah B M; Abou-El-Naga, Amoura M; Sabry, Dalia M

    2015-10-01

    Long-acting nicotine is known to exert pathological effects on almost all tissues including the cerebellar cortex. The present work was designed to elucidate the effect of nicotine on the development of cerebellar cortex of chick embryo during incubation period. The fertilized eggs of hen (Gallus gallus domesticus) were injected into the air space by a single dose of long acting nicotine (1.6 mg/kg/egg) at the 4th day of incubation. The embryos were taken out of the eggs on days 8, 12 and 16 of incubation. The cerebellum of the control and treated embryos at above ages were processed for histopathological examination. The TEM were examined at 16th day of incubation. The results of the present study revealed that, exposure to long-acting nicotine markedly influence the histogenesis of cerebellar cortex of chick embryo during the incubation period. At 8th day of incubation, nicotine delayed the differentiation of the cerebellar analge; especially the external granular layer (EGL) and inner cortical layer (ICL). Furthermore, at 12th day of incubation, the cerebellar foliation was irregular and the Purkinje cells not recognized. By 16th day of incubation, the cerebellar foliations were irregular with interrupted cerebellar cortex and irregular arrangement of Purkinje cells. Immunohistochemical analysis for antibody P53 protein revealed that the cerebellar cortex in all stages of nicotine treated groups possessed a moderate to weak reaction for P53 protein however; this reaction was markedly stronger in the cerebellar cortex of control groups. Moreover, the flow cytometric analysis confirmed that the percentage of apoptosis in control group was significantly higher compared with that of nicotine treated group. At the TEM level, the cerebellar Purkinje cells of 16th day of treated groups showed multiple subcellular alterations in compared with those of the corresponding control group. Such changes represented by appearing of vacuolated mitochondria, cisternal

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

    PubMed

    Cloney, Kellie; Franz-Odendaal, Tamara Anne

    2015-01-24

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

  20. Neural differentiation of caudal cell mass (secondary neurulation) in chick embryos: Hamburger and Hamilton Stages 16-45.

    PubMed

    Yang, Hee-Jin; Wang, Kyu-Chang; Chi, Je G; Lee, Myung-Sook; Lee, Yun-Jin; Kim, Seung-Ki; Cho, Byung-Kyu

    2003-04-14

    In an attempt to understand the events in the secondary neurulation in embryonic stage, we investigated morphological changes in the tail bud of normal developing chick embryos. Hamburger and Hamilton stage 16-45 embryos were harvested and processed for light microscopic studies. The secondary neural tube is formed by aggregation of the caudal cell mass. Cells are arranged into a cord-like mass (medullary cord), which is continuous with the primary neural tube. Multiple small cavities develop in the medullary cord, and these cavities coalesce into one single lumen. The process of coalescence is completed by stage 35, and the whole neural tube is transformed into one tube with a single continuous lumen. At this stage, the terminal portion of the neural tube is bulged dorsally. Thereafter, the caudal portion of the neural tube regresses, and the proximal portion develops into normal spinal cord. Transient occlusion of the central canal was observed at stage 40 in one sample. The sequence of events elucidated in this study can be used as base-line data for experiments concerning congenital malformations involving secondary neurulation.

  1. Effects of maternal dietary manganese and incubation temperature on hatchability, antioxidant status, and expression of heat shock proteins in chick embryos.

    PubMed

    Zhu, Y W; Lu, L; Li, W X; Zhang, L Y; Ji, C; Lin, X; Liu, H C; Odle, J; Luo, X G

    2015-12-01

    To investigate whether supplementing manganese (Mn) to the maternal diet could reduce the deleterious effect of heat stress on the developing embryo, the hatchability, antioxidant status, and expression of heat shock proteins (HSP) were evaluated in chick embryos under normal and high incubation temperatures. A completely randomized design ( = 6) with 2 maternal dietary Mn treatments (unsupplemented control basal diet versus the basal diet + 120 mg Mn/kg as inorganic Mn) × 2 incubation temperatures (normal, 37.8°C, versus high, 39.0°C) was used. High incubation temperature did not affect ( > 0.19) hatchability and embryo mortality and development but did increase ( < 0.05) activities of heart manganese superoxide dismutase (MnSOD) and liver copper zinc superoxide dismutase and liver MnSOD mRNA and protein levels in embryos. High incubation temperature also decreased ( < 0.003) HSP70 protein level in the heart but had no effects ( > 0.07) in the liver of embryos. Maternal diet with Mn supplementation not only increased ( < 0.05) the hatchability and Mn content ( < 0.001) in the yolk and embryonic tissues and the activity of MnSOD in the heart ( < 0.004) as well as relative liver weight ( < 0.05) under normal incubation temperature but also decreased ( ≤ 0.05) embryo mortality and HSP90 mRNA level in the liver and heart of embryos. Furthermore, under high incubation temperature, maternal diet Mn supplementation increased ( < 0.002) MnSOD protein expression in the liver of embryos but had no effect ( > 0.43) under normal incubation temperature. These results indicated that high incubation temperature induced self-protective responses of chick embryos with a modification of antioxidant status and a depression of HSP70 protein level. Maternal dietary supplementation of Mn could improve the hatchability as well as antioxidant ability to protect against heat challenge in embryos during incubation.

  2. The Effect of Levetiracetam on Closure of the Midline in Early Chicken Embryos.

    PubMed

    Ozgural, Onur; Armagan, Ercan; Bozkurt, Melih; Eroglu, Umit; Kahilogullari, Gokmen; Unlu, Agahan

    2015-01-01

    Genetic predisposition and some environmental factors play an important role in the development of neural tube defects. Levetiracetam is a new drug that has been approved in the treatment of partial seizures. We aimed in this study to determine the effect of levetiracetam on chick embryos. One hundred and sixty fertile non-pathogenic Super Nick eggs were incubated for 24 hours and were divided into four groups of 40 eggs each. Levetiracetam was administered via the sub-blastodermic route. The eggs were incubated for another 24 hours. All eggs were opened at the 48th hour, and the embryos were evaluated morphologically and histopathologically. The effects of levetiracetam on the embryo were correlated with the dose of levetiracetam. In the light of the results, it was determined that the use of increasing doses of levetiracetam led to defects of midline closure in early chicken embryos. Levetiracetam, a new antiepileptic drug that is effective especially on calcium ion concentration, leads to defects in midline closure in embryos in a dose-dependent manner. Further studies are needed to show the mechanism of embryonic damage and the mechanisms of its teratogenous effects associated with genetic and environmental factors.

  3. Chicken Embryos as a Potential New Model for Early Onset Type I Diabetes

    PubMed Central

    Shi, Liheng; Ko, Michael L.; Huang, Cathy Chia-Yu; Park, So-Young; Hong, Min-Pyo; Ko, Gladys Y.-P.

    2014-01-01

    Diabetic retinopathy (DR) is the leading cause of blindness among the American working population. The purpose of this study is to establish a new diabetic animal model using a cone-dominant avian species to address the distorted color vision and altered cone pathway responses in prediabetic and early diabetic patients. Chicken embryos were injected with either streptozotocin (STZ), high concentration of glucose (high-glucose), or vehicle at embryonic day 11. Cataracts occurred in varying degrees in both STZ- and high glucose-induced diabetic chick embryos at E18. Streptozotocin-diabetic chicken embryos had decreased levels of blood insulin, glucose transporter 4 (Glut4), and phosphorylated protein kinase B (pAKT). In STZ-injected E20 embryos, the ERG amplitudes of both a- and b-waves were significantly decreased, the implicit time of the a-wave was delayed, while that of the b-wave was significantly increased. Photoreceptors cultured from STZ-injected E18 embryos had a significant decrease in L-type voltage-gated calcium channel (L-VGCC) currents, which was reflected in the decreased level of L-VGCCα1D subunit in the STZ-diabetic retinas. Through these independent lines of evidence, STZ-injection was able to induce pathological conditions in the chicken embryonic retina, and it is promising to use chickens as a potential new animal model for type I diabetes. PMID:25133191

  4. Investigation of mRNA expression for secreted frizzled-related protein 2 (sFRP2) in chick embryos.

    PubMed

    Lin, Chung-Tien; Lin, Yu-Ting; Kuo, Tzong-Fu

    2007-08-01

    The roles of secreted frizzled-related protein 2 (sFRP2) in organ development of vertebrate animals are not well understood. We investigated expression of sFRP2 during embryogenesis of Arbor Acre broiler chicken eggs. Expression of sFRP2 was detected in the folds and lateral layer of developing brains. The sFRP2 signals in the developing eye were marked as a circle along the orbit. In younger embryos on days 3-6, the sFRP2 signals were consistent with growth of the sclerotome, suggesting that sFRP2 may be associated with somite development. Furthermore, with the exception of bones, sFRP2 mRNA was detectable in the interdigital tissue of embryos older than eight days as the limbs matured. This revealed that sFRP2 might play a role in myogenesis. In situ hybridization was also used to analyze the expression of sFRP2 in day 3-10 chick embryos. Signals were expressed in the gray matter of the developing brain coelom, including the optic lobe, metencephalon, myelencephalon, mesencephalon and diencephalon. The developing eyes contained an intercellular distribution of sFRP2 in the pigmented layer of the retina and photoreceptors. Furthermore, sFRP2 was expressed in the mantle layer of the neural tube and notochord. Based on these findings, it seems reasonable to suggest that sFRP2 may play an active role in embryogenesis, especially in development of the neural system, eyes, muscles and limbs.

  5. Somitic disruption of GNAS in chick embryos mimics progressive osseous heteroplasia

    PubMed Central

    Cairns, Dana M.; Pignolo, Robert J.; Uchimura, Tomoya; Brennan, Tracy A.; Lindborg, Carter M.; Xu, Meiqi; Kaplan, Frederick S.; Shore, Eileen M.; Zeng, Li

    2013-01-01

    Progressive osseous heteroplasia (POH) is a rare developmental disorder of heterotopic ossification (HO) caused by heterozygous inactivating germline mutations in the paternal allele of the GNAS gene. Interestingly, POH lesions have a bewildering mosaic distribution. Using clinical, radiographic, and photographic documentation, we found that most of the 12 individuals studied had a lesional bias toward one side or the other, even showing exclusive sidedness. Most strikingly, all had a dermomyotomal distribution of HO lesions. We hypothesized that somatic mutations in a progenitor cell of somitic origin may act on a background of germline haploinsufficiency to cause loss of heterozygosity at the GNAS locus and lead to the unilateral distribution of POH lesions. Taking advantage of the chick system, we examined our hypothesis by mimicking loss of heterozygosity of GNAS expression using dominant-negative GNAS that was introduced into a subset of chick somites, the progenitors that give rise to dermis and muscle. We observed rapid ectopic cartilage and bone induction at the axial and lateral positions in a unilateral distribution corresponding to the injected somites, which suggests that blocking GNAS activity in a targeted population of progenitor cells can lead to mosaic ectopic ossification reminiscent of that seen in POH. PMID:23863715

  6. Rediscovering the chick embryo as a model to study retinal development

    PubMed Central

    2012-01-01

    The embryonic chick occupies a privileged place among animal models used in developmental studies. Its rapid development and accessibility for visualization and experimental manipulation are just some of the characteristics that have made it a vertebrate model of choice for more than two millennia. Until a few years ago, the inability to perform genetic manipulations constituted a major drawback of this system. However, the completion of the chicken genome project and the development of techniques to manipulate gene expression have allowed this classic animal model to enter the molecular age. Such techniques, combined with the embryological manipulations that this system is well known for, provide a unique toolkit to study the genetic basis of neural development. A major advantage of these approaches is that they permit targeted gene misexpression with extremely high spatiotemporal resolution and over a large range of developmental stages, allowing functional analysis at a level, speed and ease that is difficult to achieve in other systems. This article provides a general overview of the chick as a developmental model focusing more specifically on its application to the study of eye development. Special emphasis is given to the state of the art of the techniques that have made gene gain- and loss-of-function studies in this model a reality. In addition, we discuss some methodological considerations derived from our own experience that we believe will be beneficial to researchers working with this system. PMID:22738172

  7. Fluorescence histochemical and elec-ron microscopical observations on sympathetic ganglia of the chick embryo cultured with and without hydrocortisone.

    PubMed

    Hervonen, H; Eränkö, O

    1975-01-01

    Lumbar sympathetic ganglia of 12-day-old chick embryos were cultured in organ cultures for 14 days with 1, 10 or 100 mg/l of hydrocortisone or without it. Catecholamines were demonstrated by the formaldehyde-induced fluorescence method. For electron microscopy, the cultures were fixed with glutarialdehyde and osmium tetroxide. Two types of cells with catecholamine fluoresecence were observed in the control cultures: (1) weakly fluorescent sympathetic neurons and sympathicoblasts with long nerve fibres, which were the most common cell type in the explant, and (2) brightly fluorescent cells with or without fluorescent processes, which were less common and were scattered in the explant. Hydrocortisone caused a great increase in the number of the brightly fluorescent cells. With 10 mg/l of hydrocortisone the increase was about ten-fold as compared with the control cultures. There was no change in the morphology of the cells, nor could any change be observed in the fluorescence intensity by eye. Electron microscopically the mature neurons were the most common cell type on the surface of the culture, while more immature sympathicoblasts were seen in the deeper layers. Cells were also found which contained large numbers of catecholamine-strong granular vesicles 105-275 nm in diameter. These cells were infrequent. They had round vesicular nuclei and resembled also in other respects sympathicoblasts or young nerve cells. One such cell was found in mitotic division by electron microscopy. Hydrocortisone caused a marked increase in the number of these granule-containing cells and their processes. Cells which could have been classified as the small intensely fluorescent cells of the mammalian ganglion type or their electron microscopic equivalent, the granule-containing cells were found neither in the control cultures nor in the hydrocortisone-containing cultures. It is concluded that most brightly fluorescent cells in cultured sympathetic ganglia of the chick are nerve cells

  8. 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.

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

    PubMed

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

    2014-06-01

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

  10. SDOCT Doppler velocimetry for investigating the morphological influences on blood flow in the developing chick embryo heart

    NASA Astrophysics Data System (ADS)

    Davis, Anjul M.; Rothenberg, Florence G.; Law, Tzuo H.; Taber, Larry A.; Izatt, Joseph A.

    2007-02-01

    The onset of congenital heart disease (CHD) is believed to occur at very early stages of development. Investigations in the initiation and development of CHD has been hampered by the inability to image early stage heart structure and function, in vivo. Imaging small animals using optical coherence tomography (OCT) has filled a niche between the limited penetration depth of confocal microscopy and insufficient resolution from ultrasound. Previous demonstrations of chick heart imaging using OCT have entailed excision of, or arresting the heart to prevent motion artifacts. In this summary, we introduce SDOCT Doppler velocimetry as an enhancement of Doppler OCT for in vivo measurement of localized temporal blood flow dynamics. With this technique, dynamic velocity waveforms were measured in the outflow tract of the heart tube. These flow dynamics correlate to a finite element model of pulsatile flow and may lead to a further understanding of morphological influences on early heart development.

  11. Cell death during the development of the truncus and conus of the chick embryo heart.

    PubMed Central

    Hurle, J M; Ojeda, J L

    1979-01-01

    The presence of cell death in the walls of the truncus and conus of the developing chick heart was investigated by a variety of light and electron microscopic techniques. Necrotic areas were observed in the myocardial layer of the truncus and conus and within the mesenchymal cells of the truncoconal ridges and aortopulmonary septum. These necrotic zones appeared first at Stage 25-26 and reached their maximum extent at Stages 29-32 undergoing later progressive disappearance. The morphological changes of the degenerating cells detectable under both transmission and scanning electron microscopy are also reported. The possible role of cell death in the morphogenesis of the truncus and conus is discussed. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 Fig. 13 Fig. 14 Fig. 15 PMID:500497

  12. Impact of PCOS on early embryo cleavage kinetics.

    PubMed

    Wissing, M L; Bjerge, M R; Olesen, A I G; Hoest, T; Mikkelsen, A L

    2014-04-01

    This study investigated whether polycystic ovary syndrome (PCOS) affected early embryo development assessed by time-lapse analysis of embryo kinetics from fertilization to the blastocyst stage. This was a prospective cohort study of two pronuclei (2PN) embryos from 25 hyperandrogenic PCOS patients (110 2PN embryos), 26 normoandrogenic PCOS patients (140 2PN embryos) and 20 healthy, regularly cycling women (controls, 97 2PN embryos). Patients underwent the same baseline evaluation and the same ovarian stimulation from April 2010 to February 2013. Oocytes were fertilized by intracytoplasmic sperm injection and incubated in an EmbryoScope with pictures taken every 20 min in seven focal planes. Time to 2PN breakdown, first cleavage and cleavage to 3, 4, 5, 6, 7 and 8 cells, morula and blastocyst (t₂, t₃, t₄, t₅, t₆, t₇, t₈, t(M), t(B)) were annotated. Differences in embryo kinetics between groups were assessed by mixed modelling. Compared with controls, embryos from hyperandrogenic PCOS patients were significantly delayed at 2PN breakdown, t₂, t₃, t₄ and t₇ but not at t₅, t₆, t₈, t(M) or t(B). Embryos from hyperandrogenic PCOS women had developed slower from fertilization to the 8-cell stage compared with embryos from controls. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  13. Prenatal opiate withdrawal activates the chick embryo hypothalamic pituitary-adrenal axis and dilates vitelline blood vessels via serotonin(2) receptors.

    PubMed

    Schrott, Lisa M; Baumgart, Mary Irene; Zhang, Xuewei; Sparber, Sheldon B

    2002-10-01

    Opiate withdrawal during pregnancy may occur because of voluntary or forced detoxification, or from rapid cycling associated with exposure to short-acting "street" opiates. Thus, animal modeling of prenatal withdrawal and development of potential therapeutic interventions is important. Direct developmental effects of opiates and/or withdrawal can be studied using a chick model. In ovo administration of the long-acting opiate N-desmethyl-l-alpha-noracetylmethadol (NLAAM) induces opiate dependence in the chick embryo. We examined activation of the hypothalamic-pituitary-adrenal (HPA) axis (assessed via serum corticosterone) and hemodynamic changes (assessed as changes in apparent diameter of vitelline (extraembryonic) blood vessels) after chronic NLAAM exposure and naloxone (Nx)-precipitated withdrawal during late stages of embryogenesis. Nx-precipitated withdrawal increased corticosterone 2- to 4.5-fold and diameters of vitelline blood vessels by 15 to 45%. NLAAM exposure itself did not effect these measures. In a second set of experiments, isobutylmethylxanthine (IBMX), a phosphodiesterase inhibitor, was injected into eggs with embryos. IBMX similarly increased corticosterone and vitelline vessel diameter, with a similar time course and response magnitude. Previous studies found that serotonin(2) (5-HT(2)) receptors were involved in other withdrawal manifestations, so we determined whether they were likewise involved. Pretreatment with the 5-HT(2) antagonist ritanserin completely blocked HPA axis activation and vasodilation associated with both Nx-precipitated withdrawal and IBMX administration. This indicates that 5-HT(2) receptors, directly or indirectly, mediate these withdrawal manifestations in the chick embryo.

  14. Early age thermal conditioning immediately reduces body temperature of broiler chicks in a tropical environment.

    PubMed

    De Basilio, V; Requena, F; León, A; Vilariño, M; Picard, M

    2003-08-01

    Early age thermal conditioning (TC) durably improves resistance of broilers to heat stress and reduces body temperature (Tb). Three experiments on broiler chicks were conducted to evaluate the effects of TC at 5 d of age on Tb variation measured by thermometer between 4 and 7 d of age, under a tropical environment. Because manipulation of chickens to measure Tb with a thermometer may increase Tb, a preliminary experiment on 13 3-to-4-wk-old male broilers compared Tb measured by telemetry to Tb measured in the terminal colon during three successive periods at 22, 33, and 22 degrees C. During heat exposure, Tb rapidly increased by 0.9 degrees C and plateaued over 24 h. During the last period, seven of the broilers rapidly reduced Tb to a plateau lower than the initial Tb, although six broilers exhibited more variable Tb. Measurement by thermometer underestimated on average core Tb by 0.28 degrees C at 22 degrees C and by 0.57 degrees C at 33 degrees C, whereas Tb recorded by telemetry was not affected by manipulation of the chickens. TC reduced Tb 24 h later in the three experiments. Compared to unexposed control chicks (N), 12 h of TC at 40 degrees C did not significantly reduce Tb at 7 d of age, although 24 h did. TC at 38 and 40 degrees C over 24 h significantly reduced Tb variation from 4 to 7 d of age compared to N chicks, whereas 36 degrees C did not. Withdrawing feed from the chicks for 2 h prior to measurement did not significantly reduce Tb at 4 and 7 d of age, but Tb reduction due to TC was greater in fed chicks (0.28 degrees C) than in chicks without feed (0.05 degrees C). Early age thermal conditioning at 38 to 40 degrees C at 5 d of age for 24 h reduced body temperature of 7-d-old male broilers.

  15. Efficacy of early chick nutrition with Aloe vera and Azadirachta indica on gut health and histomorphometry in chicks.

    PubMed

    Sujatha, Tamilvanan; Abhinaya, Sivasankar; Sunder, Jai; Thangapandian, Marudhai; Kundu, Anandamoy

    2017-06-01

    This study was conducted with an aim of studying the efficacy of water supplements of Aloe vera and Azadirachta indica (neem) during pre-starter age (0-2 weeks) on gut health and histomorphometry in Vanaraja chicks. A total of 192 day old Vanaraja chicks were randomly assigned to one of four herbal water treatments throughout the experimental pre-starter stage (0-2 weeks) in a completely randomized design. Each treatment was given four replicates consisting of 12 chicks per replicate. Water treatments comprised T1: Control with regular antibiotic supplement, T2: 3 ml Aloe juice per chick per day, T3: 3 ml neem extract per chick per day, T4: 1.5 ml Aloe and 1.5 ml neem per chick per day. Gut culture was done for Escherichia coli and Lactobacillus sps. and gut histomorphometry in 24 gut samples at 14 days of age. This study revealed that supplementation of A. vera and neem in water significantly (p<0.05) reduced and increased the number of gut E. coli and Lactobacillus sps. Colonies, respectively, as compared to that of control groups; Villi was significantly (p<0.05) taller and broader on 14 days of age across the jejunum of chicks fed with neem supplementation as compared to that of control chicks. Significantly lower crypt depth (p<0.05) was observed in the duodenum of Aloe supplementation. Villus height: Crypt depth ratio of duodenum and jejunum was significantly (p<0.05) increased neem and Aloe supplementation in chicks as compared to their combination and control. Immediate post hatch supplementation of Aloe juice and neem extract in chicks improved the development and health of their gut.

  16. Efficacy of early chick nutrition with Aloe vera and Azadirachta indica on gut health and histomorphometry in chicks

    PubMed Central

    Sujatha, Tamilvanan; Abhinaya, Sivasankar; Sunder, Jai; Thangapandian, Marudhai; Kundu, Anandamoy

    2017-01-01

    Aim: This study was conducted with an aim of studying the efficacy of water supplements of Aloe vera and Azadirachta indica (neem) during pre-starter age (0-2 weeks) on gut health and histomorphometry in Vanaraja chicks. Materials and Methods: A total of 192 day old Vanaraja chicks were randomly assigned to one of four herbal water treatments throughout the experimental pre-starter stage (0-2 weeks) in a completely randomized design. Each treatment was given four replicates consisting of 12 chicks per replicate. Water treatments comprised T1: Control with regular antibiotic supplement, T2: 3 ml Aloe juice per chick per day, T3: 3 ml neem extract per chick per day, T4: 1.5 ml Aloe and 1.5 ml neem per chick per day. Gut culture was done for Escherichia coli and Lactobacillus sps. and gut histomorphometry in 24 gut samples at 14 days of age. Results: This study revealed that supplementation of A. vera and neem in water significantly (p<0.05) reduced and increased the number of gut E. coli and Lactobacillus sps. Colonies, respectively, as compared to that of control groups; Villi was significantly (p<0.05) taller and broader on 14 days of age across the jejunum of chicks fed with neem supplementation as compared to that of control chicks. Significantly lower crypt depth (p<0.05) was observed in the duodenum of Aloe supplementation. Villus height: Crypt depth ratio of duodenum and jejunum was significantly (p<0.05) increased neem and Aloe supplementation in chicks as compared to their combination and control. Conclusion: Immediate post hatch supplementation of Aloe juice and neem extract in chicks improved the development and health of their gut. PMID:28717305

  17. A model system for testing gene vectors using murine tumor cells on the chorioallantoic membrane of the chick embryo.

    PubMed

    Dani, Sergio U; Espindola, Rachel

    2002-06-30

    We developed a model system for testing gene vectors, based on the growth of murine tumors on the chorioallantoic membrane (CAM) of embryonic chickens. The ability of selected murine cells to grow on the CAM was rated according to the following criteria: i) formation of tumor masses; ii) metastasis formation; iii) reproducibility; iv) yield, indicated as the number of embryos surviving to assessment time with visible tumors on the CAM; v) maintainability of the cell, both in the original host and the embryonic chick, or 'shuttle maintainability'; vi) detection by the naked eye, and vii) cost/benefit relation. The murine melanoma cell lineage, B16F10, which efficiently forms distinct, pigmented tumor masses and metastases on the CAM, performed better in this model than the murine B61 cell line. In vitro transduction of B16F10 cells with a recombinant adenovirus carrying a construct of the E. coli LacZ gene followed by inoculation onto the CAM resulted in beta-galactosidase expression in the tumor mass growing on the CAM. This model is potentially applicable to preclinical evaluation of gene vectors, especially for gene therapy of cancer.

  18. Application of high-performance liquid chromatography to the determination of glyoxylate synthesis in chick embryo liver.

    PubMed

    Qureshi, A A; Elson, C E; Lebeck, L A

    1982-11-19

    The isolation and identification of three major alpha-keto end products (glyoxylate, pyruvate, alpha-ketoglutarate) of the isocitrate lyase reaction in 18-day chick embryo liver have been described. This was accomplished by the separation of these alpha-keto acids as their 2,4-dinitrophenylhydrazones (DNPHs) by high-performance liquid chromatography (HPLC). The DNPHs of alpha-keto acids were eluted with an isocratic solvent system of methanol-water-acetic acid (60:38.5:1.5) containing 5 mM tetrabutylammonium phosphate from a reversed-phase ultrasphere C18 (IP) and from a radial compression C18 column. The separation can be completed on the radial compression column within 15-20 min as compared to 30-40 min with a conventional reversed-phase column. Retention times and peak areas were integrated for both the assay samples and reference compounds. A relative measure of alpha-keto acid in the peak was calculated by comparison with the standard. The identification of each peak was done on the basis of retention time matching, co-chromatography with authentic compounds, and stopped flow UV-VIS scanning between 240 and 440 nm. Glyoxylate represented 5% of the total product of the isocitrate lyase reaction. Day 18 parallels the peak period of embryonic hepatic glycogenesis which occurs at a time when the original egg glucose reserve has been depleted.

  19. ULTRASTRUCTURAL ORGANIZATION OF CILIA AND BASAL BODIES OF THE EPITHELIUM OF THE CHOROID PLEXUS IN THE CHICK EMBRYO

    PubMed Central

    Doolin, Paul F.; Birge, Wesley J.

    1966-01-01

    Ultrastructural studies were performed on normal and abnormal cilia and basal bodies associated with the choroidal epithelium of the chick embryo. Tissues were prepared in each of several fixatives including: 1% osmium tetroxide, in both phosphate and veronal acetate buffers; 2% glutaraldehyde, followed by postfixation in osmium tetroxide; 1% potassium permanganate in veronal acetate buffer. Normal cilia display the typical pattern of 9 peripheral doublets and 2 central fibers, as well as a system of 9 secondary fibers. The latter show distinct interconnections between peripheral and central fibers. Supernumerary fibers were found to occur in certain abnormal cilia. The basal body is complex, bearing 9 transitional fibers at the distal end and numerous cross-striated rootlets at the proximal end. The distal end of the basal body is delimited by a basal plate of moderate density. The tubular cylinder consists of 9 triple fibers. The C subfibers end at the basal plate, whereas subfibers A and B continue into the shaft of the cilium. The 9 transitional fibers radiate out from the distal end of the basal body, ending in bulblike terminal enlargements which are closely associated with the cell membrane in the area of the basal cup. One or 2 prominent basal feet project laterally from the basal body. These structures characteristically show several dense cross-bands and, on occasion, are found associated with microtubules. PMID:5335827

  20. Binding of various ovotransferrin fragments to chick-embryo red cells.

    PubMed Central

    Oratore, A; D'Andrea, G; Moreton, K; Williams, J

    1989-01-01

    1. The ability of N- and C-terminal half-molecule fragments of hen ovotransferrin to interact with chick red blood cells (CERBC) has been studied under conditions that allow binding of the transferrin to transferrin receptors to take place, but not the delivery of iron to the cell. Two kinds of half-molecule fragments were used: (a) those which can associate with one another to give a dimer resembling native transferrin and (b) those which cannot associate in this way because they lack a few amino acid residues from their C-terminal ends. 2. Neither N nor C half-molecules alone can bind to the CERBC, but, when both are present, tight binding occurs. 3. Whether or not the half-molecules can associate with one another makes little difference to receptor binding. 4. Given that one of the half-molecules is iron-saturated, the presence or absence of iron in the contralateral half-molecule again makes little difference to receptor binding. PMID:2920021

  1. Expression of neuronal antigens and related ventral and dorsal proteins in the normal spinal cord and a surgically induced open neural tube defect of the spine in chick embryos: an immunohistochemical study.

    PubMed

    Lee, Do-Hun; Phi, Ji Hoon; Chung, You-Nam; Lee, Yun-Jin; Kim, Seung-Ki; Cho, Byung-Kyu; Kim, Dong Won; Park, Moon-Sik; Wang, Kyu-Chang

    2010-05-01

    The aims of this study were to elucidate the processes of neuronal differentiation and ventrodorsal patterning in the spinal cord of the chick embryo from embryonic day (E) 3 to E17 and to study the effect of a prenatal spinal open neural tube defect (ONTD) on these processes. Expression patterns of neuronal antigens (neuronal nuclear antigen, neurofilament-associated protein (NAP), and synaptophysin) and related ventral markers [sonic hedgehog, paired box gene (PAX)6, and islet-1], and dorsal markers (bone morphogenetic protein, Notch homolog 1, and PAX7) were investigated in the normal spinal cord and in a surgically induced spinal ONTD in chick embryos. Four normal and ONTD chick embryos were used for each antigen group. There were no differences in the expression of neuronal and ventrodorsal markers between the control and ONTD groups. NAP and synaptophysin were useful for identifying dorsal structures in the distorted anatomy of the ONTD chicks.

  2. A descriptive study to provide evidence of the teratogenic and cellular effects of sibutramine and ephedrine on cardiac- and liver-tissue of chick embryos.

    PubMed

    Oberholzer, Hester Magdalena; Van Der Schoor, Ciska; Taute, Helena; Bester, Megan Jean

    2015-08-01

    Exposure to drugs during pregnancy is a major concern, as some teratogenic compounds can influence normal foetal development. Although the use of drugs during pregnancy should generally be avoided, exposure of the developing foetus to teratogens may occur unknowingly since these compounds may be hidden in products that are being marketed as "all natural." The aim of the current study was to investigate the possible teratogenic and cellular effects of sibutramine-a serotonin-norepinephrine reuptake inhibitor used in the treatment of obesity-on the heart and liver tissue of chick embryos. Ephedrine was used as a positive control. The chick embryo model was chosen because it has been used in studying developmental and experimental biology and teratology with great success. The embryos were exposed to three different concentrations of sibutramine and ephedrine respectively. The results obtained revealed that both compounds exhibited embryotoxicity when compared to the control groups. Liver and heart tissue of the exposed embryos was severely affected by these compounds in a dose-related manner. Morphology similar to that of muscle dystrophy was observed in the heart, where the muscle tissue was infiltrated by adipose and connective tissue. Severe liver steatosis was also noted. A more in-depth investigation into the molecular pathways involved might provide more information on the exact mechanism of toxicity of these products influencing embryonic development. © 2015 Wiley Periodicals, Inc.

  3. Effect of Radiofrequency Radiation Emitted from 2G and 3G Cell Phone on Developing Liver of Chick Embryo - A Comparative Study.

    PubMed

    D'Silva, Mary Hydrina; Swer, Rijied Thompson; Anbalagan, J; Rajesh, Bhargavan

    2017-07-01

    The increasing scientific evidence of various health hazards on exposure of Radiofrequency Radiation (RFR) emitted from both the cell phones and base stations have caused significant media attention and public discussion in recent years. The mechanism of interaction of RF fields with developing tissues of children and fetuses may be different from that of adults due to their smaller physical size and variation in tissue electromagnetic properties. The present study may provide an insight into the basic mechanisms by which RF fields interact with developing tissues in an embryo. To evaluate the possible tissue and DNA damage in developing liver of chick embryo following chronic exposure to Ultra-High Frequency/Radiofrequency Radiation (UHF/RFR) emitted from 2G and 3G cell phone. Fertilized chick embryos were incubated in four groups. Group A-experimental group exposed to 2G radiation (60 eggs), Group B- experimental group exposed to 3G radiation (60 eggs), Group C- sham exposed control group (60 eggs) and Group D- control group (48 eggs). On completion of scheduled duration, the embryos were collected and processed for routine histological studies to check structural changes in liver. The nuclear diameter and karyorrhexis changes of hepatocytes were analysed using oculometer and square reticule respectively. The liver procured from one batch of eggs from all the four groups was subjected to alkaline comet assay technique to assess DNA damage. The results were compared using one-way ANOVA test. In our study, the exposure of developing chick embryos to 2G and 3G cell phone radiations caused structural changes in liver in the form of dilated sinusoidal spaces with haemorrhage, increased vacuolations in cytoplasm, increased nuclear diameter and karyorrhexis and significantly increased DNA damage. The chronic exposure of chick embryo liver to RFR emitted from 2G and 3G cell phone resulted in various structural changes and DNA damage. The changes were more pronounced in 3

  4. Effect of Radiofrequency Radiation Emitted from 2G and 3G Cell Phone on Developing Liver of Chick Embryo – A Comparative Study

    PubMed Central

    Swer, Rijied Thompson; Anbalagan, J.; Rajesh, Bhargavan

    2017-01-01

    Introduction The increasing scientific evidence of various health hazards on exposure of Radiofrequency Radiation (RFR) emitted from both the cell phones and base stations have caused significant media attention and public discussion in recent years. The mechanism of interaction of RF fields with developing tissues of children and fetuses may be different from that of adults due to their smaller physical size and variation in tissue electromagnetic properties. The present study may provide an insight into the basic mechanisms by which RF fields interact with developing tissues in an embryo. Aim To evaluate the possible tissue and DNA damage in developing liver of chick embryo following chronic exposure to Ultra-High Frequency/Radiofrequency Radiation (UHF/RFR) emitted from 2G and 3G cell phone. Materials and Methods Fertilized chick embryos were incubated in four groups. Group A-experimental group exposed to 2G radiation (60 eggs), Group B- experimental group exposed to 3G radiation (60 eggs), Group C- sham exposed control group (60 eggs) and Group D– control group (48 eggs). On completion of scheduled duration, the embryos were collected and processed for routine histological studies to check structural changes in liver. The nuclear diameter and karyorrhexis changes of hepatocytes were analysed using oculometer and square reticule respectively. The liver procured from one batch of eggs from all the four groups was subjected to alkaline comet assay technique to assess DNA damage. The results were compared using one-way ANOVA test. Results In our study, the exposure of developing chick embryos to 2G and 3G cell phone radiations caused structural changes in liver in the form of dilated sinusoidal spaces with haemorrhage, increased vacuolations in cytoplasm, increased nuclear diameter and karyorrhexis and significantly increased DNA damage. Conclusion The chronic exposure of chick embryo liver to RFR emitted from 2G and 3G cell phone resulted in various structural

  5. Effect of Mobile Phone Radiation on Cardiovascular Development of Chick Embryo.

    PubMed

    Ye, W; Wang, F; Zhang, W; Fang, N; Zhao, W; Wang, J

    2016-06-01

    The biological effects on cardiovascular development of chicken embryos were examined after radiation exposure using mobile phone (900 MHz; specific absorption rate˜1.07 W/kg) intermittently 3 h per day during incubation. Samples were selected by morphological and histological methods. The results showed the rate of embryonic mortality and cardiac deformity increased significantly in exposed group (P < 0.05). No any histological pathological changes were observed on Day 5-7 (D5-D7) of incubation. A higher distribution of lipid droplets was unexpectedly present in myocardial tissue from the exposure groups on D10-D13. Soon afterwards, myofilament disruption, atrioventricular valve focal necrosis, mitochondria vacuolization and atrial natriuretic peptide (ANP) decrease appeared on D15-D21 of incubation. Comet assay data showed the haemocyte mean tail in the exposed group was significantly larger than that of the control (P < 0.01). The arterial vascular wall of exposed group was thicker (P < 0.05) than that of the control on D13, which was reversed to normal in later stages. Our findings suggest that long-term exposure of MPR may induce myocardium pathological changes, DNA damage and increased mortality; however, there was little effect on vascular development. © 2015 Blackwell Verlag GmbH.

  6. Development of a chick embryo heart cell for the cultivation of poliovirus.

    PubMed

    PRIER, J E; SULLIVAN, R

    1959-04-17

    An epithelial-like cell has been developed in line culture that apparently is stable. Although initially isolated cells were incapable of supporting the growth of poliovirus, the cells of the sixth and later passages allowed virus to propagate. The early, nonsusceptible cells were fibroblastic in appearance, in contrast to the epithelial type, poliovirussusceptible, derived cell of later passages.

  7. Toxicity of scorpion venom in chick embryo and mealworm assay depending on the use of the soluble fraction versus the whole venom.

    PubMed

    van der Valk, Tom; van der Meijden, Arie

    2014-09-01

    The LD50 is an important metric for venom studies and antivenom development. It has been shown that several variables in the protocol influence the LD50 value obtained, such as venom source, extraction and treatment and administration route. These inconsistencies reduce the utility of the results of these test for comparative studies. In scorpion venom LD50 assays, often only the soluble fraction of the venom is used, whereas other studies use the whole venom. We here tested the toxicity of the soluble fraction in isolation, and of the whole venom in two different systems: chick embryos and mealworms Tenebrio molitor. Ten microliters of venom solutions from Hadrurus arizonensis, Leiurus quinquestriatus, Androctonus australis, Grosphus grandidieri and Heterometrus laoticus were applied to five day old chicken embryos at stage 25-27. Our results showed no significant differences between the LD50 based on the whole venom versus that of only the soluble fraction and in the chicken embryo assay in four of the five scorpion species tested. H. laoticus however, showed a significantly lower LD50 value for the whole venom than the soluble fraction. In assays on mealworms however, this pattern was not seen. Nonetheless, caution may be warranted when using LD50 values obtained from only the soluble fraction. The LD50 values of the five species in this study, based on the chicken embryo assay, showed good correlation with values from the literature based on mouse studies. This suggests that the chick embryo assay may be an economic alternative to rodent assays for scorpion LD50 studies. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Effects of early environmental enrichment upon open-field behavior and timidity in the domestic chick.

    PubMed

    Jones, R B

    1982-03-01

    Early environmental enrichment was evaluated by its effect on the behavior of 7-day-old male and female domestic chicks in an open field or novel environment and in a hole-in-the-wall test of timidity. The chicks were housed in same-sex groups of 10. The bare environments contained wood litter, food, and water, whereas the enriched boxes also contained various objects. In the open field, immobility was lower while feeding, walking, jumping and vocalization were higher in the enriched birds than in those reared in the bare environment. Enrichment also decreased emergence latencies in the hole-in-the-wall box. The increased stimulation provided by environmental enrichment may decrease fearfulness in subsequent fear-inducing situations and may enhance the ability of animals to adapt to novelty.

  9. Effects of Optically Imposed Astigmatism on Early Eye Growth in Chicks

    PubMed Central

    Chu, Chin Hung Geoffrey; Kee, Chea Su

    2015-01-01

    Purpose To determine the effects of optically imposed astigmatism on early eye growth in chicks. Methods 5-day-old (P5) White Leghorn chicks were randomly assigned to either wear, monocularly, a “high magnitude” (H: +4.00DS/-8.00DC) crossed-cylindrical lens oriented at one of four axes (45, 90, 135, and 180; n = 20 in each group), or were left untreated (controls; n = 8). Two additional groups wore a “low magnitude” (L: +2.00DS/−4.00DC) cylindrical lens orientated at either axis 90 or 180 (n = 20 and n = 18, respectively). Refractions were measured at P5 and after 7 days of treatment for all chicks (P12), whereas videokeratography and ex-vivo eyeshape analysis were performed at P12 for a subset of chicks in each group (n = 8). Results Compared to controls, chicks in the treatment groups developed significant amounts of refractive astigmatism (controls: 0.03±0.22DC; treatment groups: 1.34±0.22DC to 5.51±0.26DC, one-way ANOVAs, p≤0.05) with axes compensatory to those imposed by the cylindrical lenses. H cylindrical lenses induced more refractive astigmatism than L lenses (H90 vs. L90: 5.51±0.26D vs. 4.10±0.16D; H180 vs. L180: 2.84±0.44D vs. 1.34±0.22D, unpaired two-sample t-tests, both p≤0.01); and imposing with-the-rule (H90 and L90) and against-the-rule astigmatisms (H180 and L180) resulted in, respectively, steeper and flatter corneal shape. Both corneal and internal astigmatisms were moderately to strongly correlated with refractive astigmatisms (Pearson’s r: +0.61 to +0.94, all p≤0.001). In addition, the characteristics of astigmatism were significantly correlated with multiple eyeshape parameters at the posterior segments (Pearson’s r: -0.27 to +0.45, all p≤0.05). Conclusions Chicks showed compensatory ocular changes in response to the astigmatic magnitudes imposed in this study. The correlations of changes in refractive, corneal, and posterior eyeshape indicate the involvement of anterior and posterior ocular segments during the

  10. Experimental analysis of control mechanisms in somite segmentation in avian embryos. II. Reduction of material in the gastrula stages of the chick.

    PubMed

    Bellairs, R; Veini, M

    1984-02-01

    A new theory of control of somite segmentation in chick embryos is proposed. This supposses that tiny clusters of already programmed cells are present throughout the presumptive somite area at stage 4, but that in order to fulfill their destiny they probably depend on the addition of further cells from the primitive streak. Evidence is based on the two groups of experiments: a) Experiments involving transection across the primitive streak at various stages, (which results in a 'tail' which possesses mesodermal derivatives) and across the segmental plate (which results in a 'tail' lacking mesodermal derivatives). b) Experiments in which parts of embryos have been explanted with or without their primitive streak. It is suggested that the initial clusters of pre-programmed cells move further and further posteriorly, developing into somitomeres (the precursors of true somites) only as they receive re-inforcements from the primitive streak or, ultimately, from the tail bud.

  11. Abnormal early cleavage events predict early embryo demise: sperm oxidative stress and early abnormal cleavage.

    PubMed

    Burruel, Victoria; Klooster, Katie; Barker, Christopher M; Pera, Renee Reijo; Meyers, Stuart

    2014-10-13

    Human embryos resulting from abnormal early cleavage can result in aneuploidy and failure to develop normally to the blastocyst stage. The nature of paternal influence on early embryo development has not been directly demonstrated although many studies have suggested effects from spermatozoal chromatin packaging, DNA damage, centriolar and mitotic spindle integrity, and plasma membrane integrity. The goal of this study was to determine whether early developmental events were affected by oxidative damage to the fertilizing sperm. Survival analysis was used to compare patterns of blastocyst formation based on P2 duration. Kaplan-Meier survival curves demonstrate that relatively few embryos with short (<1 hr) P2 times reached blastocysts, and the two curves diverged beginning on day 4, with nearly all of the embryos with longer P2 times reaching blastocysts by day 6 (p < .01). We determined that duration of the 2nd to 3rd mitoses were sensitive periods in the presence of spermatozoal oxidative stress. Embryos that displayed either too long or too short cytokineses demonstrated an increased failure to reach blastocyst stage and therefore survive for further development. Although paternal-derived gene expression occurs later in development, this study suggests a specific role in early mitosis that is highly influenced by paternal factors.

  12. Cross-neutralization of antibodies induced by vaccination with Purified Chick Embryo Cell Vaccine (PCECV) against different Lyssavirus species

    PubMed Central

    Malerczyk, Claudius; Freuling, Conrad; Gniel, Dieter; Giesen, Alexandra; Selhorst, Thomas; Müller, Thomas

    2014-01-01

    Background: Rabies is a neglected zoonotic disease caused by viruses belonging to the genus lyssavirus. In endemic countries of Asia and Africa, where the majority of the estimated 60,000 human rabies deaths occur, it is mainly caused by the classical rabies virus (RABV) transmitted by dogs. Over the last decade new species within the genus lyssavirus have been identified. Meanwhile 15 (proposed or classified) species exist, including Australian bat lyssavirus (ABLV), European bat lyssavirus (EBLV-1 and -2), Duvenhage virus (DUVV), as well as Lagos bat virus (LBV) and Mokola virus (MOKV) and recently identified novel species like Bokeloh bat lyssavirus (BBLV), Ikoma bat lyssavirus (IKOV) or Lleida bat lyssavirus (LLBV). The majority of these lyssavirus species are found in bat reservoirs and some have caused human infection and deaths. Previous work has demonstrated that Purified Chick Embryo Cell Rabies Vaccine (PCECV) not only induces immune responses against classical RABV, but also elicits cross-neutralizing antibodies against ABLV, EBLV-1 and EBLV-2. Material & Methods: Using the same serum samples as in our previous study, this study extension investigated cross-neutralizing activities of serum antibodies measured by rapid fluorescent focus inhibition test (RFFIT) against selected other non-classical lyssavirus species of interest, namely DUVV and BBLV, as well as MOKV and LBV. Results: Antibodies developed after vaccination with PCECV have neutralizing capability against BBLV and DUVV in the same range as against ABLV and EBLV-1 and -2. As expected, for the phylogenetically more distant species LBV no cross-neutralizing activity was found. Interestingly, 15 of 94 serum samples (16%) with a positive neutralizing antibody titer against RABV displayed specific cross-neutralizing activity (65-fold lower than against RABV) against one specific MOKV strain (Ethiopia isolate), which was not seen against a different strain (Nigeria isolate). Conclusion: Cross

  13. Cross-neutralization of antibodies induced by vaccination with Purified Chick Embryo Cell Vaccine (PCECV) against different Lyssavirus species.

    PubMed

    Malerczyk, Claudius; Freuling, Conrad; Gniel, Dieter; Giesen, Alexandra; Selhorst, Thomas; Müller, Thomas

    2014-01-01

    Rabies is a neglected zoonotic disease caused by viruses belonging to the genus lyssavirus. In endemic countries of Asia and Africa, where the majority of the estimated 60,000 human rabies deaths occur, it is mainly caused by the classical rabies virus (RABV) transmitted by dogs. Over the last decade new species within the genus lyssavirus have been identified. Meanwhile 15 (proposed or classified) species exist, including Australian bat lyssavirus (ABLV), European bat lyssavirus (EBLV-1 and -2), Duvenhage virus (DUVV), as well as Lagos bat virus (LBV) and Mokola virus (MOKV) and recently identified novel species like Bokeloh bat lyssavirus (BBLV), Ikoma bat lyssavirus (IKOV) or Lleida bat lyssavirus (LLBV). The majority of these lyssavirus species are found in bat reservoirs and some have caused human infection and deaths. Previous work has demonstrated that Purified Chick Embryo Cell Rabies Vaccine (PCECV) not only induces immune responses against classical RABV, but also elicits cross-neutralizing antibodies against ABLV, EBLV-1 and EBLV-2. Using the same serum samples as in our previous study, this study extension investigated cross-neutralizing activities of serum antibodies measured by rapid fluorescent focus inhibition test (RFFIT) against selected other non-classical lyssavirus species of interest, namely DUVV and BBLV, as well as MOKV and LBV. Antibodies developed after vaccination with PCECV have neutralizing capability against BBLV and DUVV in the same range as against ABLV and EBLV-1 and -2. As expected, for the phylogenetically more distant species LBV no cross-neutralizing activity was found. Interestingly, 15 of 94 serum samples (16%) with a positive neutralizing antibody titer against RABV displayed specific cross-neutralizing activity (65-fold lower than against RABV) against one specific MOKV strain (Ethiopia isolate), which was not seen against a different strain (Nigeria isolate). Cross-neutralizing activities partly correlate with the

  14. HNK-1 immunoreactivity during early morphogenesis of the head region in a nonmodel vertebrate, crocodile embryo

    NASA Astrophysics Data System (ADS)

    Kundrát, Martin

    2008-11-01

    The present study examines HNK-1 immunoidentification of a population of the neural crest (NC) during early head morphogenesis in the nonmodel vertebrate, the crocodile ( Crocodylus niloticus) embryos. Although HNK-1 is not an exclusive NC marker among vertebrates, temporospatial immunoreactive patterns found in the crocodile are almost consistent with NC patterns derived from gene expression studies known in birds (the closest living relatives of crocodiles) and mammals. In contrast to birds, the HNK-1 epitope is immunoreactive in NC cells at the neural fold level in crocodile embryos and therefore provides sufficient base to assess early migratory events of the cephalic NC. I found that crocodile NC forms three classic migratory pathways in the head: mandibular, hyoid, and branchial. Further, I demonstrate that, besides this classic phenotype, there is also a forebrain-derived migratory population, which consolidates into a premandibular stream in the crocodile. In contrast to the closely related chick model, crocodilian premandibular and mandibular NC cells arise from the open neural tube suggesting that species-specific heterochronic behavior of NC may be involved in the formation of different vertebrate facial phenotypes.

  15. The use of the chick embryo chorioallantoic membrane as experimental model to study virus growth and to test the clonal selection hypothesis. The contribution of Sir Mac Farlane Burnet.

    PubMed

    Ribatti, Domenico

    2018-06-07

    Sir Mac Farlane Burnet was the most honored of all Australian scientists. In 1960, Burnet shared the Nobel Prize for Medicine with Peter Medawar of Britain for the discovery of acquired immunological tolerance. He developed techniques for growing influenza viruses in the chorioallantoic membrane of the chick embryo. This became a standard laboratory practice. He continued to work with chick embryos long after the use of cell cultures had become general. His virology research resulted in significant discoveries concerning the nature and replication of viruses and their interaction with the immune system. Copyright © 2018 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

  16. Motor neurons with limb-innervating character in the cervical spinal cord are sculpted by apoptosis based on the Hox code in chick embryo.

    PubMed

    Mukaigasa, Katsuki; Sakuma, Chie; Okada, Tomoaki; Homma, Shunsaku; Shimada, Takako; Nishiyama, Keiji; Sato, Noboru; Yaginuma, Hiroyuki

    2017-12-15

    In the developing chick embryo, a certain population of motor neurons (MNs) in the non-limb-innervating cervical spinal cord undergoes apoptosis between embryonic days 4 and 5. However, the characteristics of these apoptotic MNs remain undefined. Here, by examining the spatiotemporal profiles of apoptosis and MN subtype marker expression in normal or apoptosis-inhibited chick embryos, we found that this apoptotic population is distinguishable by Foxp1 expression. When apoptosis was inhibited, the Foxp1 + MNs survived and showed characteristics of lateral motor column (LMC) neurons, which are of a limb-innervating subtype, suggesting that cervical Foxp1 + MNs are the rostral continuation of the LMC. Knockdown and misexpression of Foxp1 did not affect apoptosis progression, but revealed the role of Foxp1 in conferring LMC identity on the cervical MNs. Furthermore, ectopic expression of Hox genes that are normally expressed in the brachial region prevented apoptosis, and directed Foxp1 + MNs to LMC neurons at the cervical level. These results indicate that apoptosis in the cervical spinal cord plays a role in sculpting Foxp1 + MNs committed to LMC neurons, depending on the Hox expression pattern. © 2017. Published by The Company of Biologists Ltd.

  17. Comparison of safety and immunogenicity of purified chick embryo cell vaccine using Zagreb and Essen regimens in patients with category II exposure in China.

    PubMed

    Hu, Quan; Liu, Man-Qing; Zhu, Zheng-Gang; Zhu, Ze-Rong; Lu, Sha

    2014-01-01

    The aim was to compare the safety and immunogenicity of purified chick embryo cell vaccine (PCECV) with Zagreb 2-1-1 and Essen 1-1-1-1-1 regimens in patients with WHO category II exposure in China. Side effects including systemic and local symptoms were recorded for all patients during vaccination with purified chick embryo cell vaccine (PCECV) under Zagreb 2-1-1 or Essen 1-1-1-1-1 regimens, and the rabies neutralization antibody titers in patients' serum at days 0, 7, 14, 45, 365 post-immunization were measured to determine the immunogenicity. Fever and pain were the most common events for systemic and local symptoms respectively, and most side effects (86.78%, 105/121) occurred after the first dose of vaccination. Safety analysis showed differences in side effects in<5-year-old patients between Zagreb and Essen regimens, especially after the first dose of vaccination (P = 0.043). Immunogenicity analysis indicated that Zagreb can achieve higher neutralization antibody titers and a greater seroconversion rate in a shorter time but had less persistence than Essen. When compared with the Essen regimen, the Zagreb regimen had a different immunogenicity in all study subjects, and different safety profile in young children, and a further study with a larger population and longer surveillance is warranted.

  18. 3D Reconstruction of Chick Embryo Vascular Geometries Using Non-invasive High-Frequency Ultrasound for Computational Fluid Dynamics Studies.

    PubMed

    Tan, Germaine Xin Yi; Jamil, Muhammad; Tee, Nicole Gui Zhen; Zhong, Liang; Yap, Choon Hwai

    2015-11-01

    Recent animal studies have provided evidence that prenatal blood flow fluid mechanics may play a role in the pathogenesis of congenital cardiovascular malformations. To further these researches, it is important to have an imaging technique for small animal embryos with sufficient resolution to support computational fluid dynamics studies, and that is also non-invasive and non-destructive to allow for subject-specific, longitudinal studies. In the current study, we developed such a technique, based on ultrasound biomicroscopy scans on chick embryos. Our technique included a motion cancelation algorithm to negate embryonic body motion, a temporal averaging algorithm to differentiate blood spaces from tissue spaces, and 3D reconstruction of blood volumes in the embryo. The accuracy of the reconstructed models was validated with direct stereoscopic measurements. A computational fluid dynamics simulation was performed to model fluid flow in the generated construct of a Hamburger-Hamilton (HH) stage 27 embryo. Simulation results showed that there were divergent streamlines and a low shear region at the carotid duct, which may be linked to the carotid duct's eventual regression and disappearance by HH stage 34. We show that our technique has sufficient resolution to produce accurate geometries for computational fluid dynamics simulations to quantify embryonic cardiovascular fluid mechanics.

  19. Precision matters for position decoding in the early fly embryo

    NASA Astrophysics Data System (ADS)

    Petkova, Mariela D.; Tkacik, Gasper; Wieschaus, Eric F.; Bialek, William; Gregor, Thomas

    Genetic networks can determine cell fates in multicellular organisms with precision that often reaches the physical limits of the system. However, it is unclear how the organism uses this precision and whether it has biological content. Here we address this question in the developing fly embryo, in which a genetic network of patterning genes reaches 1% precision in positioning cells along the embryo axis. The network consists of three interconnected layers: an input layer of maternal gradients, a processing layer of gap genes, and an output layer of pair-rule genes with seven-striped patterns. From measurements of gap gene protein expression in hundreds of wild-type embryos we construct a ``decoder'', which is a look-up table that determines cellular positions from the concentration means, variances and co-variances. When we apply the decoder to measurements in mutant embryos lacking various combinations of the maternal inputs, we predict quantitative changes in the output layer such as missing, altered or displaced stripes. We confirm these predictions by measuring pair-rule expression in the mutant embryos. Our results thereby show that the precision of the patterning network is biologically meaningful and a necessary feature for decoding cell positions in the early fly embryo.

  20. Layer chicken embryo survival to hatch when administered an in ovo vaccination of strain F Mycoplasma gallisepticum and locations of bacteria prevalence in the newly hatched chick.

    PubMed

    Elliott, K E C; Branton, S L; Evans, J D; Gerard, P D; Peebles, E D

    2017-09-01

    Mycoplasma gallisepticum (MG) is a bacterial pathogen that causes production losses in layer chickens. To combat MG, multiage layer facilities vaccinate pullets by either spray or eye-drop vaccination. The objective in this study was to evaluate the use of in ovo vaccination as a potential alternative for MG vaccination. Layer embryos at 18 d of incubation were either not-injected (control), or were hand-injected with either commercial Marek's disease vaccine diluent alone or with a high, medium, low, or very low dosage of a live attenuated strain F (FMG) vaccine suspended in the commercial diluent. Hatch success and residual egg embryonic mortality were determined after 23 d of incubation. Six hatched chicks per treatment were swabbed for the detection of FMG at 4 different sites (trachea, mouth and esophagus, yolk sac membrane, and the lumen of the duodenal loop) via real-time PCR. Embryos were found to be administered 106 CFU per dose in the high treatment, 104 CFU/dose in the medium treatment, 102 CFU/dose in the low treatment, and between 5.06 and 5.93 CFU/dose in the very low treatment. Hatch of embryonated eggs was decreased by the medium and high doses (P = 0.02). These embryos died while pipping. No FMG was detected in the control and diluent-injected chicks. In the FMG treatments, FMG was found in all sites and dosages, with a greater number of positive chicks found in the higher FMG dosage treatments. These findings indicate the potential practicality of vaccinating layer embryos with FMG by in ovo injection based on the observed hatch success at lower dosages. Also, once injected into the amnion, the bacteria are present in the upper respiratory and gastrointestinal tracts as well the yolk sac membrane and the small intestine of hatchlings. Future research will need to ascertain the effects of FMG administered by in ovo injection on posthatch immunity and mortality. © 2017 Poultry Science Association Inc.

  1. Embryo sac formation and early embryo development in Agave tequilana (Asparagaceae).

    PubMed

    González-Gutiérrez, Alejandra G; Gutiérrez-Mora, Antonia; Rodríguez-Garay, Benjamín

    2014-01-01

    Agave tequilana is an angiosperm species that belongs to the family Asparagaceae (formerly Agavaceae). Even though there is information regarding to some aspects related to the megagametogenesis of A. tequilana, this is the first report describing the complete process of megasporogenesis, megagametogenesis, the early embryo and endosperm development process in detail. The objective of this work was to study and characterize all the above processes and the distinctive morphological changes of the micropylar and chalazal extremes after fertilization in this species. The agave plant material for the present study was collected from commercial plantations in the state of Jalisco, Mexico. Ovules and immature seeds, previously fixed in FAA and kept in ethanol 70%, were stained based on a tissue clarification technique by using a Mayer's-Hematoxylin solution. The tissue clarification technique was successfully used for the characterization of the megasporogenesis, megagametogenesis, mature embryo sac formation, the early embryo and endosperm development processes by studying intact cells. The embryo sac of A. tequilana was confirmed to be of the monosporic Polygonum-type and an helobial endosperm formation. Also, the time-lapse of the developmental processes studied was recorded.

  2. Chromatin Immunoprecipitation in Early Mouse Embryos.

    PubMed

    García-González, Estela G; Roque-Ramirez, Bladimir; Palma-Flores, Carlos; Hernández-Hernández, J Manuel

    2018-01-01

    Epigenetic regulation is achieved at many levels by different factors such as tissue-specific transcription factors, members of the basal transcriptional apparatus, chromatin-binding proteins, and noncoding RNAs. Importantly, chromatin structure dictates the availability of a specific genomic locus for transcriptional activation as well as the efficiency with which transcription can occur. Chromatin immunoprecipitation (ChIP) is a method that allows elucidating gene regulation at the molecular level by assessing if chromatin modifications or proteins are present at a specific locus. Initially, the majority of ChIP experiments were performed on cultured cell lines and more recently this technique has been adapted to a variety of tissues in different model organisms. Using ChIP on mouse embryos, it is possible to document the presence or absence of specific proteins and chromatin modifications at genomic loci in vivo during mammalian development and to get biological meaning from observations made on tissue culture analyses. We describe here a ChIP protocol on freshly isolated mouse embryonic somites for in vivo analysis of muscle specific transcription factor binding on chromatin. This protocol has been easily adapted to other mouse embryonic tissues and has also been successfully scaled up to perform ChIP-Seq.

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

    NASA Astrophysics Data System (ADS)

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

    2013-03-01

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

  4. Exposure of chick embryos to cadmium changes the extra-embryonic vascular branching pattern and alters expression of VEGF-A and VEGF-R2

    SciT

    Gheorghescu, Anna Kaskova; Tywoniuk, Bartlomiej; Complex and Adaptive Systems Laboratory, University College Dublin, Belfield, Dublin 4

    Cadmium (Cd) has several industrial applications, and is found in tobacco products, a notable source of human exposure. Vascular endothelial cells are key targets of Cd toxicity. Here, we aim to quantify the alteration to vascular branching pattern following Cd exposure in the chick extra-embryonic membrane (EEM) using fractal analysis, and explore molecular cues to angiogenesis such as VEGF-A and VEGF-R2 expression following Cd treatment. Chicken embryos were incubated for 60 h to Hamburger–Hamilton developmental stage 16–17, then explanted and treated with 50 μL of 50 μmol cadmium acetate (CdAc) or an equivalent volume of equimolar sodium acetate (NaAc). Imagesmore » of embryos and their area vasculosa (AV) were captured and analyzed at 4 different time points (4, 8, 24 and 48 h) following treatment. Vascular branching in the AV was quantified using its fractal dimension (D{sub f}), estimated using a box counting method. Gallinaceous VEGF ELISA was used to measure the VEGF-A concentration in the EEM following treatment, with determination of the relative expression of VEGF-A and VEGF-R2 using quantitative real-time RT-PCR. Vascular branching increased monotonically in the control group at all time points. The anti-angiogenic effect of Cd exposure on the AV was reflected by a significant reduction in D{sub f} when compared with controls. D{sub f} was more markedly reduced in cultures with abnormal embryos. The expression of VEGF-A protein, and VEGF-A and VEGF-R2 mRNA were reduced in Cd-exposed EEMs. Both molecules contribute to growth, vessel sprouting and branching processes, which supports our findings using fractal analysis. - Highlights: • The chick area vasculosa was undersized in embryos exposed to 50 μM cadmium acetate. • Fractal dimension was reduced in the AV after Cd exposure, indicating decreased branching. • VEGF-A protein was decreased in Cd-treated extraembryonic membranes. • VEGF-A and VEGF-R2 mRNA was decreased in Cd

  5. Early stages of memory formation in filial imprinting: Fos-like immunoreactivity and behavior in the domestic chick.

    PubMed

    Suge, R; McCabe, B J

    2004-01-01

    Early stages of memory formation in filial imprinting were studied in domestic chicks. Chicks trained for 15 min showed strong imprinting, demonstrated by a strong preference for their training stimulus, and the time course of this preference over 2 days after training was similar to that of chicks trained for 60 min. The chicks therefore learned characteristics of the training stimulus very early during training. The intermediate and medial hyperstriatum ventrale (IMHV) is a part of the chick forebrain that is crucial for imprinting. Previous experiments have shown a learning-specific increase in Fos-like immunoreactivity, used as a marker of neuronal activity, in the IMHV after training for 60 min. The time course of Fos expression in the IMHV was measured after training for 15 min and 60 min. The same pattern of expression was found for both training times, showing a peak 120 min after the start of training. The time course of expression was stimulus-dependent. Fos expression in the IMHV, but not the hippocampus, was significantly correlated with strength of imprinting. It is concluded that the learning-specific change in Fos expression in the IMHV is associated with very early components of memory formation.

  6. Egg storage duration and hatch window affect gene expression of nutrient transporters and intestine morphological parameters of early hatched broiler chicks.

    PubMed

    Yalcin, S; Gursel, I; Bilgen, G; Izzetoglu, G T; Horuluoglu, B H; Gucluer, G

    2016-05-01

    In recent years, researchers have given emphasis on the differences in physiological parameters between early and late hatched chicks within a hatch window. Considering the importance of intestine development in newly hatched chicks, however, changes in gene expression of nutrient transporters in the jejunum of early hatched chicks within a hatch window have not been studied yet. This study was conducted to determine the effects of egg storage duration before incubation and hatch window on intestinal development and expression of PepT1 (H+-dependent peptide transporter) and SGLT1 (sodium-glucose co-transporter) genes in the jejunum of early hatched broiler chicks within a 30 h of hatch window. A total of 1218 eggs obtained from 38-week-old Ross 308 broiler breeder flocks were stored for 3 (ES3) or 14 days (ES14) and incubated at the same conditions. Eggs were checked between 475 and 480 h of incubation and 40 chicks from each egg storage duration were weighed; chick length and rectal temperature were measured. The chicks were sampled to evaluate morphological parameters and PepT1 and SGLT1 expression. The remaining chicks that hatched between 475 and 480 h were placed back in the incubator and the same measurements were conducted with those chicks at the end of hatch window at 510 h of incubation. Chick length, chick dry matter content, rectal temperature and weight of small intestine segments increased, whereas chick weight decreased during the hatch window. The increase in the jejunum length and villus width and area during the hatch window were higher for ES3 than ES14 chicks. PepT1 expression was higher for ES3 chicks compared with ES14. There was a 10.2 and 17.6-fold increase in PepT1 and SGLT1 expression of ES3 chicks at the end of hatch window, whereas it was only 2.3 and 3.3-fold, respectively, for ES14 chicks. These results suggested that egg storage duration affected development of early hatched chicks during 30 h of hatch window. It can be concluded that

  7. Effects of male and female sex steroids on the development of normal and the transient Froriep's dorsal root ganglia of the chick embryo.

    PubMed

    Liu, Jiali; Chen, Dawei; Goldstein, Ronald S; Cui, Sheng

    2005-03-22

    Sex steroids can influence developmental processes and support the survival of neurons in the embryonic central nervous system. Recent studies have shown that estrogen receptors are also expressed in the peripheral nervous system, in the dorsal root ganglia (DRG) of chick embryos. However, no studies have examined the effects of sex steroids on development of embryonic DRG. In the present study, 0.2 microg, 1.0 microg, 5.0 microg 10 microg, 20 microg, 25 microg, and 40 microg doses of testosterone or estradiol were delivered to chick embryos at Hamburger and Hamilton stage 18 (E3). The actions of these doses of sex steroids on the development of the C5DRG (fifth cervical ganglion, a "normal" DRG) and C2DRG (a transient ganglion known as a "Froriep's DRG") were then evaluated by quantifying ganglionic volumes, cell number, proliferation, and apoptosis after 1 day of growth to stage 23. We found that both testosterone and estradiol promoted proliferation of cells in both normal DRG and the Froriep's ganglia. By contrast, estradiol significantly increased the number of apoptotic cells, while testosterone strongly inhibited apoptosis. These actions of sex steroids on DRG development were dose-dependent, and C5DRG and C2DRG showed different sensitivities to the applied sex steroids. In addition, the present results demonstrated that specific ER and AR inhibitors (tamoxifen and flutamide) did not influence the effects of 5 microg E2 and 5 microg T on C2 and C5DRG significantly. These results demonstrate that male and female sex steroids can modulate DRG development through an epigenetic mechanism, as had been shown for the central nervous system.

  8. Embryotoxic effects of crude oil in mallard ducks and chicks

    Hoffman, David J.

    1978-01-01

    Recent studies in this laboratory have revealed that surface applications of microliter amounts of some crude and fuel oils that coat less than 10% of the egg surface reduce hatching considerably in different avian species. Applications of paraffin compounds that coat equal areas of the egg surface do not reduce hatching suggesting that toxicity is due to causes other than asphyxia. In the present study, 1–10 μl of South Louisiana crude oil, an API reference oil, were applied to the surface of fertile mallard (Anas platyrhynchos) and chicken (Gallus gallus) eggs. Early embryolethality was greater in mallard embryos than in chick embryos, but later embryolethality that coincided with the time of rapid outgrowth of the chorioallantoic membrane was more prevalent in chick embryos. The overall incidence of embryolethality was similar in both species. Retardation of growth as reflected by embryonic body weight, crown-rump length, beak length, and general appearance was more pronounced in chick than mallard embryos. Teratogenic defects were more frequent in chick embryos, and incomplete or abnormal ossification of the skull was the most common. External application of equivalent amounts of a mixture of paraffin compounds present in crude oil had virtually no embryotoxic effects in either species, suggesting that other components including aromatic hydrocarbons and organometallics may cause the embryotoxicity.

  9. Effect of zinc imprinting and replacing inorganic zinc with organic zinc on early performance of broiler chicks

    PubMed Central

    Timmons, J.; Ao, T.; Paul, M.; Macalintal, L.; Pescatore, A.; Cantor, A.; Ford, M.; Dawson, K. A.

    2017-01-01

    Abstract The goal of this study was to determine the effects of feeding a zinc (Zn) deficient diet to broiler chicks for 96 h post-hatch followed by feeding diets with different Zn sources and supplemental levels (5 to 21 d) on the growth performance, tissue, and excreta Zn content. At the start of the study, four hundred 20-day-old male broiler chicks were divided into two groups. One group was fed a corn soybean meal based diet containing 25 mg of Zn/kg (imprinting diet, ID). The second group was fed the basal diet supplemented with 40 mg of Zn/kg from Zn oxide (ZnO) (non-imprinting diet, NID). Both groups were fed these diets for 96 h. At d 5, chicks from each group were randomly assigned to the dietary treatments consisting of the basal diet alone or the basal diet supplemented with 8 or 40 mg/kg Zn as ZnO or Zn proteinate. Main effects of post-hatch Zn ID were observed on feed intake and G:F. ID decreased (P < 0.05) feed intake and improved (P < 0.05) the gain to feed ratio (G:F) of 14 and 21 d old chicks compared to G:F of chicks fed NID. Additionally, G:F for 14 and 21 d was improved (P < 0.05) by interaction of Zn source × level. Furthermore, at d 21 chicks fed the ID had a lower (P < 0.05) Zn content in the tibia ash and excreta, and a higher (P < 0.05) Zn content in the pancreas tissue compared to chicks fed NID. These results suggest that Zn imprinting can affect body Zn stores and early performance. PMID:27664197

  10. Effect of zinc imprinting and replacing inorganic zinc with organic zinc on early performance of broiler chicks.

    PubMed

    Mwangi, S; Timmons, J; Ao, T; Paul, M; Macalintal, L; Pescatore, A; Cantor, A; Ford, M; Dawson, K A

    2017-04-01

    The goal of this study was to determine the effects of feeding a zinc (Zn) deficient diet to broiler chicks for 96 h post-hatch followed by feeding diets with different Zn sources and supplemental levels (5 to 21 d) on the growth performance, tissue, and excreta Zn content. At the start of the study, four hundred 20-day-old male broiler chicks were divided into two groups. One group was fed a corn soybean meal based diet containing 25 mg of Zn/kg (imprinting diet, ID). The second group was fed the basal diet supplemented with 40 mg of Zn/kg from Zn oxide (ZnO) (non-imprinting diet, NID). Both groups were fed these diets for 96 h. At d 5, chicks from each group were randomly assigned to the dietary treatments consisting of the basal diet alone or the basal diet supplemented with 8 or 40 mg/kg Zn as ZnO or Zn proteinate. Main effects of post-hatch Zn ID were observed on feed intake and G:F. ID decreased (P < 0.05) feed intake and improved (P < 0.05) the gain to feed ratio (G:F) of 14 and 21 d old chicks compared to G:F of chicks fed NID. Additionally, G:F for 14 and 21 d was improved (P < 0.05) by interaction of Zn source × level. Furthermore, at d 21 chicks fed the ID had a lower (P < 0.05) Zn content in the tibia ash and excreta, and a higher (P < 0.05) Zn content in the pancreas tissue compared to chicks fed NID. These results suggest that Zn imprinting can affect body Zn stores and early performance. © The Author 2016. Published by Oxford University Press on behalf of Poultry Science Association.

  11. Contraction and stress-dependent growth shape the forebrain of the early chicken embryo.

    PubMed

    Garcia, Kara E; Okamoto, Ruth J; Bayly, Philip V; Taber, Larry A

    2017-01-01

    During early vertebrate development, local constrictions, or sulci, form to divide the forebrain into the diencephalon, telencephalon, and optic vesicles. These partitions are maintained and exaggerated as the brain tube inflates, grows, and bends. Combining quantitative experiments on chick embryos with computational modeling, we investigated the biophysical mechanisms that drive these changes in brain shape. Chemical perturbations of contractility indicated that actomyosin contraction plays a major role in the creation of initial constrictions (Hamburger-Hamilton stages HH11-12), and fluorescent staining revealed that F-actin is circumferentially aligned at all constrictions. A finite element model based on these findings shows that the observed shape changes are consistent with circumferential contraction in these regions. To explain why sulci continue to deepen as the forebrain expands (HH12-20), we speculate that growth depends on wall stress. This idea was examined by including stress-dependent growth in a model with cerebrospinal fluid pressure and bending (cephalic flexure). The results given by the model agree with observed morphological changes that occur in the brain tube under normal and reduced eCSF pressure, quantitative measurements of relative sulcal depth versus time, and previously published patterns of cell proliferation. Taken together, our results support a biphasic mechanism for forebrain morphogenesis consisting of differential contractility (early) and stress-dependent growth (late). Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. The chronic infusion of nicotine into the developing chick embryo does not alter the density of (-)-[3H]nicotine-binding sites or vestibular function

    NASA Technical Reports Server (NTRS)

    Roll, R. L.; Jones, T. A.; Benowitz, N. L.; Morley, B. J.

    1993-01-01

    (-)-Nicotine (1.2 mg/day) or saline was infused into chick embryos (Gallus domesticus) for 10 days beginning 12 h beyond the eight day of incubation (E8 + 12 h). Twelve h beyond the eighteenth day of incubation (E18 + 12 h), the eggs were opened to access the embryos and subcutaneous skull electrodes placed. Short latency vestibular response thresholds and input/output functions were determined to assess neurophysiological consequences of chronic nicotine administration. Samples of serum and extraembryonic (amniotic and albumen) fluid were analyzed by gas chromatography-mass spectrometry to determine the levels of nicotine and its major metabolite, cotinine. The brains were removed and divided into diencephalon and mesencephalon and the density of (-)-[3H]nicotine binding sites in each brain area was measured. Nicotine and cotinine were found in the serum and extraembryonic fluid, but nicotinic receptors were not up-regulated in the brains of animals infused with nicotine in comparison to controls. Vestibular response thresholds also did not differ between nicotine-treated and control animals.

  13. [Specification of cell destiny in early Caenorhabditis elegans embryo].

    PubMed

    Schierenberg, E

    1997-02-01

    Embryogenesis of the nematode Caenorhabditis elegans has been described completely on a cell-by-cell basis and found to be essentially invariant. With this knowledge in hands, micromanipulated embryos and mutants have been analyzed for cell lineage defects and the distribution of specific gene products. The results challenge the classical view of cell-autonomous development in nematodes and indicate that the early embryo of C. elegans is a highly dynamic system. A network of inductive events between neighboring cells is being revealed, which is necessary to assign different developmental programs to blastomeres. In those cases where molecules involved in these cell-cell interactions have been identified, homologies to cell surface receptors, ligands and transcription factors found in other systems have become obvious.

  14. Embryo density and medium volume effects on early murine embryo development.

    PubMed

    Canseco, R S; Sparks, A E; Pearson, R E; Gwazdauskas, F C

    1992-10-01

    One-cell mouse embryos were used to determine the effects of drop size and number of embryos per drop for optimum development in vitro. Embryos were collected from immature C57BL6 female mice superovulated with pregnant mare serum gonadotropin and human chorionic gonadotropin and mated by CD1 males. Groups of 1, 5, 10, or 20 embryos were cultured in 5-, 10-, 20-, or 40-microliters drops of CZB under silicon oil at 37.5 degrees C in a humidified atmosphere of 5% CO2 and 95% air. Development score for embryos cultured in 10 microliters was higher than that of embryos cultured in 20 or 40 microliters. Embryos cultured in groups of 5, 10, or 20 had higher development scores than embryos cultured singly. The highest development score was obtained by the combination of 5 embryos per 10-microliters drop. The percentage of live embryos in 20 or 40 microliters was lower than that of embryos cultured in 10 microliters. Additionally, the percentage of live embryos cultured singly was lower than that of embryos cultured in groups. Our results suggest that a stimulatory interaction occurs among embryos possibly exerted through the secretion of growth factors. This effect can be diluted if the embryos are cultured in large drops or singly.

  15. Expression profiling of the mouse early embryo: Reflections and Perspectives

    PubMed Central

    Ko, Minoru S. H.

    2008-01-01

    Laboratory mouse plays important role in our understanding of early mammalian development and provides invaluable model for human early embryos, which are difficult to study for ethical and technical reasons. Comprehensive collection of cDNA clones, their sequences, and complete genome sequence information, which have been accumulated over last two decades, have provided even more advantages to mouse models. Here the progress in global gene expression profiling in early mouse embryos and, to some extent, stem cells are reviewed and the future directions and challenges are discussed. The discussions include the restatement of global gene expression profiles as snapshot of cellular status, and subsequent distinction between the differentiation state and physiological state of the cells. The discussions then extend to the biological problems that can be addressed only through global expression profiling, which include: bird’s-eye view of global gene expression changes, molecular index for developmental potency, cell lineage trajectory, microarray-guided cell manipulation, and the possibility of delineating gene regulatory cascades and networks. PMID:16739220

  16. PITX2 and NODAL expression during axis formation in the early rabbit embryo.

    PubMed

    Plöger, Ruben; Viebahn, Christoph

    2018-04-26

    Attaining molecular and morphological axial polarity during gastrulation is a fundamental early requirement for normal development of the embryo. In mammals, the first morphological sign of the anterior-posterior axis appears anteriorly in the form of the anterior marginal crescent (or anterior visceral endoderm) while in the avian the first such sign is the Koller's sickle at the posterior pole of the embryonic disc. Despite this inverse mode of axis formation many genes and molecular pathways involved in various steps of this process seem to be evolutionary conserved amongst amniotes, the nodal gene being a well-known example with its functional involvement prior and during gastrulation. The pitx2 gene, however, is a new candidate described in the chick as an early marker for anterior-posterior polarity and as regulator of axis formation including twinning. To find out whether pitx2 has retained its inductive and early marker function during the evolution of mammals, this study analyzes pitx2 and nodal expression at parallel stages during formation of the anterior-posterior polarity in the early rabbit embryo using whole-mount in situ hybridization and serial light-microscopical sections. At a late pre-gastrulation stage a localized reduction of nodal expression presages the position of the anterior pole of the embryonic disc and thus serves as the earliest molecular marker of anterior-posterior polarity known so far. pitx2 is expressed in a polarized manner in the anterior marginal crescent and in the posterior half of the embryonic disc during further development only while nodal expression in the anterior segment of the posterior pitx2 expression domain helps to define the so-called anterior streak domain (ASD), a novel progenitor region of the anterior half of the primitive streak. The expression patterns of both genes thus serve as signs of a conserved involvement in early axis formation in amniotes and, possibly, in twinning in mammals as well. Copyright

  17. Efficient harvesting methods for early-stage snake and turtle embryos.

    PubMed

    Matsubara, Yoshiyuki; Kuroiwa, Atsushi; Suzuki, Takayuki

    2016-04-01

    Reptile development is an intriguing research target for understating the unique morphogenesis of reptiles as well as the evolution of vertebrates. However, there are numerous difficulties associated with studying development in reptiles. The number of available reptile eggs is usually quite limited. In addition, the reptile embryo is tightly adhered to the eggshell, making it a challenge to isolate reptile embryos intact. Furthermore, there have been few reports describing efficient procedures for isolating intact embryos especially prior to pharyngula stage. Thus, the aim of this review is to present efficient procedures for obtaining early-stage reptilian embryos intact. We first describe the method for isolating early-stage embryos of the Japanese striped snake. This is the first detailed method for obtaining embryos prior to oviposition in oviparous snake species. Second, we describe an efficient strategy for isolating early-stage embryos of the soft-shelled turtle. © 2016 Japanese Society of Developmental Biologists.

  18. The effect of flurbiprofen on the development of anencephaly in early stage chicken embryos.

    PubMed

    Özeren, Ersin; Er, Uygur; Güvenç, Yahya; Demirci, Adnan; Arıkök, Ata Türker; Şenveli, Engin; Ergün, Rüçhan Behzat

    2015-04-01

    The study investigated the effect of flurbiprofen on the development of anencephaly in early stage chicken embryos. We looked at four groups with a total of 36 embryos. There was a control group, a normal saline group, a normal-dose group and a high-dose group with ten, ten, eight and eight eggs with embryo respectively. Two embryos in the control group, studied with light microscopy at 48 h, were consistent with 28-29 hours' incubation in the Hamburger-Hamilton System. They had open neural tubes. The other embryos in this group were considered normal. One embryo in the normal saline group was on the occlusion stage at 48 h. One embryo showed an open neural tube. They were compatible with 28-29 hours' incubation in the Hamburger-Hamilton system. The remaining eight embryos showed normal development. In the normal dose group, one embryo showed underdevelopment of the embryonic disc and the embryo was dead. In four embryos, the neural tubes were open. One cranial malformation was found that was complicated with anencephaly in one embryo. In two embryos the neural tubes were closed, as they showed normal development, and they reached their expected stages according to the Hamburger-Hamilton classification. There was no malformation or growth retardation. Four experimental embryos were anencephalic in the high dose group, and three embryos had open neural tubes. One embryo exhibited both anencephaly and a neural tube closure defect. None of the embryos in this group showed normal development. Even the usual therapeutic doses of flurbiprofen increased the risk of neural tube defect. Flurbiprofen was found to significantly increase the risk of anencephaly. The provision of improved technical materials and studies with larger sample sizes will reveal the stage of morphological disruption during the development of embryos.

  19. [Studies of the eating behavior of Japanese quail chicks in the early postnatal period].

    PubMed

    Khekhneva, A V; Gur'eva, T S; Dadasheva, O A; Sychev, V N

    2006-01-01

    Time of the eating reaction (response to video signals) and eating behavior shortly after hatching were studied in Japanese quail chicks whose embryonic development took place under normal or changed gravity. Chicks partially incubated in a changed gravity showed a much slower eating reaction when compared with the chicks the prenatal development of which occurred under the normal gravity. In the chicks incubated at 1 g and placed in individual cages immediately after dominating afferentation for the eating behavior was visual Observations in the study will be used as a basis for designing a technology for handling and maintenance of hatchlings of the Japanese quail as a potential heterotrophic component of space life support systems.

  20. Antibodies induced by vaccination with purified chick embryo cell culture vaccine (PCECV) cross-neutralize non-classical bat lyssavirus strains.

    PubMed

    Malerczyk, Claudius; Selhorst, Thomas; Tordo, Noël; Moore, Susan; Müller, Thomas

    2009-08-27

    Tissue-culture vaccines like purified chick embryo cell vaccine (PCECV) have been shown to provide protection against classical rabies virus (RABV) via pre-exposure or post-exposure prophylaxis. A cross-neutralization study was conducted using a panel of 100 human sera, to determine, to what extent after vaccination with PCECV protection exists against non-classical bat lyssavirus strains like European bat lyssavirus (EBLV) type 1 and 2 and Australian bat lyssavirus (ABLV). Virus neutralizing antibody (VNA) concentrations against the rabies virus variants CVS-11, ABLV, EBLV-1 and EBLV-2 were determined by using a modified rapid fluorescent focus inhibition test. For ABLV and EBLV-2, the comparison to CVS-11 revealed almost identical results (100% adequate VNA concentrations >or=0.5 IU/mL; correlation coefficient r(2)=0.69 and 0.77, respectively), while for EBLV-1 more scattering was observed (97% adequate VNA concentrations; r(2)=0.50). In conclusion, vaccination with PCECV produces adequate VNA concentrations against classical RABV as well as non-classical lyssavirus strains ABLV, EBLV-1, and EBLV-2.

  1. 3D chick embryo chorioallantoic membrane model as an in vivo model to study morphological and histopathological features of feline fibrosarcomas.

    PubMed

    Zabielska-Koczywąs, Katarzyna; Wojtkowska, Agata; Dolka, Izabella; Małek, Anna; Walewska, Magdalena; Wojtalewicz, Anna; Żbikowski, Artur; Lechowski, Roman

    2017-06-26

    The chick embryo chorioallantoic membrane (CAM) model is well described in human medicine as a cost-effective, easy to perform preclinical oncological model for observing pro- and antiangiogenic response, tumor biology and metastasis. The main objective of this article was to present the modification of the CAM assay in order to evaluate tumor growth from two feline fibrosarcoma cell lines (FFS1, FFS3) and describe their morphological and histopathological features. The authors described morphological and histopathological features of two feline fibrosarcoma cell lines (FFS1 and FFS3) grown on the CAM. Tumors from the FFS1 cell line showed high malignancy (grade III), while tumors from the FFS3 cell line were grade II. Proliferation markers (Ki-67 and PCNA) were determined and the positive correlation between PCNA and tumor grade (r = 0.8247; p < 0.001) was demonstrated, as opposed to Ki-67. The results obtained indicate that PCNA may be helpful to evaluate the tumor grade, better than Ki-67, for feline fibrosarcomas. However, further investigations of proliferation marker, in bigger number of feline spontaneous fibrosarcomas and feline fibrosarcomas grown on the CAM from different cell lines, are needed to confirm these observations.

  2. Derivation of feline vaccine-associated fibrosarcoma cell line and its growth on chick embryo chorioallantoic membrane - a new in vivo model for veterinary oncological studies.

    PubMed

    Zabielska, K; Lechowski, R; Król, M; Pawłowski, K M; Motyl, T; Dolka, I; Zbikowski, A

    2012-12-01

    Feline vaccine associated fibrosarcomas are the second most common skin tumor in cats. Methods of treatment are: surgery, chemotherapy and radiotherapy. Nevertheless, the usage of cytostatics in feline vaccine associated sarcoma therapy is limited due to their adverse side effects, high toxicity and low biodistribution after i.v. injection. Therefore, much research on new therapeutic drugs is being conducted. In human medicine, the chick embryo chorioallantoic membrane (CAM) model is used as a cheap and easy to perform assay to assess new drug effectiveness in cancer treatment. Various human cell lines have different tumors growth on CAM. In veterinary medicine such model has not been described yet. In the present article derivation of feline vaccine associated fibrosarcoma cell line and its growth on CAM is described. The cell line and the tumor grown were confirmed by histopathological and immunohistochemical examination. As far as we believe, this is the first attempt to create such model, which may be used for further in vivo studies in veterinary oncology.

  3. Characterization of a photoproduct of 7,12-dimethylbenz[alpha]anthracene and its effects on chick-embryo cells in culture.

    PubMed Central

    Warshawsky, D; Kerns, E; Bissell, M J; Calvin, M

    1977-01-01

    A common impurity of 7,12-dimethylbenz[alpha]anthracene was more effective than 7,12-dimethylbenz[alpha]anthracene in inducing morphological alterations, and in causing an increase in glucose uptake, DNA synthesis and cell number in chick-embryo fibroblasts. Gradual morphological transformation follows the increase in DNA synthesis after 2 days when either primary or secondary cultures are treated with 3 microgram of the compound/ml. The compound, isolated from 7,12-dimethylbenz[alpha]anthracene by alumina column chromatography, was characterized by t.l.c., mass spectroscopy, carbon-hydrogen analysis, u.v. and nuclear-magnetic-resonance spectroscopy and thermal decomposition. It was the photo-oxidation product of 7,12-dimethylbenz[alpha]anthracene, 7,12-epidioxy-7,12-dimethylbenz[alpha]anthracene. It is suggested that some of the biological effects observed after treatment of cultures with 7,12-dimethylbenz[alpha]anthracene may be due in part to the presence of the photo-oxidation product. PMID:407902

  4. The role of Mixer in patterning the early Xenopus embryo.

    PubMed

    Kofron, Matt; Wylie, Chris; Heasman, Janet

    2004-05-01

    The transcription factor VegT, is required in early Xenopus embryos for the formation of both the mesoderm and endoderm germ layers. Inherited as a maternal mRNA localized only in vegetal cells, VegT activates the transcription of a large number of transcription factors, as well as signaling ligands that induce cells in the vegetal mass to form endoderm, and the marginal zone to form mesoderm. It is important now to understand the extent to which transcription factors downstream of VegT play individual, or overlapping, roles in the specification and patterning of the endoderm and mesoderm. In addition, it is important to understand the mechanism that specifies the boundary between endoderm and mesoderm. One of the downstream targets of VegT, the homeodomain protein Mixer, is expressed at high levels at the mesoderm/endoderm boundary at the late blastula stage. We therefore examined its functions by blocking its translation using morpholino oligos. In Mixer-depleted embryos, the expression of many signaling ligands and transcription factors was affected. In particular, we found that the expression of several genes, including several normally expressed in mesoderm, was upregulated. Functional assays of Mixer-depleted vegetal cells showed that they have increased mesoderm-inducing activity. This demonstrates that Mixer plays an essential role in controlling the amount of mesoderm induction by the vegetal cells.

  5. Detailed analysis of the δ-crystallin mRNA-expressing region in early development of the chick pituitary gland.

    PubMed

    Inoue, Makiko; Shiina, Tomoya; Aizawa, Sayaka; Sakata, Ichiro; Takagi, Hiroyasu; Sakai, Takafumi

    2012-06-01

    Although δ-crystallin (δ-crys), also known as lens protein, is transiently expressed in Rathke's pouch (RP) of the chick embryo, detailed temporal and spatial expression patterns have been obscure. In this study, to understand the relationship between the δ-crys mRNA-expressing region and RP formation, we examined the embryonic expression pattern of δ-crys mRNA in the primordium of the adenohypophysis. δ-crys mRNA expression was initially found at stage 15 anterior to the foregut and posterior to the invaginated oral ectoderm. After RP formation, the δ-crys mRNA was expressed in the post-ventral region of RP and the anterior region of RP. δ-crys mRNA expression was then restricted to the cephalic lobe of the pituitary gland. From stage 20, the δ-crys and alpha-glycoprotein subunit (αGSU) mRNA-expressing regions were almost completely overlapping. The αGSU mRNA-expressing region is thought to be the primordium of the pars tuberalis, and these regions were overlapped with the Lhx3 mRNA-expressing region. The intensity of δ-crys mRNA expression gradually decreased with development and completely disappeared by stage 34. These results suggest that the embryonic chick pituitary gland consists of two different regions labeled with δ-crys and Lhx3.

  6. 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.

  7. Early Cambrian Pentamerous Cubozoan Embryos from South China

    PubMed Central

    Han, Jian; Kubota, Shin; Li, Guoxiang; Yao, Xiaoyong; Yang, Xiaoguang; Shu, Degan; Li, Yong; Kinoshita, Shunichi; Sasaki, Osamu; Komiya, Tsuyoshi; Yan, Gang

    2013-01-01

    Background Extant cubozoans are voracious predators characterized by their square shape, four evenly spaced outstretched tentacles and well-developed eyes. A few cubozoan fossils are known from the Middle Cambrian Marjum Formation of Utah and the well-known Carboniferous Mazon Creek Formation of Illinois. Undisputed cubozoan fossils were previously unknown from the early Cambrian; by that time probably all representatives of the living marine phyla, especially those of basal animals, should have evolved. Methods Microscopic fossils were recovered from a phosphatic limestone in the Lower Cambrian Kuanchuanpu Formation of South China using traditional acetic-acid maceration. Seven of the pre-hatched pentamerous cubozoan embryos, each of which bears five pairs of subumbrellar tentacle buds, were analyzed in detail through computed microtomography (Micro-CT) and scanning electron microscopy (SEM) without coating. Results The figured microscopic fossils are unequivocal pre-hatching embryos based on their spherical fertilization envelope and the enclosed soft-tissue that has preserved key anatomical features arranged in perfect pentaradial symmetry, allowing detailed comparison with modern cnidarians, especially medusozoans. A combination of features, such as the claustrum, gonad-lamella, suspensorium and velarium suspended by the frenula, occur exclusively in the gastrovascular system of extant cubozoans, indicating a cubozoan affinity for these fossils. Additionally, the interior anatomy of these embryonic cubozoan fossils unprecedentedly exhibits the development of many new septum-derived lamellae and well-partitioned gastric pockets unknown in living cubozoans, implying that ancestral cubozoans had already evolved highly specialized structures displaying unexpected complexity at the dawn of the Cambrian. The well-developed endodermic lamellae and gastric pockets developed in the late embryonic stages of these cubozoan fossils are comparable with extant pelagic

  8. A multicenter prospective study to assess the effect of early cleavage on embryo quality, implantation, and live-birth rate.

    PubMed

    de los Santos, Maria José; Arroyo, Gemma; Busquet, Ana; Calderón, Gloria; Cuadros, Jorge; Hurtado de Mendoza, Maria Victoria; Moragas, Marta; Herrer, Raquel; Ortiz, Agueda; Pons, Carme; Ten, Jorge; Vilches, Miguel Angel; Figueroa, Maria José

    2014-04-01

    To investigate the impact of early cleavage (EC) on embryo quality, implantation, and live-birth rates. Prospective cross-sectional study. Multicenter study. Seven hundred embryo transfers and 1,028 early-stage human embryos. None. Implantation according to the presence of EC and embryo quality. The presence of EC is associated with embryo quality, especially in cycles with autologous oocytes. However, the use of EC as an additional criterion for selecting an embryo for transfer does not appear to significantly improve likelihood of implantation. Furthermore, embryos that presented EC had live-birth rates per implanted embryo similar to those that did not show any sign of cleavage. At least for conventional embryo culture and morphologic evaluations, the additional evaluation of EC in embryos may not be valuable to improve embryo implantation. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  9. 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…

  10. The effects of early post-hatching changes of imprinting object on the pattern of monocular/unihemispheric sleep of domestic chicks.

    PubMed

    Bobbo, Daniela; Vallortigara, Giorgio; Mascetti, Gian Gastone

    2006-06-03

    The pattern of monocular/unihemispheric sleep (Mo-Un sleep) was studied behaviourally in male and female chicks after early post-hatching changes of the imprinting object. Chicks were reared with an imprinting object on day 1 post-hatching which was removed or changed on day 2. On day 1, time spent in binocular sleep (both eyes closed) was similar in male and female chicks, though the number of episodes was lower in females than in males. There was no eye-closure bias in the pattern of Mo-Un sleep (one eye shut and the other open) in chicks of both sexes. On day 2, chicks subjected to the removal of imprinting object showed less time and number of episodes of binocular sleep than control chicks and chicks subjected to changes of imprinting object. There was no eye-closure bias in control chicks whilst a significant bias for more right Mo-Un sleep was recorded in chicks after removal and changes of imprinting object of both sexes. It is suggested that the removal or changes of imprinting object would cause a decrease of binocular sleep and trigger processes associated to secondary imprinting involving the left hemisphere. The bias for more right Mo-Un sleep (right eye-closure) could be the by-product of consolidation processes of secondary imprinting memories in the left hemisphere and/or of more left eye-opening as a result of periodical awakening of right hemisphere to control the environment after a stressful condition such as the removal or change of imprinting object.

  11. Early embryo development in Fucus distichus is auxin sensitive

    NASA Technical Reports Server (NTRS)

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

    2002-01-01

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

  12. Early Embryo Development in Fucus distichus Is Auxin Sensitive1

    PubMed Central

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

    2002-01-01

    Auxin and polar auxin transport have been implicated in controlling embryo development in land plants. The goal of these studies was to determine if auxin and auxin transport are also important during the earliest stages of development in embryos of the brown alga Fucus distichus. Indole-3-acetic acid (IAA) was identified in F. distichus embryos and mature tissues by gas chromatography-mass spectroscopy. F. distichus embryos accumulate [3H]IAA and an inhibitor of IAA efflux, naphthylphthalamic acid (NPA), elevates IAA accumulation, suggesting the presence of an auxin efflux protein complex similar to that found in land plants. F. distichus embryos normally develop with a single unbranched rhizoid, but growth on IAA leads to formation of multiple rhizoids and growth on NPA leads to formation of embryos with branched rhizoids, at concentrations that are active in auxin accumulation assays. The effects of IAA and NPA are complete before 6 h after fertilization (AF), which is before rhizoid germination and cell division. The maximal effects of IAA and NPA are between 3.5 and 5 h AF and 4 and 5.5 h AF, respectively. Although, the location of the planes of cell division was significantly altered in NPA- and IAA-treated embryos, these abnormal divisions occurred after abnormal rhizoid initiation and branching was observed. The results of this study suggest that auxin acts in the formation of apical basal patterns in F. distichus embryo development. PMID:12226509

  13. Physiological response of broiler embryos to different incubator temperature profiles and maternal flock age during incubation. 1. Embryonic metabolism and day-old chick quality.

    PubMed

    Hamidu, J A; Torres, C A; Johnson-Dahl, M L; Korver, D R

    2018-05-18

    Broiler strain, maternal age, and incubation temperature influence embryo metabolism. Hatching eggs were obtained from young (Y; 28 to 34 wk, $\\bar{\\rm x}$ = 31.2 wk), mid (M; 36 to 45 wk, $\\bar{\\rm x}$ = 40.5 wk) and old (O; 49 to 54 wk, $\\bar{\\rm x}$ = 51.4 wk) Ross 708 (n = 88; Experiment 1) and Ross 308 [(n = 45; Experiment 2: (Y; 25 to 34 wk, $\\bar{\\rm x}$ = 30.5 wk), (M; 35 to 44 wk, $\\bar{\\rm x}$ = 40.2 wk), and (O; 49 to 54 wk, $\\bar{\\rm x}$ = 51.6 wk)] breeders. Eggs were stored for 2 to 4 d (18°C, 73% RH), and incubated for 14 d at 37.5°C and 56% RH. At 15 d (E15), 8 fertile eggs per flock age were incubated in individual metabolic chambers at 36.0, 36.5, 37.0, or 37.5°C until E21.5. Each temperature was repeated one additional time. O2 consumption and CO2 production were used to calculate embryonic heat production (EHP). Embryo temperature was measured as eggshell temperature (EST). Initial egg weight was used as a covariate; significance was assessed at P < 0.05. In Ross 708, daily EHP tended to be higher in M and O than Y treatments at E16; EHP of M was higher than Y and O eggs at E18; M and O were higher than O eggs at E19. Incubation at 37.0°C resulted in the highest EHP from E15 to E21, except at E17. Embryos at 37.5°C had reduced EHP beyond E17. Daily EST from E15 to E21 was higher at 37.5 and 37.0°C than at 36.0 and 36.5°C. In Ross 308, daily EST was highest at 37.5°C except at E20. Incubation temperature and EST were highly correlated (R2 = 0.90 to 0.89; P < 0.001). Ross 708 chicks were longer and hatched earlier at 37.0°C than at 36.0 and 37.5°C. EST and EHP increased with incubation temperature in Ross 708. In Ross 308, maternal flock age and incubation temperature did not impact EHP. However, EST was highest at 37.5°C except at E20. Ross 708 was more sensitive to incubation temperature than Ross 308.

  14. Radiation induced abnormalities in early in vitro mouse embryos

    SciT

    Kirkpatrick, J.F.

    1973-08-01

    Female mice were superovulated and mated, and the two-cell embryos were collected and cultured in vitro. The embryos were exposed to x-irradiation (0 to 491 rads) during the two-cell stage before the appearance of the next cleavage plate, placed in new unirradiated culture medium and observed during subsequent development. Morphological abnormalities, which occurred as a result of irradiation, included fragmentation, disintegration, granlation, incomplete cleavage, cleavage cessation, nuclear degeneration and pycnosis and cytoplasmic vacuolization. There was no damage to the zona pellucida. The types of abnormalities indicate an agreement with the results of previous in vivo studies. A distinct correlation existedmore » between morphological abnormalities and embryo death. The greatest number of abnormalities resulted within five hours following irradiation, but increased through 20 hours post-exposure. At doses above 300 rads, the magnitude of damage was greater in the in vitro embryos than that shown in previous in vivo studies. (auth)« less

  15. Expression of renin–angiotensin system components in the early bovine embryo

    PubMed Central

    Pijacka, Wioletta; Hunter, Morag G; Broughton Pipkin, Fiona; Luck, Martin R

    2012-01-01

    The renin–angiotensin system (RAS), mainly associated with the regulation of blood pressure, has been recently investigated in female reproductive organs and the developing foetus. Angiotensin II (Ang II) influences oviductal gamete movements and foetal development, but there is no information about RAS in the early embryo. The aim of this study was to determine whether RAS components are present in the pre-implantation embryo, to determine how early they are expressed and to investigate their putative role at this stage of development. Bovine embryos produced in vitro were used for analysis of RAS transcripts (RT-PCR) and localisation of the receptors AGTR1 and AGTR2 (immunofluorescent labelling). We also investigated the effects of Ang II, Olmesartan (AGTR1 antagonist) and PD123319 (AGTR2 antagonist) on oocyte cleavage, embryo expansion and hatching. Pre-implanted embryos possessed AGTR1 and AGTR2 but not the other RAS components. Both receptors were present in the trophectoderm and in the inner cell mass of the blastocyst. AGTR1 was mainly localised in granular-like structures in the cytoplasm, suggesting its internalisation into clathrin-coated vesicles, and AGTR2 was found mainly in the nuclear membrane and in the mitotic spindle of dividing trophoblastic cells. Treating embryos with PD123319 increased the proportion of hatched embryos compared with the control. These results, the first on RAS in the early embryo, suggest that the pre-implanted embryo responds to Ang II from the mother rather than from the embryo itself. This may be a route by which the maternal RAS influences blastocyst hatching and early embryonic development. PMID:23781300

  16. Tlx-1 and Tlx-3 homeobox gene expression in cranial sensory ganglia and hindbrain of the chick embryo: markers of patterned connectivity.

    PubMed

    Logan, C; Wingate, R J; McKay, I J; Lumsden, A

    1998-07-15

    Recent evidence suggests that in vertebrates the formation of distinct neuronal cell types is controlled by specific families of homeodomain transcription factors. Furthermore, the expression domains of a number of these genes correlates with functionally integrated neuronal populations. We have isolated two members of the divergent T-cell leukemia translocation (HOX11/Tlx) homeobox gene family from chick, Tlx-1 and Tlx-3, and show that they are expressed in differentiating neurons of both the peripheral and central nervous systems. In the peripheral nervous system, Tlx-1 and Tlx-3 are expressed in overlapping domains within the placodally derived components of a number of cranial sensory ganglia. Tlx-3, unlike Tlx-1, is also expressed in neural crest-derived dorsal root and sympathetic ganglia. In the CNS, both genes are expressed in longitudinal columns of neurons at specific dorsoventral levels of the hindbrain. Each column has distinct anterior and/or posterior limits that respect inter-rhombomeric boundaries. Tlx-3 is also expressed in D2 and D3 neurons of the spinal cord. Tlx-1 and Tlx-3 expression patterns within the peripheral and central nervous systems suggest that Tlx proteins may be involved not only in the differentiation and/or survival of specific neuronal populations but also in the establishment of neuronal circuitry. Furthermore, by analogy with the LIM genes, Tlx family members potentially define sensory columns early within the developing hindbrain in a combinatorial manner.

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

    PubMed Central

    2011-01-01

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

  18. Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds

    PubMed Central

    Raissig, Michael T.; Gagliardini, Valeria; Jaenisch, Johan; Grossniklaus, Ueli; Baroux, Célia

    2013-01-01

    In flowering plants, the embryo develops within a nourishing tissue - the endosperm - surrounded by the maternal seed integuments (or seed coat). As a consequence, the isolation of plant embryos at early stages (1 cell to globular stage) is technically challenging due to their relative inaccessibility. Efficient manual dissection at early stages is strongly impaired by the small size of young Arabidopsis seeds and the adhesiveness of the embryo to the surrounding tissues. Here, we describe a method that allows the efficient isolation of young Arabidopsis embryos, yielding up to 40 embryos in 1 hr to 4 hr, depending on the downstream application. Embryos are released into isolation buffer by slightly crushing 250-750 seeds with a plastic pestle in an Eppendorf tube. A glass microcapillary attached to either a standard laboratory pipette (via a rubber tube) or a hydraulically controlled microinjector is used to collect embryos from droplets placed on a multi-well slide on an inverted light microscope. The technical skills required are simple and easily transferable, and the basic setup does not require costly equipment. Collected embryos are suitable for a variety of downstream applications such as RT-PCR, RNA sequencing, DNA methylation analyses, fluorescence in situ hybridization (FISH), immunostaining, and reporter gene assays. PMID:23770918

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

    PubMed

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

    2018-01-01

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

  20. Chronology of early embryonic development and embryo uterine migration in alpacas.

    PubMed

    Picha, Y; Tibary, A; Memon, M; Kasimanickam, R; Sumar, J

    2013-03-01

    The objectives were to: (1) describe the chronology of early embryonic development from ovulation to entry into the uterus; and (2) to determine the timing of embryo migration to the left uterine horn when ovulation occurred from the right ovary. The experiment was conducted in Peru. Females (n = 132) were randomly assigned to 15 experimental groups. All females were mated to an intact male, given 50 μg GnRH im (Cystorelin) and ovulation time determined by transrectal ultrasonography, conducted every 6 hours, starting 24 hours postmating. Animals were slaughtered at a specific intervals postovulation and reproductive tracts were recovered and subjected to oviductal and uterine flushing for females slaughtered between 1 and 6 days postovulation (dpo; Day 0 = ovulation) and uterine flushing for females slaughtered from 7 to 15 dpo for recovery of oocytes/embryos. Season of mating did not influence the interval from mating to ovulation (winter: 29 ± 6 hours vs. summer: 30 ± 6 hours; P = 0.49). Ovulation rates for females mated during winter and summer were 92% versus 100%, respectively (P = 0.05). Fertilization rates for winter and summer mated females were 72% and 82% (P = 0.29). Unfertilized ova were not retained in the uterine tube. All embryos collected were in the uterine tube ipsilateral to the side of ovulation between 1 and 5 dpo. Embryos reached the uterus on 6 dpo. Embryos began to elongate on 9 dpo; at this time, 83% of embryos derived from right-ovary ovulations were collected from the left uterine horn. Embryos occupied the entire uterine cavity by 10 dpo. In conclusion, we characterized early embryo development and location of embryo during its early developmental stages in alpaca. This was apparently the first report regarding chronology of embryo development and migration to the left horn in alpaca which merits further investigation regarding its role in maternal recognition of pregnancy. Copyright © 2013 Elsevier Inc. All rights reserved.

  1. The "waiting period" of sensory and motor axons in early chick hindlimb: its role in axon pathfinding and neuronal maturation.

    PubMed

    Wang, G; Scott, S A

    2000-07-15

    During embryonic development motor axons in the chick hindlimb grow out slightly before sensory axons and wait in the plexus region at the base of the limb for approximately 24 hr before invading the limb itself (Tosney and Landmesser, 1985a). We have investigated the role of this waiting period by asking, Is the arrest of growth cones in the plexus region a general property of both sensory and motor axons? Why do axons wait? Does eliminating the waiting period affect the further development of motor and sensory neurons? Here we show that sensory axons, like motor axons, pause in the plexus region and that neither sensory nor motor axons require cues from the other population to wait in or exit from the plexus region. By transplanting older or younger donor limbs to host embryos, we show that host axons innervate donor limbs on a schedule consistent with the age of the grafted limbs. Thus, axons wait in the plexus region for maturational changes to occur in the limb rather than in the neurons themselves. Both sensory and motor axons innervate their appropriate peripheral targets when the waiting period is eliminated by grafting older donor limbs. Therefore, axons do not require a prolonged period in the plexus region to sort out and project appropriately. Eliminating the waiting period does, however, accelerate the onset of naturally occurring cell death, but it does not enhance the development of central projections or the biochemical maturation of sensory neurons.

  2. FT-IR microscopic mappings of early mineralization in chick limb bud mesenchymal cell cultures

    NASA Technical Reports Server (NTRS)

    Boskey, A. L.; Camacho, N. P.; Mendelsohn, R.; Doty, S. B.; Binderman, I.

    1992-01-01

    Chick limb bud mesenchymal cells differentiate into chondrocytes and form a cartilaginous matrix in culture. In this study, the mineral formed in different areas within cultures supplemented with 4 mM inorganic phosphate, or 2.5, 5.0, and 10 mM beta-glycerophosphate (beta GP), was characterized by Fourier-transform infrared (FT-IR) microscopy. The relative mineral-to-matrix ratios, and distribution of crystal sizes at specific locations throughout the matrix were measured from day 14 to day 30. The only mineral phase detected was a poorly crystalline apatite. Cultures receiving 4 mM inorganic phosphate had smaller crystals which were less randomly distributed around the cartilage nodules than those in the beta GP-treated cultures. beta GP-induced mineral consisted of larger, more perfect apatite crystals. In cultures receiving 5 or 10 mM beta GP, the relative mineral-to-matrix ratios (calculated from the integrated intensities of the phosphate and amide I bands, respectively) were higher than in the cultures with 4 mM inorganic phosphate or in the in vivo calcified chick cartilage.

  3. Early Activation of MAPK and Apoptosis in Nutritive Embryos of Calyptraeid Gastropods.

    PubMed

    Lesoway, Maryna P; Collin, Rachel; Abouheif, Ehab

    2017-07-01

    Investigation of alternative phenotypes, different morphologies produced by a single genome, has contributed novel insights into development and evolution. Yet, the mechanisms underlying developmental switch points between alternative phenotypes remain poorly understood. The calyptraeid snails Crepidula navicella and Calyptraea lichen produce two phenotypes: viable and nutritive embryos, where nutritive embryos arrest their development after gastrulation and are ingested by their viable siblings as a form of intracapsular nutrition. Here, we investigate the activity of mitogen-activated protein kinase (MAPK, ERK1/2) and apoptosis during early cleavage. MAPK and apoptosis, found in a previous transcriptomic study, are known to be involved in organization of other spiralian embryos and nutritive embryo development, respectively. In the model Crepidula fornicata, MAPK activation begins at the 16-cell stage. In contrast, we discovered in C. navicella and C. lichen that many embryos begin MAPK activation at the one-cell stage. A subset of embryos shows a similar pattern of MAPK activation to C. fornicata at later stages. In all stages where MAPK is detected, the activation pattern is highly variable, frequently occurring in all quadrants or in multiple tiers of cells. We also detected apoptosis in cleaving embryos, while C. fornicata and Crepidula lessoni, which do not produce nutritive embryos, show no signs of apoptosis during cleavage. Our results show that MAPK and apoptosis are expressed during early development in species with nutritive embryos, and raises the possibility that these processes may play a role and even interact with one another in producing the nutritive embryo phenotype. © 2017 Wiley Periodicals, Inc.

  4. Metabolic and mitochondrial dysfunction in early mouse embryos following maternal dietary protein intervention.

    PubMed

    Mitchell, Megan; Schulz, Samantha L; Armstrong, David T; Lane, Michelle

    2009-04-01

    Dietary supply of nutrients, both periconception and during pregnancy, influence the growth and development of the fetus and offspring and their health into adult life. Despite the importance of research efforts surrounding the developmental origins of health and disease hypothesis, the biological mechanisms involved remain elusive. Mitochondria are of major importance in the oocyte and early embryo, particularly as a source of ATP generation, and perturbations in their function have been related to reduced embryo quality. The present study examined embryo development following periconception exposure of females to a high-protein diet (HPD) or a low-protein diet (LPD) relative to a medium-protein diet (MPD; control), and we hypothesized that perturbed mitochondrial metabolism in the mouse embryo may be responsible for the impaired embryo and fetal development reported by others. Although the rate of development to the blastocyst stage did not differ between diets, both the HPD and LPD reduced the number of inner cell mass cells in the blastocyst-stage embryo. Furthermore, mitochondrial membrane potential was reduced and mitochondrial calcium levels increased in the 2-cell embryo. Embryos from HPD females had elevated levels of reactive oxygen species and ADP concentrations, indicative of metabolic stress and, potentially, the uncoupling of oxidative phosphorylation, whereas embryos from LPD females had reduced mitochondrial clustering around the nucleus, suggestive of an overall quietening of metabolism. Thus, although periconception dietary supply of different levels of protein is permissive of development, mitochondrial metabolism is altered in the early embryo, and the nature of the perturbation differs between HPD and LPD exposure.

  5. Excess caffeine exposure impairs eye development during chick embryogenesis

    PubMed Central

    Ma, Zheng-lai; Wang, Guang; Cheng, Xin; Chuai, Manli; Kurihara, Hiroshi; Lee, Kenneth Ka Ho; Yang, Xuesong

    2014-01-01

    Caffeine has been an integral component of our diet and medicines for centuries. It is now known that over consumption of caffeine has detrimental effects on our health, and also disrupts normal foetal development in pregnant mothers. In this study, we investigated the potential teratogenic effect of caffeine over-exposure on eye development in the early chick embryo. Firstly, we demonstrated that caffeine exposure caused chick embryos to develop asymmetrical microphthalmia and induced the orbital bone to develop abnormally. Secondly, caffeine exposure perturbed Pax6 expression in the retina of the developing eye. In addition, it perturbed the migration of HNK-1+ cranial neural crest cells. Pax6 is an important gene that regulates eye development, so altering the expression of this gene might be the cause for the abnormal eye development. Thirdly, we found that reactive oxygen species (ROS) production was significantly increased in eye tissues following caffeine treatment, and that the addition of anti-oxidant vitamin C could rescue the eyes from developing abnormally in the presence of caffeine. This suggests that excess ROS induced by caffeine is one of the mechanisms involved in the teratogenic alterations observed in the eye during embryogenesis. In sum, our experiments in the chick embryo demonstrated that caffeine is a potential teratogen. It causes asymmetrical microphthalmia to develop by increasing ROS production and perturbs Pax6 expression. PMID:24636305

  6. Mortality, size of the gonads, and ultrastructure of primordial germ cell in chick embryos treated with gamma-irradiation or injected with donor cells.

    PubMed

    Maeda, T; Clark, M E; Etches, R J

    1998-06-01

    The effects of injection and/or gamma-irradiation prior to injection on mortality, size of the gonads, and ultrastructure of primordial germ cell (PGC) were examined after 5 d of incubation. The mortality of embryos injected with donor cells was significantly higher than that of control and irradiated embryos. All irradiated embryos were alive, although their development was delayed compared to those not exposed to irradiation. The size of the gonads of embryos injected with donor cells were similar to those of control embryos, however, the size of the gonads in irradiated embryos was significantly smaller than those of control embryos. The number of PGC in the gonads was significantly decreased by irradiation. There was no notable effect of irradiation or injection on the nuclei and cytoplasmic organelles in PGC.

  7. 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.

  8. Early detection and staging of spontaneous embryo resorption by ultrasound biomicroscopy in murine pregnancy.

    PubMed

    Flores, Luis E; Hildebrandt, Thomas B; Kühl, Anja A; Drews, Barbara

    2014-05-10

    Embryo resorption is a major problem in human medicine, agricultural animal production and in conservation breeding programs. Underlying mechanisms have been investigated in the well characterised mouse model. However, post mortem studies are limited by the rapid disintegration of embryonic structures. A method to reliably identify embryo resorption in alive animals has not been established yet. In our study we aim to detect embryos undergoing resorption in vivo at the earliest possible stage by ultra-high frequency ultrasound. In a longitudinal study, we monitored 30 pregnancies of wild type C57BI/6 mice using ultra-high frequency ultrasound (30-70 MHz), so called ultrasound biomicroscopy (UBM). We compared the sonoembryology of mouse conceptuses under spontaneous resorption and neighbouring healthy conceptuses and correlated the live ultrasound data with the respective histology. The process of embryo resorption comprised of four stages: first, the conceptus exhibited growth retardation, second, bradycardia and pericardial edema were observed, third, further development ceased and the embryo died, and finally embryo remnants were resorbed by maternal immune cells. In early gestation (day 7 and 8), growth retardation was characterized by a small embryonic cavity. The embryo and its membranes were ill defined or did not develop at all. The echodensity of the embryonic fluid increased and within one to two days, the embryo and its cavity disappeared and was transformed into echodense tissue surrounded by fluid filled caverns. In corresponding histologic preparations, fibrinoid material interspersed with maternal granulocytes and lacunae filled with maternal blood were observed. In later stages (day 9-11) resorption prone embryos were one day behind in their development compared to their normal siblings. The space between Reichert's membrane and inner yolk sac membrane was enlarged The growth retarded embryos exhibited bradycardia and ultimately cessation of heart

  9. Early detection and staging of spontaneous embryo resorption by ultrasound biomicroscopy in murine pregnancy

    PubMed Central

    2014-01-01

    Background Embryo resorption is a major problem in human medicine, agricultural animal production and in conservation breeding programs. Underlying mechanisms have been investigated in the well characterised mouse model. However, post mortem studies are limited by the rapid disintegration of embryonic structures. A method to reliably identify embryo resorption in alive animals has not been established yet. In our study we aim to detect embryos undergoing resorption in vivo at the earliest possible stage by ultra-high frequency ultrasound. Methods In a longitudinal study, we monitored 30 pregnancies of wild type C57BI/6 mice using ultra-high frequency ultrasound (30-70 MHz), so called ultrasound biomicroscopy (UBM). We compared the sonoembryology of mouse conceptuses under spontaneous resorption and neighbouring healthy conceptuses and correlated the live ultrasound data with the respective histology. Results The process of embryo resorption comprised of four stages: first, the conceptus exhibited growth retardation, second, bradycardia and pericardial edema were observed, third, further development ceased and the embryo died, and finally embryo remnants were resorbed by maternal immune cells. In early gestation (day 7 and 8), growth retardation was characterized by a small embryonic cavity. The embryo and its membranes were ill defined or did not develop at all. The echodensity of the embryonic fluid increased and within one to two days, the embryo and its cavity disappeared and was transformed into echodense tissue surrounded by fluid filled caverns. In corresponding histologic preparations, fibrinoid material interspersed with maternal granulocytes and lacunae filled with maternal blood were observed. In later stages (day 9–11) resorption prone embryos were one day behind in their development compared to their normal siblings. The space between Reichert’s membrane and inner yolk sac membrane was enlarged The growth retarded embryos exhibited bradycardia and

  10. Early embryonic survival and embryo development in two lines of rabbits divergently selected for uterine capacity.

    PubMed

    Peiró, R; Santacreu, M A; Climent, A; Blasco, A

    2007-07-01

    The aim of this work is to study early embryo survival and development in 2 lines divergently selected for high and low uterine capacity throughout 10 generations. A total of 162 female rabbits from the high line and 133 from the low line were slaughtered at 25, 48, or 62 h of gestation. There were no differences in ovulation rate and fertilization rate between lines in any of the 3 stages of gestation. Embryo survival, estimated as the number of normal embryos recovered at a constant ovulation rate, was similar in both lines at 25 and 48 h. Embryo survival was greater in the high line [D (posterior mean of the difference between the high and low lines) = 0.57 embryos] at 62 h of gestation. There was no difference in embryonic stage of development at 25 h, but at 48 and 62 h of gestation, the high line, compared with the low line, had a greater percentage of early morulae (83 vs. 72%) and compacted morulae (55 vs. 38%). Divergent selection for uterine capacity appeared to modify embryo development, at least from 48 h of gestation, and embryo survival from 62 h.

  11. The early Cambrian fossil embryo Pseudooides is a direct-developing cnidarian, not an early ecdysozoan

    PubMed Central

    2017-01-01

    Early Cambrian Pseudooides prima has been described from embryonic and post-embryonic stages of development, exhibiting long germ-band development. There has been some debate about the pattern of segmentation, but this interpretation, as among the earliest records of ecdysozoans, has been generally accepted. Here, we show that the ‘germ band’ of P. prima embryos separates along its mid axis during development, with the transverse furrows between the ‘somites’ unfolding into the polar aperture of the ten-sided theca of Hexaconularia sichuanensis, conventionally interpreted as a scyphozoan cnidarian; co-occurring post-embryonic remains of ecdysozoans are unrelated. We recognize H. sichuanensis as a junior synonym of P. prima as a consequence of identifying these two form-taxa as distinct developmental stages of the same organism. Direct development in P. prima parallels the co-occuring olivooids Olivooides, and Quadrapyrgites and Bayesian phylogenetic analysis of a novel phenotype dataset indicates that, despite differences in their tetra-, penta- and pseudo-hexa-radial symmetry, these hexangulaconulariids comprise a clade of scyphozoan medusozoans, with Arthrochites and conulariids, that all exhibit direct development from embryo to thecate polyp. The affinity of hexangulaconulariids and olivooids to extant scyphozoan medusozoans indicates that the prevalence of tetraradial symmetry and indirect development are a vestige of a broader spectrum of body-plan symmetries and developmental modes that was manifest in their early Phanerozoic counterparts. PMID:29237861

  12. The early Cambrian fossil embryo Pseudooides is a direct-developing cnidarian, not an early ecdysozoan.

    PubMed

    Duan, Baichuan; Dong, Xi-Ping; Porras, Luis; Vargas, Kelly; Cunningham, John A; Donoghue, Philip C J

    2017-12-20

    Early Cambrian Pseudooides prima has been described from embryonic and post-embryonic stages of development, exhibiting long germ-band development. There has been some debate about the pattern of segmentation, but this interpretation, as among the earliest records of ecdysozoans, has been generally accepted. Here, we show that the 'germ band' of P. prima embryos separates along its mid axis during development, with the transverse furrows between the 'somites' unfolding into the polar aperture of the ten-sided theca of Hexaconularia sichuanensis , conventionally interpreted as a scyphozoan cnidarian; co-occurring post-embryonic remains of ecdysozoans are unrelated. We recognize H. sichuanensis as a junior synonym of P. prima as a consequence of identifying these two form-taxa as distinct developmental stages of the same organism. Direct development in P. prima parallels the co-occuring olivooids Olivooides, and Quadrapyrgites and Bayesian phylogenetic analysis of a novel phenotype dataset indicates that, despite differences in their tetra-, penta- and pseudo-hexa-radial symmetry, these hexangulaconulariids comprise a clade of scyphozoan medusozoans, with Arthrochites and conulariids, that all exhibit direct development from embryo to thecate polyp. The affinity of hexangulaconulariids and olivooids to extant scyphozoan medusozoans indicates that the prevalence of tetraradial symmetry and indirect development are a vestige of a broader spectrum of body-plan symmetries and developmental modes that was manifest in their early Phanerozoic counterparts. © 2017 The Authors.

  13. The early-stage diagnosis of albinic embryos by applying optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Yang, Bor-Wen; Wang, Shih-Yuan; Wang, Yu-Yen; Cai, Jyun-Jhang; Chang, Chung-Hao

    2013-09-01

    Albinism is a kind of congenital disease of abnormal metabolism. Poecilia reticulata (guppy fish) is chosen as the model to study the development of albinic embryos as it is albinic, ovoviviparous and with short life period. This study proposed an imaging method for penetrative embryo investigation using optical coherence tomography. By imaging through guppy mother’s reproduction purse, we found the embryo’s eyes were the early-developed albinism features. As human’s ocular albinism typically appear at about four weeks old, it is the time to determine if an embryo will grow into an albino.

  14. Hippocampus and medial striatum dissociation during goal navigation by geometry or features in the domestic chick: An immediate early gene study.

    PubMed

    Mayer, Uwe; Pecchia, Tommaso; Bingman, Verner Peter; Flore, Michele; Vallortigara, Giorgio

    2016-01-01

    We employed a standard reference memory task to study the involvement of the hippocampal formation (HF) of domestic chicks that used the boundary geometry of a test environment to orient to and locate a reward. Using the immediate early gene product c-Fos as a neuronal activity marker, we found enhanced HF activation in chicks that learned to locate rewarded corners using the shape of a rectangular arena compared to chicks trained to solve the task by discriminating local features in a square-shaped arena. We also analyzed neuronal activity in the medial part of the medial striatum (mMSt). Surprisingly, in mMSt we observed a reverse pattern, with higher activity in the chicks that were trained to locate the goal by local features. Our results identify two seemingly parallel, memory systems in chicks, with HF central to the processing of spatial-geometrical information and mMSt important in supporting local feature discrimination. © 2015 Wiley Periodicals, Inc.

  15. Analysis of NUAK1 and NUAK2 expression during early chick development reveals specific patterns in the developing head.

    PubMed

    Bekri, Abdelhamid; Billaud, Marc; Thélu, Jacques

    2014-01-01

    Several human diseases are associated with the NUAK1 and NUAK2 genes. These genes encode kinases, members of the AMPK-related kinases (ARK) gene family. Both NUAK1 and NUAK2 are known targets of the serine threonine kinase LKB1, a tumor suppressor involved in regulating cell polarity. While much is known about their functions in disease, their expression pattern in normal development has not been extensively studied. Here, we present the expression patterns for NUAK1 and NUAK2 in the chick during early-stage embryogenesis, until day 3 (Hamburger and Hamilton stage HH20). Several embryonic structures, in particular the nascent head, showed distinct expression levels. NUAK1 expression was first detected at stage HH6 in the rostral neural folds. It was then expressed (HH7-11) throughout the encephalalon, predominantly in the telencephalon and mesencephalon. NUAK1 expression was also detected in the splanchnic endoderm area at HH8-10, and in the vitellin vein derived from this area, but not in the heart. NUAK2 expression was first detected at stage HH6 in the neural folds. It was then found throughout the encephalon at stage HH20. Particular attention was paid in this study to the dorsal ectoderm at stages HH7 and HH8, where a local deficit or accumulation of NUAK2 mRNA were found to correlate with the direction of curvature of the neural plate. This is the first description of NUAK1 and NUAK2 expression patterns in the chick during early development; it reveals non-identical expression profiles for both genes in neural development.

  16. Expression of hLAMP-1-Positive Particles During Early Heart Development in the Chick.

    PubMed

    Abd-Elhamid, T H; Conway, M L; Sinning, A R

    2017-10-01

    Heart development requires coordinated activity of various factors, the disturbance of which can lead to congenital heart defects. Heart lectin-associated matrix protein-1 (hLAMP-1) is a matrix protein expressed within Hensen's node at Hamburger-Hamilton (HH) stage 4, in the lateral mesoderm by HH stages 5-6 and enhanced within the left pre-cardiac field at HH stage 7. At HH stages 15-16, hLAMP-1 expression is observed in the atrioventricular canal and the outflow tract. Also, the role of hLAMP-1 in induction of mesenchyme formation in chick heart has been well documented. To further elucidate the role of this molecule in heart development, we examined its expression patterns during HH stages 8-14 in the chick. In this regard, we immunostained sections of the heart during HH stages 8-14 with antibodies specific to hLAMP-1. Our results showed prominent expression of hLAMP-1-positive particles in the extracellular matrix associated with the pre-cardiac mesoderm, the endoderm, ectoderm as well as neuroectoderm at HH stages 8-9. After formation of the linear heart tube at HH stage 10, the expression of hLAMP-1-stained particles disappears in those regions of original contact between the endoderm and heart forming fields due to rupture of the dorsal mesocardium while their expression becomes confined to the arterial and venous poles of the heart tube. This expression pattern is maintained until HH stage 14. This expression pattern suggests that hLAMP-1 may be involved in the formation of the endocardial tube. © 2017 Blackwell Verlag GmbH.

  17. MHC class I target recognition, immunophenotypes and proteomic profiles of natural killer cells within the spleens of day-14 chick embryos

    Chicken natural killer (NK) cells are not well defined, so little is known about the molecular interactions controlling their activity. At day 14 of embryonic development, chick spleens are a rich source of T-cellfree CD8aa+, CD3_ cells with natural killing activity. Cell-mediated cytotoxicity assay...

  18. Laminarin improves developmental competence of porcine early stage embryos by inhibiting oxidative stress.

    PubMed

    Jiang, Hao; Liang, Shuang; Yao, Xue-Rui; Jin, Yong-Xun; Shen, Xing-Hui; Yuan, Bao; Zhang, Jia-Bao; Kim, Nam-Hyung

    2018-04-23

    Laminarin (LMA), a β-glucan mixture with good biocompatibility, improves the growth performance and immune response when used as food additives and nutraceuticals. The aim of the present research was to explore the effects of LMA on porcine early stage embryo development, as well as the underlying mechanisms. The results showed that the developmental competence of porcine early stage embryos was dramatically improved after LMA supplementation during the in vitro culture period. The presence of 20 μg/mL LMA during the in vitro culture period significantly improved cleavage rate, blastocyst formation rates, hatching rate, and total cell number in the blastocyst compared to that in the control group. Notably, LMA attenuated the intracellular reactive oxygen species generation induced by H 2 O 2 . Furthermore, LMA not only increased intracellular glutathione levels, but also ameliorated mitochondrial membrane potential. In addition, the expression of a zygotic genome activation related gene (YAP1), pluripotency-related genes (OCT4, NANOG, and SOX2), and hatching-related genes (COX2, GATA4, and ITGA5) were up-regulated following LMA supplementation during porcine early stage embryo development. These results demonstrate that LMA has beneficial effects on the development of porcine early stage embryos via regulation of oxidative stress. This evidence provides a novel method for embryo development improvement associated with exposure to LMA. Copyright © 2018 Elsevier Inc. All rights reserved.

  19. Wounded cells drive rapid epidermal repair in the early Drosophila embryo

    PubMed Central

    Fernandez-Gonzalez, Rodrigo; Zallen, Jennifer A.

    2013-01-01

    Epithelial tissues are protective barriers that display a remarkable ability to repair wounds. Wound repair is often associated with an accumulation of actin and nonmuscle myosin II around the wound, forming a purse string. The role of actomyosin networks in generating mechanical force during wound repair is not well understood. Here we investigate the mechanisms of force generation during wound repair in the epidermis of early and late Drosophila embryos. We find that wound closure is faster in early embryos, where, in addition to a purse string around the wound, actomyosin networks at the medial cortex of the wounded cells contribute to rapid wound repair. Laser ablation demonstrates that both medial and purse-string actomyosin networks generate contractile force. Quantitative analysis of protein localization dynamics during wound closure indicates that the rapid contraction of medial actomyosin structures during wound repair in early embryos involves disassembly of the actomyosin network. By contrast, actomyosin purse strings in late embryos contract more slowly in a mechanism that involves network condensation. We propose that the combined action of two force-generating structures—a medial actomyosin network and an actomyosin purse string—contributes to the increased efficiency of wound repair in the early embryo. PMID:23985320

  20. Sensitivity of early mouse embryos to (/sup 3/H)thymidine

    SciT

    Spindle, A.; Wu, K.; Pedersen, R.A.

    1982-12-01

    Effects of intranuclear radiation on the developmental capacity of early mouse embryos were studied by exposing embryos to (/sup 3/H)thymidine and counting the number of embryos forming blastocysts, trophoblast outgrowths, inner cell masses (ICMs), and two-layer ICMs (differentiated into primary endoderm and ectoderm). When embryos were cultured from the 2-cell stage for 8 days in the continuous presence of (/sup 3/H)thymidine, concentrations as low as 0.2 nCi/ml reduced the number of embryos forming two-layer ICMs. At 1 nCi/ml, the number of both ICMs and two-layer ICMs were reduced, and at 10 nCi/ml the number of embryos developing to all threemore » post-blastocyst endpoints was reduced. Blastocyst formation was not affected even at the highst concentration (/sup 3/H)thymidine and then cultured further in unlabelled medium, the effects were similar to those of 8-day exposure. When embryos were exposed to (/sup 3/H)thymidine for 24 h at various developmental stages, effects were less severe than when they were exposed continuously for 3 or 8 days, and the sensitivity of embryos differed between stages. The 24-h exposure of immunosurgically isolated ICMS to (/sup 3/H)thymidine revealed that the high sensitivity of the ICM to (/sup 3/H)thymidine persists through the late blastocyst stage and declines progressively thereafter. Autoradiography indicated that the change in radiosensitivity of embryos or ICMs is generally related to their ability to incorporate (/sup 3/H)thymidine into the DNA.« less

  1. Paternal Diet-Induced Obesity Retards Early Mouse Embryo Development, Mitochondrial Activity and Pregnancy Health

    PubMed Central

    Binder, Natalie K.; Hannan, Natalie J.; Gardner, David K.

    2012-01-01

    Worldwide, 48% of adult males are overweight or obese. An association between infertility and excessive body weight is now accepted, although focus remains primarily on females. It has been shown that parental obesity results in compromised embryo development, disproportionate changes in embryo metabolism and reduced blastocyst cell number. The aim of this study was to determine whether paternal obesity has negative effects on the resultant embryo. Specifically, using in vitro fertilisation (IVF), we wanted to isolate the functional effects of obesity on sperm by examining the subsequent embryo both pre- and post-implantation. Epididymal sperm was collected from age matched normal and obese C57BL/6 mice and cryopreserved for subsequent IVF with oocytes collected from Swiss females (normal diet/weight). Obesity was induced in male mice by feeding a high fat diet of 22% fat for 10 weeks. Resultant embryos were cultured individually and development monitored using time-lapse microscopy. Paternal obesity resulted in a significant delay in preimplantation embryo development as early as syngamy (P<0.05). Metabolic parameters were measured across key developmental stages, demonstrating significant reduction in mitochondrial membrane potential (P<0.01). Blastocysts were stained to determine trophectoderm (TE) and inner cell mass (ICM) cell numbers, revealing significant differences in the ratio of cell allocation to TE and ICM lineages (P<0.01). Functional studies examining blastocyst attachment, growth and implantation demonstrated that blastocysts derived from sperm of obese males displayed significantly reduced outgrowth on fibronectin in vitro (P<0.05) and retarded fetal development in vivo following embryo transfer (P<0.05). Taken together, these data clearly demonstrate that paternal obesity has significant negative effects on the embryo at a variety of key early developmental stages, resulting in delayed development, reduced placental size and smaller offspring

  2. Early bovine embryos regulate oviduct epithelial cell gene expression during in vitro co-culture.

    PubMed

    Schmaltz-Panneau, Barbara; Cordova, Amanda; Dhorne-Pollet, Sophie; Hennequet-Antier, Christelle; Uzbekova, Sveltlana; Martinot, Emmanuelle; Doret, Sarah; Martin, Patrice; Mermillod, Pascal; Locatelli, Yann

    2014-10-01

    In mammals, the oviduct may participate to the regulation of early embryo development. In vitro co-culture of early bovine embryos with bovine oviduct epithelial cells (BOEC) has been largely used to mimic the maternal environment. However, the mechanisms of BOEC action have not been clearly elucidated yet. The aim of this study was to determine the response of BOEC cultures to the presence of developing bovine embryos. A 21,581-element bovine oligonucleotide array was used compare the gene expression profiles of confluent BOEC cultured for 8 days with or without embryos. This study revealed 34 differentially expressed genes (DEG). Of these 34 genes, IFI6, ISG15, MX1, IFI27, IFI44, RSAD2, IFITM1, EPSTI1, USP18, IFIT5, and STAT1 expression increased to the greatest extent due to the presence of embryos with a major impact on antiviral and immune response. Among the mRNAs at least 25 are already described as induced by interferons. In addition, transcript levels of new candidate genes involved in the regulation of transcription, modulation of the maternal immune system and endometrial remodeling were found to be increased. We selected 7 genes and confirmed their differential expression by quantitative RT-PCR. The immunofluorescence imaging of cellular localization of STAT1 protein in BOEC showed a nuclear translocation in the presence of embryos, suggesting the activation of interferon signaling pathway. This first systematic study of BOEC transcriptome changes in response to the presence of embryos in cattle provides some evidences that these cells are able to adapt their transcriptomic profile in response to embryo signaling. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. Differential muscular myosin heavy chain expression of the pectoral and pelvic girdles during early growth in the king penguin (Aptenodytes patagonicus) chick.

    PubMed

    Erbrech, Aude; Robin, Jean-Patrice; Guérin, Nathalie; Groscolas, René; Gilbert, Caroline; Martrette, Jean-Marc

    2011-06-01

    Continuous growth, associated with a steady parental food supply, is a general pattern in offspring development. So that young chicks can acquire their locomotor independence, this period is usually marked by a fast maturation of muscles, during which different myosin heavy chain (MyHC) isoforms are expressed. However, parental food provisioning may fluctuate seasonally, and offspring therefore face a challenge to ensure the necessary maturation of their tissues when energy is limited. To address this trade-off we investigated muscle maturation in both the pectoral and pelvic girdles of king penguin chicks. This species has an exceptionally long rearing period (1 year), which is prolonged when parental food provisioning is drastically reduced during the sub-Antarctic winter. Approximately 1 month post hatching, chicks acquire a functional pedestrian locomotion, which uses pelvic muscles, whereas swimming, which uses the pectoral muscles, only occurs 1 year later. We therefore tested the hypothesis that the MyHC content of the leg muscles reaches a mature state before those of the pectoral muscles. We found that leg muscle MyHC composition changed with the progressive acquisition of pedestrian locomotion, whereas pectoral muscle fibres reached their mature MyHC profile as early as hatching. Contrary to our predictions, the acquisition of the adult profile in pectoral muscles could be related to an early maturation of the contractile muscular proteins, presumably associated with early thermoregulatory capacities of chicks, necessary for survival in their cold environment. This differential maturation appears to reconcile both the locomotor and environmental constraints of king penguin chicks during growth.

  4. Exceptional preservation of tiny embryos documents seed dormancy in early angiosperms.

    PubMed

    Friis, Else Marie; Crane, Peter R; Pedersen, Kaj Raunsgaard; Stampanoni, Marco; Marone, Federica

    2015-12-24

    The rapid diversification of angiosperms through the Early Cretaceous period, between about 130-100 million years ago, initiated fundamental changes in the composition of terrestrial vegetation and is increasingly well understood on the basis of a wealth of palaeobotanical discoveries over the past four decades and their integration with improved knowledge of living angiosperms. Prevailing hypotheses, based on evidence both from living and from fossil plants, emphasize that the earliest angiosperms were plants of small stature with rapid life cycles that exploited disturbed habitats in open, or perhaps understorey, conditions. However, direct palaeontogical data relevant to understanding the seed biology and germination ecology of Early Cretaceous angiosperms are sparse. Here we report the discovery of embryos and their associated nutrient storage tissues in exceptionally well-preserved angiosperm seeds from the Early Cretaceous. Synchrotron radiation X-ray tomographic microscopy of the fossil embryos from many taxa reveals that all were tiny at the time of dispersal. These results support hypotheses based on extant plants that tiny embryos and seed dormancy are basic for angiosperms as a whole. The minute size of the fossil embryos, and the modest nutrient storage tissues dictated by the overall small seed size, is also consistent with the interpretation that many early angiosperms were opportunistic, early successional colonizers of disturbance-prone habitats.

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

    PubMed

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

    2009-03-01

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

  6. Expression of voltage-activated calcium channels in the early zebrafish embryo.

    PubMed

    Sanhueza, Dayán; Montoya, Andro; Sierralta, Jimena; Kukuljan, Manuel

    2009-05-01

    Increases in cytosolic calcium concentrations regulate many cellular processes, including aspects of early development. Calcium release from intracellular stores and calcium entry through non-voltage-gated channels account for signalling in non-excitable cells, whereas voltage-gated calcium channels (CaV) are important in excitable cells. We report the expression of multiple transcripts of CaV, identified by its homology to other species, in the early embryo of the zebrafish, Danio rerio, at stages prior to the differentiation of excitable cells. CaV mRNAs and proteins were detected as early as the 2-cell stages, which indicate that they arise from both maternal and zygotic transcription. Exposure of embryos to pharmacological blockers of CaV does not perturb early development significantly, although late effects are appreciable. These results suggest that CaV may have a role in calcium homeostasis and control of cellular process during early embryonic development.

  7. Chick ex ovo culture and ex ovo CAM assay: how it really works.

    PubMed

    Dohle, Daniel S; Pasa, Susanne D; Gustmann, Sebastian; Laub, Markus; Wissler, Josef H; Jennissen, Herbert P; Dünker, Nicole

    2009-11-30

    Chicken eggs in the early phase of breeding are between in vitro and in vivo systems and provide a vascular test environment not only to study angiogenesis but also to study tumorigenesis. After the chick chorioallantoic membrane (CAM) has developed, its blood vessel network can be easily accessed, manipulated and observed and therefore provides an optimal setting for angiogenesis assays. Since the lymphoid system is not fully developed until late stages of incubation, the chick embryo serves as a naturally immunodeficient host capable of sustaining grafted tissues and cells without species-specific restrictions. In addition to nurturing developing allo- and xenografts, the CAM blood vessel network provides a uniquely supportive environment for tumor cell intravasation, dissemination, and vascular arrest and a repository where arrested cells extravasate to form micro metastatic foci. For experimental purposes, in most of the recent studies the CAM was exposed by cutting a window through the egg shell and experiments were carried out in ovo, resulting in significant limitations in the accessibility of the CAM and possibilities for observation and photo documentation of effects. When shell-less cultures of the chick embryo were used(1-4), no experimental details were provided and, if published at all, the survival rates of these cultures were low. We refined the method of ex ovo culture of chick embryos significantly by introducing a rationally controlled extrusion of the egg content. These ex ovo cultures enhance the accessibility of the CAM and chick embryo, enabling easy in vivo documentation of effects and facilitating experimental manipulation of the embryo. This allows the successful application to a large number of scientific questions: (1) As an improved angiogenesis assay(5,6), (2) an experimental set up for facilitated injections in the vitreous of the chick embryo eye(7-9), (3) as a test environment for dissemination and intravasation of dispersed tumor

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

    PubMed Central

    Jarvis, Gavin E.

    2017-01-01

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

  9. 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

  10. Changes in Oscillatory Dynamics in the Cell Cycle of Early Xenopus laevis Embryos

    PubMed Central

    Tsai, Tony Y.-C.; Theriot, Julie A.; Ferrell, James E.

    2014-01-01

    During the early development of Xenopus laevis embryos, the first mitotic cell cycle is long (∼85 min) and the subsequent 11 cycles are short (∼30 min) and clock-like. Here we address the question of how the Cdk1 cell cycle oscillator changes between these two modes of operation. We found that the change can be attributed to an alteration in the balance between Wee1/Myt1 and Cdc25. The change in balance converts a circuit that acts like a positive-plus-negative feedback oscillator, with spikes of Cdk1 activation, to one that acts like a negative-feedback-only oscillator, with a shorter period and smoothly varying Cdk1 activity. Shortening the first cycle, by treating embryos with the Wee1A/Myt1 inhibitor PD0166285, resulted in a dramatic reduction in embryo viability, and restoring the length of the first cycle in inhibitor-treated embryos with low doses of cycloheximide partially rescued viability. Computations with an experimentally parameterized mathematical model show that modest changes in the Wee1/Cdc25 ratio can account for the observed qualitative changes in the cell cycle. The high ratio in the first cycle allows the period to be long and tunable, and decreasing the ratio in the subsequent cycles allows the oscillator to run at a maximal speed. Thus, the embryo rewires its feedback regulation to meet two different developmental requirements during early development. PMID:24523664

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

    Gilbert Levesque

    1963-01-01

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

  12. Effects of embryo size at transfer (whole versus demi) and early pregnancy progesterone supplementation on embryo growth and pregnancy-specific protein bovine concentrations in recipient dairy heifers.

    PubMed

    Lopes-da-Costa, L; Chagas e Silva, J; Deloche, M C; Jeanguyot, N; Humblot, P; Horta, A E M

    2011-08-01

    The objectives of this study were to evaluate embryonic size and survival, plasma progesterone (P4) and pregnancy-specific protein bovine (PSPB) concentrations in early pregnancies (n = 99) following the transfer of one whole (n = 66) or one demi (n = 33) embryo to recipient virgin dairy heifers. The experiment was designed to evaluate the fixed effects of embryo size at transfer (whole or demi embryo) on Day 7 of the estrous cycle (Day 0 = estrus) and P4 supplementation between Days 7 to 19 through an intravaginal device (yes or no) on plasma P4 and PSPB concentrations and on embryo measurements. Plasma P4 concentrations were measured by RIA on Days 0, 7, 14, 19, 21, 25, 35, 42, 49, 56 and 63 of pregnancy and, PSPB concentrations were measured by ELISA on Days 7, 21, 25, 35, 42, 49, 56 and 63. The presence of an embryonic vesicle was detected on Day 25, embryonic/fetal movements and heartbeat were evaluated on Days 42 and 63 and embryo measurements [crown-rump length (CRL) and width at mid body] were obtained on Day 42 through ultrasonography. In non-supplemented pregnancies, Day 42 whole embryos had higher (P < 0.05) CRL and width than demi embryos, but the difference averaged only 1 to 2 mm. In P4 supplemented pregnancies, whole and demi embryos attained a similar size on Day 42 of pregnancy. Embryo size at transfer, early exogenous P4 supplementation and their interactions had no effects (P > 0.05) on plasma P4 concentrations. However, the post-hoc LSD evaluation showed that plasma P4 concentrations on Day 25 were higher (P < 0.001) in whole than in demi embryo derived pregnancies and, that exogenous P4 supplementation increased (P < 0.05) plasma P4 concentrations on Day 19 of pregnancy. The plasma PSPB detection rate on Days 7 to 63 of pregnancy was similar in pregnancies resulting from the transfer of whole and demi embryos. From a total of 93 recipients remaining pregnant until Day 63, plasma PSPB was constantly undetectable on Day 7, was detected in 4% of

  13. The influence of early embryo traits on human embryonic stem cell derivation efficiency.

    PubMed

    O'Leary, Thomas; Heindryckx, Björn; Lierman, Sylvie; Van der Jeught, Margot; Menten, Björn; Deforce, Dieter; Cornelissen, Ria; de Sousa Lopes, Susana Chuva; De Sutter, Petra

    2011-05-01

    Despite its prognostic value in in vitro fertilization, early embryo morphology is not reported on in the derivation of human embryonic stem cell (hESC) lines. Standard hESC derivation does rely on blastocyst development and its efficiency is highly correlated to inner cell mass (ICM) quality. Poor-quality embryos (PQEs) donated for hESC derivation may have a range of cleavage-stage abnormalities that are known to compromise further development. This study was implemented to determine whether specific PQEs traits influence the efficiency of good-quality ICMs to derive new hESC lines. We found that although the types of PQEs investigated were all able to make blastocysts with good-quality ICMs, the ICMs were unequal in their ability to derive hESCs. Good-quality ICMs from embryos with multiple poor-quality traits were unable to generate hESC lines, in contrast to good-quality ICMs from embryos with a single poor-quality trait. In addition, our data suggest a direct correlation between the number of ICM cells present in the blastocyst and its capacity to derive new hESC lines. This study is the first to demonstrate that ICM quality alone is an incomplete indicator of hESC derivation and that application of in vitro fertilization-based early embryo scoring can help predict hESC derivation efficiency. Experiments aiming to quantify, improve upon, or compare hESC derivation efficiency should thus take into consideration early embryo morphology scoring for the comparison of groups with equal developmental competence.

  14. Immunological mechanisms to establish embryo tolerance in early bovine pregnancy.

    PubMed

    Groebner, A E; Schulke, K; Schefold, J C; Fusch, G; Sinowatz, F; Reichenbach, H D; Wolf, E; Meyer, H H D; Ulbrich, S E

    2011-01-01

    A well-balanced immunological interaction between mother and the semi-allogenic embryo is of particular importance. The objective of the present study was to analyse mechanisms of immune tolerance in bovine pregnancy during peri-implantation. Simmental heifers inseminated with either cryopreserved spermatozoa or seminal plasma were killed 12, 15 or 18 days after oestrus. Uteri were flushed for the recovery of conceptuses and the ipsilateral intercaruncular endometrium was sampled for gene expression analysis. Indoleamine 2,3-dioxygenase (IDO) mRNA, coding for the initial enzyme of the kynurenine pathway, was 18-fold (P < 0.001) more abundant in the endometrium of Day 18 pregnant v. non-pregnant animals. Tandem mass spectrometry revealed a decrease of endometrial l-tryptophan (P = 0.0008), but an increase of l-kynurenine concentration (P = 0.005) from Day 12 to Day 18, suggesting increasing IDO activity (P < 0.03). An in vitro coculture model of endometrial cells showed an induction of IDO expression following interferon-τ exposure primarily in stroma cells, which was confirmed by in situ hybridisation localising IDO mRNA mainly in deep stroma cells. Immunohistochemical analysis revealed fewer CD45-positive leucocytes in the zona basalis of pregnant animals. Elevated IDO activity may reduce the presence of leucocytes in the pregnant endometrium, providing a possible mechanism for protecting the semi-allogenic conceptus from maternal rejection.

  15. Influence of follicle characteristics at ovulation on early embryo survival

    Reproductive losses are significant in domestic animals and represent huge economic inefficiencies in livestock production. Across livestock species, embryonic mortality is the greatest contributor to reproductive losses. In most livestock species, early pregnancy diagnosis is an impediment to mak...

  16. Nucleolus Precursor Bodies and Ribosome Biogenesis in Early Mammalian Embryos: Old Theories and New Discoveries.

    PubMed

    Fulka, Helena; Aoki, Fugaku

    2016-06-01

    In mammals, mature oocytes and early preimplantation embryos contain transcriptionally inactive structures termed nucleolus precursor bodies instead of the typical fibrillo-granular nucleoli. These nuclear organelles are essential and strictly of maternal origin. If they are removed from oocytes, the resulting embryos are unable to replace them and consequently fail to develop. Historically, nucleolus precursor bodies have been perceived as a passive repository site of nucleolar proteins that are required for embryos to form fully functional nucleoli. Recent results, however, contradict this long-standing dogma and show that these organelles are dispensable for nucleologenesis and ribosome biogenesis. In this article, we discuss the possible roles of nucleolus precursor bodies and propose how they might be involved in embryogenesis. Furthermore, we argue that these organelles are essential only shortly after fertilization and suggest that they might actively participate in centromeric chromatin establishment. © 2016 by the Society for the Study of Reproduction, Inc.

  17. Expression of genes involved in early cell fate decisions in human embryos and their regulation by growth factors.

    PubMed

    Kimber, S J; Sneddon, S F; Bloor, D J; El-Bareg, A M; Hawkhead, J A; Metcalfe, A D; Houghton, F D; Leese, H J; Rutherford, A; Lieberman, B A; Brison, D R

    2008-05-01

    Little is understood about the regulation of gene expression in human preimplantation embryos. We set out to examine the expression in human preimplantation embryos of a number of genes known to be critical for early development of the murine embryo. The expression profile of these genes was analysed throughout preimplantation development and in response to growth factor (GF) stimulation. Developmental expression of a number of genes was similar to that seen in murine embryos (OCT3B/4, CDX2, NANOG). However, GATA6 is expressed throughout preimplantation development in the human. Embryos were cultured in IGF-I, leukaemia inhibitory factor (LIF) or heparin-binding EGF-like growth factor (HBEGF), all of which are known to stimulate the development of human embryos. Our data show that culture in HBEGF and LIF appears to facilitate human embryo expression of a number of genes: ERBB4 (LIF) and LIFR and DSC2 (HBEGF) while in the presence of HBEGF no blastocysts expressed EOMES and when cultured with LIF only two out of nine blastocysts expressed TBN. These data improve our knowledge of the similarities between human and murine embryos and the influence of GFs on human embryo gene expression. Results from this study will improve the understanding of cell fate decisions in early human embryos, which has important implications for both IVF treatment and the derivation of human embryonic stem cells.

  18. Early development of Xenopus embryos is affected by simulated gravity

    NASA Technical Reports Server (NTRS)

    Yokota, Hiroki; Neff, Anton W.; Malacinski, George M.

    1994-01-01

    Early amphibian (Xenopus laevis) development under clinostat-simulated weightlessness and centrifuge-simulated hypergravity was studied. The results revealed significant effects on (i) 'morphological patterning' such as the cleavage furrow pattern in the vegetal hemisphere at the eight-cell stage and the shape of the dorsal lip in early gastrulae and (ii) 'the timing of embryonic events' such as the third cleavage furrow completion and the dorsal lip appearance. Substantial variations in sensitivity to simulated force fields were observed, which should be considered in interpreting spaceflight data.

  19. Elevated aminopeptidase N affects sperm motility and early embryo development

    PubMed Central

    Ryu, Do-Yeal; Kwon, Woo-Sung

    2017-01-01

    Aminopeptidase N (APN) is a naturally occurring ectopeptidase present in mammalian semen. Previous studies have demonstrated that APN adversely affects male fertility through the alteration of sperm motility. This enzyme constitutes 0.5 to 1% of the seminal plasma proteins, which can be transferred from the prostasomes to sperms by a fusion process. In the present study, we investigated the molecular mechanism of action of APN and its role in regulating sperm functions and male fertility. In this in vitro study, epididymal mouse spermatozoa were incubated in a capacitating media (pH 7) containing 20 ng/mL of recombinant mouse APN for 90 min. Our results demonstrated that the supplementation of recombinant APN in sperm culture medium significantly increased APN activity, and subsequently altered motility, hyperactivated motility, rapid and medium swimming speeds, viability, and the acrosome reaction of mouse spermatozoa. These effects were potentially caused by increased toxicity in the spermatozoa. Further, altered APN activity in sperm culture medium affected early embryonic development. Interestingly, the effect of elevated APN activity in sperm culture medium was independent of protein tyrosine phosphorylation and protein kinase A activity. On the basis of these results, we concluded that APN plays a significant role in the regulation of several sperm functions and early embryonic development. In addition, increased APN activity could potentially lead to several adverse consequences related to male fertility. PMID:28859152

  20. Embryo selection using time-lapse analysis (Early Embryo Viability Assessment) in conjunction with standard morphology: a prospective two-center pilot study.

    PubMed

    Kieslinger, Dorit C; De Gheselle, Stefanie; Lambalk, Cornelis B; De Sutter, Petra; Kostelijk, E Hanna; Twisk, Jos W R; van Rijswijk, Joukje; Van den Abbeel, Etienne; Vergouw, Carlijn G

    2016-11-01

    Does prospective embryo selection using the results from the Eava Test (Early Embryo Viability Assessment) in combination with standard morphology increase the pregnancy rate of IVF and ICSI patients compared to embryo selection based on morphology only? Embryo selection using the Eeva Test plus standard morphology on Day 3 results in comparable pregnancy rates as conventional morphological embryo selection. Time-lapse monitoring of embryo development may represent a superior way to culture and select embryos in vitro. The Eeva Test records the development of each embryo with a cell-tracking system and predicts the likelihood (High, Medium or Low) that an embryo will form a blastocyst based on an automated analysis of early cell division timings. This trial was designed as a prospective, observational, two-center pilot study with a propensity matched control group. The analysis involved 280 of 302 enrolled patients who were included in the Eeva Test group in 2013 and 560 control patients who were treated in the years 2011-2013. The majority of transfers (98%) were single embryo transfers. Two academic hospitals (VUmc Amsterdam and UZ Gent) enrolled patients <41 years old, with <3 previous attempts and ≥5 normally fertilized eggs. Propensity matching was used to identify a propensity matched control group from a cohort of 1777 patients based on age, cycle number, oocyte number and number of fertilized oocytes. There was no difference in patient baseline characteristics between the two groups. The ongoing pregnancy rate (OPR) of patients enrolled in the Eeva Test group (34.3%; 96/280) did not differ significantly from the OPR in the propensity matched control group (34.6%, 194/560; P = 0.92). However, significantly less top quality embryos (eight-cell embryos with ≤25% fragmentation) were transferred in the Eeva Test group compared to the propensity matched control group (70.4% vs. 82.3%; P < 0.001). The transfer of Eeva High and Medium embryos resulted in a

  1. Central cell-derived peptides regulate early embryo patterning in flowering plants.

    PubMed

    Costa, Liliana M; Marshall, Eleanor; Tesfaye, Mesfin; Silverstein, Kevin A T; Mori, Masashi; Umetsu, Yoshitaka; Otterbach, Sophie L; Papareddy, Ranjith; Dickinson, Hugh G; Boutiller, Kim; VandenBosch, Kathryn A; Ohki, Shinya; Gutierrez-Marcos, José F

    2014-04-11

    Plant embryogenesis initiates with the establishment of an apical-basal axis; however, the molecular mechanisms accompanying this early event remain unclear. Here, we show that a small cysteine-rich peptide family is required for formation of the zygotic basal cell lineage and proembryo patterning in Arabidopsis. EMBRYO SURROUNDING FACTOR 1 (ESF1) peptides accumulate before fertilization in central cell gametes and thereafter in embryo-surrounding endosperm cells. Biochemical and structural analyses revealed cleavage of ESF1 propeptides to form biologically active mature peptides. Further, these peptides act in a non-cell-autonomous manner and synergistically with the receptor-like kinase SHORT SUSPENSOR to promote suspensor elongation through the YODA mitogen-activated protein kinase pathway. Our findings demonstrate that the second female gamete and its sexually derived endosperm regulate early embryonic patterning in flowering plants.

  2. Redundant roles of Sox17 and Sox18 in early cardiovascular development of mouse embryos

    SciT

    Sakamoto, Youhei; Hara, Kenshiro; Kanai-Azuma, Masami

    Sox7, -17 and -18 constitute the Sox subgroup F (SoxF) of HMG box transcription factor genes, which all are co-expressed in developing vascular endothelial cells in mice. Here we characterized cardiovascular phenotypes of Sox17/Sox18-double and Sox17-single null embryos during early-somite stages. Whole-mount PECAM staining demonstrated the aberrant heart looping, enlarged cardinal vein and mild defects in anterior dorsal aorta formation in Sox17 single-null embryos. The Sox17/Sox18 double-null embryos showed more severe defects in formation of anterior dorsal aorta and head/cervical microvasculature, and in some cases, aberrant differentiation of endocardial cells and defective fusion of the endocardial tube. However, the posteriormore » dorsal aorta and allantoic microvasculature was properly formed in all of the Sox17/Sox18 double-null embryos. The anomalies in both anterior dorsal aorta and head/cervical vasculature corresponded with the weak Sox7 expression sites. This suggests the region-specific redundant activities of three SoxF members along the anteroposterior axis of embryonic vascular network.« less

  3. Microspore embryogenesis in wheat: new marker genes for early, middle and late stages of embryo development.

    PubMed

    Sánchez-Díaz, Rosa Angélica; Castillo, Ana María; Vallés, María Pilar

    2013-09-01

    Microspore embryogenesis involves reprogramming of the pollen immature cell towards embryogenesis. We have identified and characterized a collection of 14 genes induced along different morphological phases of microspore-derived embryo development in wheat (Triticum aestivum L.) anther culture. SERKs and FLAs genes previously associated with somatic embryogenesis and reproductive tissues, respectively, were also included in this analysis. Genes involved in signalling mechanisms such as TaTPD1-like and TAA1b, and two glutathione S-transferase (GSTF2 and GSTA2) were induced when microspores had acquired a 'star-like' morphology or had undergone the first divisions. Genes associated with control of plant development and stress response (TaNF-YA, TaAGL14, TaFLA26, CHI3, XIP-R; Tad1 and WALI6) were activated before exine rupture. When the multicellular structures have been released from the exine, TaEXPB4, TaAGP31-like and an unknown embryo-specific gene TaME1 were induced. Comparison of gene expression, between two wheat cultivars with different response to anther culture, showed that the profile of genes activated before exine rupture was shifted to earlier stages in the low responding cultivar. This collection of genes constitutes a value resource for study mechanism of intra-embryo communication, early pattern formation, cell wall modification and embryo differentiation.

  4. DYZ1 copy number variation, Y chromosome polymorphism and early recurrent spontaneous abortion/early embryo growth arrest.

    PubMed

    Yan, Junhao; Fan, Lingling; Zhao, Yueran; You, Li; Wang, Laicheng; Zhao, Han; Li, Yuan; Chen, Zi-Jiang

    2011-12-01

    To find the association between recurrent spontaneous abortion (RSA)/early embryo growth arrest and Y chromosome polymorphism. Peripheral blood samples of the male patients of big Y chromosome, small Y chromosome and other male patients whose partners suffered from unexplained RSA/early embryo growth arrest were collected. PCR and real-time fluorescent quantitative PCR were used to test the deletion and the copy number variation of DYZ1 region in Y chromosome of the patients. A total of 79 big Y chromosome patients (48 of whose partners suffered from RSA or early embryo growth arrest), 7 small Y chromosome patients, 106 other male patients whose partners had suffered from unexplained RSA or early embryo growth arrest, and 100 normal male controls were enrolled. There was no fraction deletion of DYZ1 detected both in big Y patients and in normal men. Of RSA patients, 1 case showed deletion of 266bp from the gene locus 25-290bp, and 2 cases showed deletion of 773bp from 1347 to 2119bp. Of only 7 small Y chromosome patients, 2 cases showed deletion of 266bp from 25 to 290bp, and 4 cases showed deletion of 773bp from 1347 to 2119bp and 275bp from 3128 to 3420bp. The mean of DYZ1 copies was 3900 in normal control men; the mean in big Y patients was 5571, in RSA patients was 2655, and in small Y patients was 1059. All of the others were significantly different (P<0.01) compared with normal control men, which meant that DYZ1 copy number in normal control men was less than that of big Y chromosome patients, and was more than that of unexplained early RSA patients and small Y patients. The integrity and copy number variation of DYZ1 are closely related to the Y chromosome length under microscope. The cause of RSA/early embryo growth arrest in some couples may be the increase (big Y patients) or decrease of DYZ1 copy number in the husbands' Y chromosome. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  5. Chicken interferome: avian interferon-stimulated genes identified by microarray and RNA-seq of primary chick embryo fibroblasts treated with a chicken type I interferon (IFN-α).

    PubMed

    Giotis, Efstathios S; Robey, Rebecca C; Skinner, Natalie G; Tomlinson, Christopher D; Goodbourn, Stephen; Skinner, Michael A

    2016-08-05

    Viruses that infect birds pose major threats-to the global supply of chicken, the major, universally-acceptable meat, and as zoonotic agents (e.g. avian influenza viruses H5N1 and H7N9). Controlling these viruses in birds as well as understanding their emergence into, and transmission amongst, humans will require considerable ingenuity and understanding of how different species defend themselves. The type I interferon-coordinated response constitutes the major antiviral innate defence. Although interferon was discovered in chicken cells, details of the response, particularly the identity of hundreds of stimulated genes, are far better described in mammals. Viruses induce interferon-stimulated genes but they also regulate the expression of many hundreds of cellular metabolic and structural genes to facilitate their replication. This study focusses on the potentially anti-viral genes by identifying those induced just by interferon in primary chick embryo fibroblasts. Three transcriptomic technologies were exploited: RNA-seq, a classical 3'-biased chicken microarray and a high density, "sense target", whole transcriptome chicken microarray, with each recognising 120-150 regulated genes (curated for duplication and incorrect assignment of some microarray probesets). Overall, the results are considered robust because 128 of the compiled, curated list of 193 regulated genes were detected by two, or more, of the technologies.

  6. Gc protein-derived macrophage-activating factor (GcMAF) stimulates cAMP formation in human mononuclear cells and inhibits angiogenesis in chick embryo chorionallantoic membrane assay.

    PubMed

    Pacini, Stefania; Morucci, Gabriele; Punzi, Tiziana; Gulisano, Massimo; Ruggiero, Marco

    2011-04-01

    The effects of Gc protein-derived macrophage-activating factor (GcMAF) have been studied in cancer and other conditions where angiogenesis is deregulated. In this study, we demonstrate for the first time that the mitogenic response of human peripheral blood mononuclear cells (PBMCs) to GcMAF was associated with 3'-5'-cyclic adenosine monophosphate (cAMP) formation. The effect was dose dependent, and maximal stimulation was achieved using 0.1 ng/ml. Heparin inhibited the stimulatory effect of GcMAF on PBMCs. In addition, we demonstrate that GcMAF (1 ng/ml) inhibited prostaglandin E(1)- and human breast cancer cell-stimulated angiogenesis in chick embryo chorionallantoic membrane (CAM) assay. Finally, we tested different GcMAF preparations on CAM, and the assay proved to be a reliable, reproducible and inexpensive method to determine the relative potencies of different preparations and their stability; we observed that storage at room temperature for 15 days decreased GcMAF potency by about 50%. These data could prove useful for upcoming clinical trials on GcMAF.

  7. A randomized open-labeled study to demonstrate the non-inferiority of purified chick-embryo cell rabies vaccine administered in the Zagreb regimen (2-1-1) compared with the Essen regimen in Chinese adults.

    PubMed

    Ma, Jingchen; Wang, Hongchang; Li, Jun; Chang, Likuan; Xie, Yun; Liu, Zhonglin; Zhao, Yuliang; Malerczyk, Claudius; Claudius, Malerczyk

    2014-01-01

    The Zagreb regimen has been used for 20 years in various countries. In China, until 2010, the Zagreb schedule was only approved for purified chick embryo cell vaccine (PCECV) and purified Vero cell rabies vaccines (PVRV). In this phase III clinical trial, we aimed to demonstrate the safety and immunogenic non-inferiority of the Zagreb regimen compared with the Essen regimen in healthy adult Chinese immunized with PCECV (Rabipur®). The study enrolled 825 subjects aged 18 to 50 years; serum samples were collected on Days 0, 7, 14, 42, and at 13 months to assess rabies virus neutralizing antibody (RVNA) concentrations. Solicited and unsolicited local and systemic reactions were recorded for 6 days following the day of vaccination, and collected throughout the entire study period (Day 1 until Month 13). The Zagreb regimen was non-inferior to the Essen regimen with regard to RVNA concentrations after 7, 14, and 42 days, and 13 months of immunization. The non-inferiority of seroconversion was established at Days 14 and 42. The incidence of local and systemic reactions was similar between groups, and mostly of mild or moderate severity. Vaccine-related adverse events occurred more frequently in the Essen group than in the Zagreb group. Vaccination with PCECV under a 2-1-1 regimen is as safe and immunogenic as under the traditional 5-dose Essen regimen for rabies post-exposure prophylaxis, and is a more cost-effective option, has a more practical vaccination schedule, and can potentially increase compliance.

  8. Shorter Exposures to Harder X-Rays Trigger Early Apoptotic Events in Xenopus laevis Embryos

    PubMed Central

    Dong, JiaJia; Mury, Sean P.; Drahos, Karen E.; Moscovitch, Marko

    2010-01-01

    Background A long-standing conventional view of radiation-induced apoptosis is that increased exposure results in augmented apoptosis in a biological system, with a threshold below which radiation doses do not cause any significant increase in cell death. The consequences of this belief impact the extent to which malignant diseases and non-malignant conditions are therapeutically treated and how radiation is used in combination with other therapies. Our research challenges the current dogma of dose-dependent induction of apoptosis and establishes a new parallel paradigm to the photoelectric effect in biological systems. Methodology/Principal Findings We explored how the energy of individual X-ray photons and exposure time, both factors that determine the total dose, influence the occurrence of cell death in early Xenopus embryo. Three different experimental scenarios were analyzed and morphological and biochemical hallmarks of apoptosis were evaluated. Initially, we examined cell death events in embryos exposed to increasing incident energies when the exposure time was preset. Then, we evaluated the embryo's response when the exposure time was augmented while the energy value remained constant. Lastly, we studied the incidence of apoptosis in embryos exposed to an equal total dose of radiation that resulted from increasing the incoming energy while lowering the exposure time. Conclusions/Significance Overall, our data establish that the energy of the incident photon is a major contributor to the outcome of the biological system. In particular, for embryos exposed under identical conditions and delivered the same absorbed dose of radiation, the response is significantly increased when shorter bursts of more energetic photons are used. These results suggest that biological organisms display properties similar to the photoelectric effect in physical systems and provide new insights into how radiation-mediated apoptosis should be understood and utilized for therapeutic

  9. Are Early Somatic Embryos of the Norway Spruce (Picea abies (L.) Karst.) Organised?

    PubMed Central

    Petrek, Jiri; Zitka, Ondrej; Adam, Vojtech; Bartusek, Karel; Anjum, Naser A.; Pereira, Eduarda; Havel, Ladislav; Kizek, Rene

    2015-01-01

    Background Somatic embryogenesis in conifer species has great potential for the forestry industry. Hence, a number of methods have been developed for their efficient and rapid propagation through somatic embryogenesis. Although information is available regarding the previous process-mediated generation of embryogenic cells to form somatic embryos, there is a dearth of information in the literature on the detailed structure of these clusters. Methodology/Principal Findings The main aim of this study was to provide a more detailed structure of the embryogenic tissue clusters obtained through the in vitro propagation of the Norway spruce (Picea abies (L.) Karst.). We primarily focused on the growth of early somatic embryos (ESEs). The data on ESE growth suggested that there may be clear distinctions between their inner and outer regions. Therefore, we selected ESEs collected on the 56th day after sub-cultivation to dissect the homogeneity of the ESE clusters. Two colourimetric assays (acetocarmine and fluorescein diacetate/propidium iodide staining) and one metabolic assay based on the use of 2,3,5-triphenyltetrazolium chloride uncovered large differences in the metabolic activity inside the cluster. Next, we performed nuclear magnetic resonance measurements. The ESE cluster seemed to be compactly aggregated during the first four weeks of cultivation; thereafter, the difference between the 1H nuclei concentration in the inner and outer clusters was more evident. There were clear differences in the visual appearance of embryos from the outer and inner regions. Finally, a cluster was divided into six parts (three each from the inner and the outer regions of the embryo) to determine their growth and viability. The innermost embryos (centripetally towards the cluster centre) could grow after sub-cultivation but exhibited the slowest rate and required the longest time to reach the common growth rate. To confirm our hypothesis on the organisation of the ESE cluster, we

  10. The first whole transcriptomic exploration of pre-oviposited early chicken embryos using single and bulked embryonic RNA-sequencing.

    PubMed

    Hwang, Young Sun; Seo, Minseok; Choi, Hee Jung; Kim, Sang Kyung; Kim, Heebal; Han, Jae Yong

    2018-04-01

    The chicken is a valuable model organism, especially in evolutionary and embryology research because its embryonic development occurs in the egg. However, despite its scientific importance, no transcriptome data have been generated for deciphering the early developmental stages of the chicken because of practical and technical constraints in accessing pre-oviposited embryos. Here, we determine the entire transcriptome of pre-oviposited avian embryos, including oocyte, zygote, and intrauterine embryos from Eyal-giladi and Kochav stage I (EGK.I) to EGK.X collected using a noninvasive approach for the first time. We also compare RNA-sequencing data obtained using a bulked embryo sequencing and single embryo/cell sequencing technique. The raw sequencing data were preprocessed with two genome builds, Galgal4 and Galgal5, and the expression of 17,108 and 26,102 genes was quantified in the respective builds. There were some differences between the two techniques, as well as between the two genome builds, and these were affected by the emergence of long intergenic noncoding RNA annotations. The first transcriptome datasets of pre-oviposited early chicken embryos based on bulked and single embryo sequencing techniques will serve as a valuable resource for investigating early avian embryogenesis, for comparative studies among vertebrates, and for novel gene annotation in the chicken genome.

  11. Long-term primary culture of neurons taken from chick embryo brain: A model to study neural cell biology, synaptogenesis and its dynamic properties.

    PubMed

    Kumar, Awanish; Mallick, Birendra Nath

    2016-04-01

    Studying neuronal growth, development and synaptogenesis are among the hot research topics. However, it is faced with various challenges and technical limitations that include but not limited to donor's species and health, threat to life, age of embryo, glial contamination, real-time tracking, and follow-up. We have successfully standardized a method for long-term primary culture of neurons collected from post-fertilized 9 day incubated chicken embryo brain overcoming the limitations mentioned above. Fertilized eggs were incubated in the laboratory and neurons from the embryonic brain were collected and low-density culture, apparently without glial contamination, was studied at least for 35 days in vitro (DIV). Neurons were characterized by double immunostaining using stringent neuronal and glial markers. Neuronal differentiation, cytomorphology, neurite and axon formation, development and maturation, spine formation and synaptogenesis were tracked in real-time in a stage and time dependent manner. The neurons were transfected with Synaptophysin-RFP to label synaptic vesicles, which were followed in real-time under live-cell imaging. Every step was carried out under controlled laboratory conditions. Eggs are easily available, easy to handle, neurons from desired day of incubation could be conveniently studied for long period in apparently glia-free condition. In addition to common factors affecting primary culture, selection of culture media and cover glass coating are other key factors affecting neuronal cultures. We describe an inexpensive, simpler pure primary neuronal culture method for studying neuronal cell-biology, synaptogenesis, vesicular dynamics and it has potential to grow 3D-multilayered brain in vitro. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Moral uncertainty in bioethical argumentation: a new understanding of the pro-life view on early human embryos.

    PubMed

    Żuradzki, Tomasz

    2014-12-01

    In this article, I present a new interpretation of the pro-life view on the status of early human embryos. In my understanding, this position is based not on presumptions about the ontological status of embryos and their developmental capabilities but on the specific criteria of rational decisions under uncertainty and on a cautious response to the ambiguous status of embryos. This view, which uses the decision theory model of moral reasoning, promises to reconcile the uncertainty about the ontological status of embryos with the certainty about normative obligations. I will demonstrate that my interpretation of the pro-life view, although seeming to be stronger than the standard one, has limited scope and cannot be used to limit destructive research on human embryos.

  13. Early developmental gene regulation in Strongylocentrotus purpuratus embryos in response to elevated CO₂ seawater conditions.

    PubMed

    Hammond, LaTisha M; Hofmann, Gretchen E

    2012-07-15

    Ocean acidification, or the increased uptake of CO(2) by the ocean due to elevated atmospheric CO(2) concentrations, may variably impact marine early life history stages, as they may be especially susceptible to changes in ocean chemistry. Investigating the regulatory mechanisms of early development in an environmental context, or ecological development, will contribute to increased understanding of potential organismal responses to such rapid, large-scale environmental changes. We examined transcript-level responses to elevated seawater CO(2) during gastrulation and the initiation of spiculogenesis, two crucial developmental processes in the purple sea urchin, Strongylocentrotus purpuratus. Embryos were reared at the current, accepted oceanic CO(2) concentration of 380 microatmospheres (μatm), and at the elevated levels of 1000 and 1350 μatm, simulating predictions for oceans and upwelling regions, respectively. The seven genes of interest comprised a subset of pathways in the primary mesenchyme cell gene regulatory network (PMC GRN) shown to be necessary for the regulation and execution of gastrulation and spiculogenesis. Of the seven genes, qPCR analysis indicated that elevated CO(2) concentrations only had a significant but subtle effect on two genes, one important for early embryo patterning, Wnt8, and the other an integral component in spiculogenesis and biomineralization, SM30b. Protein levels of another spicule matrix component, SM50, demonstrated significant variable responses to elevated CO(2). These data link the regulation of crucial early developmental processes with the environment that these embryos would be developing within, situating the study of organismal responses to ocean acidification in a developmental context.

  14. Endogenous Nod-Factor-Like Signal Molecules Promote Early Somatic Embryo Development in Norway Spruce1

    PubMed Central

    Dyachok, Julia V.; Wiweger, Malgorzata; Kenne, Lennart; von Arnold, Sara

    2002-01-01

    Embryogenic cultures of Norway spruce (Picea abies) are composed of pro-embryogenic masses (PEMs) and somatic embryos of various developmental stages. Auxin is important for PEM formation and proliferation. In this report we show that depletion of auxin blocks PEM development and causes large-scale cell death. Extracts of the media conditioned by embryogenic cultures stimulate development of PEM aggregates in auxin-deficient cultures. Partial characterization of the conditioning factor shows that it is a lipophilic, low-molecular-weight molecule, which is sensitive to chitinase and contains GlcNAc residues. On the basis of this information, we propose that the factor is a lipophilic chitin oligosaccharide (LCO). The amount of LCO correlates to the developmental stages of PEMs and embryos, with the highest level in the media conditioned by developmentally blocked cultures. LCO is not present in nonembryogenic cultures. Cell death, induced by withdrawal of auxin, is suppressed by extra supply of endogenous LCO or Nod factor from Rhizobium sp. NGR234. The effect can be mimicked by a chitotetraose or chitinase from Streptomyces griseus. Taken together, our data suggest that endogenous LCO acts as a signal molecule stimulating PEM and early embryo development in Norway spruce. PMID:11842156

  15. The effects of the early uterine environment on the subsequent development of embryo and fetus.

    PubMed

    Barnes, F L

    2000-01-15

    Synchrony between the embryo and the uterine endometrium is essential for the establishment of pregnancy and birth in people and livestock. When asynchronous conditions occur a variety of complication result that include failure of the embryo to implant, early embryonic mortality, retarded development and growth, and accelerated development and growth. These complications all appear to be induced within the first week of embryo development and not withstanding the immediate endpoint of large or small size at birth, may alter the course of development throughout the life of the animal. Progesterone appears to play a causative role in establishing the abnormal growth of the fetus by decelerating or accelerating embryonic development. This may act through increasing the transport of blood born growth factors into the uterine lumen or by stimulating the release of growth factors from the endometrium directly. It can not be ruled out that progesterone mediated abundance of, or absence of, appropriate nutrition may bring about the same lifelong outcome. In vitro culture situations that include serum and/or co-culture can also bring about these abnormalities of growth. It is hypothesized that exposure to growth factors "out of phase" may result in an irreversible induction of abnormal development. The described abnormalities that occur in sheep and cattle have not yet been described for children resulting from IVF.

  16. Studies on trypsin-like enzymes in sperm and early embryos

    SciT

    Penn, A.

    1975-12-09

    Results are reported from a study of acrosomal proteinase, a trypsin-like enzyme (TLE), found in the acrosome of all eutherian mammals studied to date. It has been implicated in the dissolution of a passage for the sperm through the zona pellucida of the egg, a step necessary for in vivo fertilization. A cytochemical procedure employing autoradiographic film as a gelatin substrate is described for in situ detection and localization of acrosomal proteolytic activity. A role for TLE in the early development of embryos is suggested. (CH)

  17. Somatic mutations reveal asymmetric cellular dynamics in the early human embryo

    DOE PAGES

    Ju, Young Seok; Martincorena, Inigo; Gerstung, Moritz; ...

    2017-03-22

    Somatic cells acquire mutations throughout the course of an individual’s life. Mutations occurring early in embryogenesis are often present in a substantial proportion of, but not all, cells in postnatal humans and thus have particular characteristics and effects. Depending on their location in the genome and the proportion of cells they are present in, these mosaic mutations can cause a wide range of genetic disease syndromes and predispose carriers to cancer. They have a high chance of being transmitted to offspring as de novo germline mutations and, in principle, can provide insights into early human embryonic cell lineages and theirmore » contributions to adult tissues. Although it is known that gross chromosomal abnormalities are remarkably common in early human embryos, our understanding of early embryonic somatic mutations is very limited. Here we use whole-genome sequences of normal blood from 241 adults to identify 163 early embryonic mutations. We estimate that approximately three base substitution mutations occur per cell per cell-doubling event in early human embryogenesis and these are mainly attributable to two known mutational signatures. We used the mutations to reconstruct developmental lineages of adult cells and demonstrate that the two daughter cells of many early embryonic cell-doubling events contribute asymmetrically to adult blood at an approximately 2:1 ratio. As a result, this study therefore provides insights into the mutation rates, mutational processes and developmental outcomes of cell dynamics that operate during early human embryogenesis.« less

  18. Somatic mutations reveal asymmetric cellular dynamics in the early human embryo

    SciT

    Ju, Young Seok; Martincorena, Inigo; Gerstung, Moritz

    Somatic cells acquire mutations throughout the course of an individual’s life. Mutations occurring early in embryogenesis are often present in a substantial proportion of, but not all, cells in postnatal humans and thus have particular characteristics and effects. Depending on their location in the genome and the proportion of cells they are present in, these mosaic mutations can cause a wide range of genetic disease syndromes and predispose carriers to cancer. They have a high chance of being transmitted to offspring as de novo germline mutations and, in principle, can provide insights into early human embryonic cell lineages and theirmore » contributions to adult tissues. Although it is known that gross chromosomal abnormalities are remarkably common in early human embryos, our understanding of early embryonic somatic mutations is very limited. Here we use whole-genome sequences of normal blood from 241 adults to identify 163 early embryonic mutations. We estimate that approximately three base substitution mutations occur per cell per cell-doubling event in early human embryogenesis and these are mainly attributable to two known mutational signatures. We used the mutations to reconstruct developmental lineages of adult cells and demonstrate that the two daughter cells of many early embryonic cell-doubling events contribute asymmetrically to adult blood at an approximately 2:1 ratio. As a result, this study therefore provides insights into the mutation rates, mutational processes and developmental outcomes of cell dynamics that operate during early human embryogenesis.« less

  19. BMP inhibition by DAN in Hensen's node is a critical step for the establishment of left-right asymmetry in the chick embryo.

    PubMed

    Katsu, Kenjiro; Tokumori, Daisuke; Tatsumi, Norifumi; Suzuki, Atsushi; Yokouchi, Yuji

    2012-03-01

    During left-right (L-R) axis formation, Nodal is expressed in the node and has a central role in the transfer of L-R information in the vertebrate embryo. Bone morphogenetic protein (BMP) signaling also has an important role for maintenance of gene expression around the node. Several members of the Cerberus/Dan family act on L-R patterning by regulating activity of the transforming growth factor-β (TGF-β) family. We demonstrate here that chicken Dan plays a critical role in L-R axis formation. Chicken Dan is expressed in the left side of the node shortly after left-handed Shh expression and before the appearance of asymmetrically expressed genes in the lateral plate mesoderm (LPM). In vitro experiments revealed that DAN inhibited BMP signaling but not NODAL signaling. SHH had a positive regulatory effect on Dan expression while BMP4 had a negative effect. Using overexpression and RNA interference-mediated knockdown strategies, we demonstrate that Dan is indispensable for Nodal expression in the LPM and for Lefty-1 expression in the notochord. In the perinodal region, expression of Dan and Nodal was independent of each other. Nodal up-regulation by DAN required NODAL signaling, suggesting that DAN might act synergistically with NODAL. Our data indicate that Dan plays an essential role in the establishment of the L-R axis by inhibiting BMP signaling around the node. Copyright © 2012. Published by Elsevier Inc.

  20. Transcriptome analyses of rhesus monkey preimplantation embryos reveal a reduced capacity for DNA double-strand break repair in primate oocytes and early embryos

    PubMed Central

    Wang, Xinyi; Liu, Denghui; He, Dajian; Suo, Shengbao; Xia, Xian; He, Xiechao; Han, Jing-Dong J.; Zheng, Ping

    2017-01-01

    Preimplantation embryogenesis encompasses several critical events including genome reprogramming, zygotic genome activation (ZGA), and cell-fate commitment. The molecular basis of these processes remains obscure in primates in which there is a high rate of embryo wastage. Thus, understanding the factors involved in genome reprogramming and ZGA might help reproductive success during this susceptible period of early development and generate induced pluripotent stem cells with greater efficiency. Moreover, explaining the molecular basis responsible for embryo wastage in primates will greatly expand our knowledge of species evolution. By using RNA-seq in single and pooled oocytes and embryos, we defined the transcriptome throughout preimplantation development in rhesus monkey. In comparison to archival human and mouse data, we found that the transcriptome dynamics of monkey oocytes and embryos were very similar to those of human but very different from those of mouse. We identified several classes of maternal and zygotic genes, whose expression peaks were highly correlated with the time frames of genome reprogramming, ZGA, and cell-fate commitment, respectively. Importantly, comparison of the ZGA-related network modules among the three species revealed less robust surveillance of genomic instability in primate oocytes and embryos than in rodents, particularly in the pathways of DNA damage signaling and homology-directed DNA double-strand break repair. This study highlights the utility of monkey models to better understand the molecular basis for genome reprogramming, ZGA, and genomic stability surveillance in human early embryogenesis and may provide insights for improved homologous recombination-mediated gene editing in monkey. PMID:28223401

  1. Supplementation with CTGF, SDF1, NGF, and HGF promotes ovine in vitro oocyte maturation and early embryo development.

    PubMed

    Wang, D H; Ren, J; Zhou, C J; Han, Z; Wang, L; Liang, C G

    2018-05-17

    The strategies for improving the in vitro maturation (IVM) of domestic animal oocytes focus on promoting nuclear and cytoplasmic maturation. The identification of paracrine factors and their supplementation in the culture medium represent effective approaches for oocyte maturation and embryo development. This study investigated the effects of paracrine factor supplementation including connective tissue growth factor (CTGF), nerve growth factor (NGF), hepatocyte growth factor (HGF), and stromal derived factor 1 (SDF1) on ovine oocytes and early parthenogenetic embryos using an in vitro culture system. First, we identified the optimal concentrations of CTGF (30 ng/mL), SDF1 (10 ng/mL), NGF (3 ng/mL), and HGF (100 ng/mL) for promoting oocyte maturation, which combined, induced nuclear maturation in 94.19% of oocytes. This combination also promoted cumulus cell expansion and inhibited oocyte/cumulus apoptosis, while enabling a larger proportion (33.04%) of embryos to develop into blastocysts than in the controls and prevented embryo apoptosis. These novel findings demonstrate that the paracrine factors CTGF, SDF1, NGF, and HGF facilitate ovine oocyte and early parthenogenetic embryo development in vitro. Thus, supplementation with these factors may help optimize the IVM of ovine oocytes and early parthenogenetic embryo development strategies. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. A randomized open-labeled study to demonstrate the non-inferiority of purified chick-embryo cell rabies vaccine administered in the Zagreb regimen (2-1-1) compared with the Essen regimen in Chinese adults

    PubMed Central

    Ma, Jingchen; Wang, Hongchang; Li, Jun; Chang, Likuan; Xie, Yun; Liu, Zhonglin; Zhao, Yuliang; Claudius, Malerczyk

    2014-01-01

    The Zagreb regimen has been used for 20 years in various countries. In China, until 2010, the Zagreb schedule was only approved for purified chick embryo cell vaccine (PCECV) and purified Vero cell rabies vaccines (PVRV). In this phase III clinical trial, we aimed to demonstrate the safety and immunogenic non-inferiority of the Zagreb regimen compared with the Essen regimen in healthy adult Chinese immunized with PCECV (Rabipur®). The study enrolled 825 subjects aged 18 to 50 years; serum samples were collected on Days 0, 7, 14, 42, and at 13 months to assess rabies virus neutralizing antibody (RVNA) concentrations. Solicited and unsolicited local and systemic reactions were recorded for 6 days following the day of vaccination, and collected throughout the entire study period (Day 1 until Month 13). The Zagreb regimen was non-inferior to the Essen regimen with regard to RVNA concentrations after 7, 14, and 42 days, and 13 months of immunization. The non-inferiority of seroconversion was established at Days 14 and 42. The incidence of local and systemic reactions was similar between groups, and mostly of mild or moderate severity. Vaccine-related adverse events occurred more frequently in the Essen group than in the Zagreb group. Vaccination with PCECV under a 2-1-1 regimen is as safe and immunogenic as under the traditional 5-dose Essen regimen for rabies post-exposure prophylaxis, and is a more cost-effective option, has a more practical vaccination schedule, and can potentially increase compliance. PMID:25483635

  3. Comparative study on the immunogenicity and safety of a purified chick embryo cell rabies vaccine (PCECV) administered according to two different simulated post exposure intramuscular regimens (Zagreb versus Essen)

    PubMed Central

    Mahendra, BJ; Narayana, DH Ashwath; Agarkhedkar, Sharad; Ravish, HS; Harish, BR; Agarkhedkar, Shalaka; Madhusudana, SN; Belludi, Ashwin; Ahmed, Khaleel; Jonnalagedda, Rekha; Vakil, Hoshang; Bhusal, Chiranjiwi; Arora, Ashwani Kumar

    2015-01-01

    Despite availability of effective rabies vaccines, India has the highest global mortality rate for rabies. Low socio-economic communities are most affected due to lack of awareness of the disease and poor compliance to post-exposure prophylactic regimens. Currently, the only approved intramuscular regimen for post-exposure prophylaxis (PEP) against rabies in India is the Essen regimen, which consists of 5 injections administered over 5 separate days in a period of one month. The high number of doses and clinical visits, however, are major reasons for non-compliance, and thus a shorter regimen would be beneficial. In a simulated PEP trial in healthy, adult subjects, this study evaluated whether purified chick embryo cell vaccine (PCECV), administered according to the WHO-recommended 4-dose/3 visit Zagreb vaccination regimen is of equal immunogenicity and safety as the standard Essen regimen in Indian subjects. Two hundred and 50 healthy adults were enrolled and randomized into a Zagreb or Essen group, each receiving PCECV according to their respective regimen. Blood samples were collected on Days 0, 7, 14 and 42 and analyzed using the rapid fluorescent focus inhibition test (RFFIT). By Day 14, all subjects across both groups attained rabies virus neutralizing antibody (RVNA) concentrations of ≥ 0.5IU/ml. The Zagreb regimen was then demonstrated to be immunologically non-inferior to the Essen regimen by Day 14, which was the primary endpoint of the study. No safety issues were noted and the occurrence of adverse events was similar in both groups (17% and 15%, respectively). NCT01365494. CTRI No.: CTRI/2011/07/001857 PMID:25692792

  4. Comparative study on the immunogenicity and safety of a purified chick embryo cell rabies vaccine (PCECV) administered according to two different simulated post exposure intramuscular regimens (Zagreb versus Essen).

    PubMed

    Mahendra, B J; Narayana, Dh Ashwath; Agarkhedkar, Sharad; Ravish, H S; Harish, B R; Agarkhedkar, Shalaka; Madhusudana, S N; Belludi, Ashwin; Ahmed, Khaleel; Jonnalagedda, Rekha; Vakil, Hoshang; Bhusal, Chiranjiwi; Arora, Ashwani Kumar

    2015-01-01

    Despite availability of effective rabies vaccines, India has the highest global mortality rate for rabies. Low socio-economic communities are most affected due to lack of awareness of the disease and poor compliance to post-exposure prophylactic regimens. Currently, the only approved intramuscular regimen for post-exposure prophylaxis (PEP) against rabies in India is the Essen regimen, which consists of 5 injections administered over 5 separate days in a period of one month. The high number of doses and clinical visits, however, are major reasons for non-compliance, and thus a shorter regimen would be beneficial. In a simulated PEP trial in healthy, adult subjects, this study evaluated whether purified chick embryo cell vaccine (PCECV), administered according to the WHO-recommended 4-dose/3 visit Zagreb vaccination regimen is of equal immunogenicity and safety as the standard Essen regimen in Indian subjects. Two hundred and 50 healthy adults were enrolled and randomized into a Zagreb or Essen group, each receiving PCECV according to their respective regimen. Blood samples were collected on Days 0, 7, 14 and 42 and analyzed using the rapid fluorescent focus inhibition test (RFFIT). By Day 14, all subjects across both groups attained rabies virus neutralizing antibody (RVNA) concentrations of ≥ 0.5IU/ml. The Zagreb regimen was then demonstrated to be immunologically non-inferior to the Essen regimen by Day 14, which was the primary endpoint of the study. No safety issues were noted and the occurrence of adverse events was similar in both groups (17% and 15%, respectively). NCT01365494. CTRI No.: CTRI/2011/07/001857.

  5. Effects of early neonatal development and delayed feeding immediately post-hatch on the hepatic lipogenic program in broiler chicks.

    PubMed

    Richards, Mark P; Proszkowiec-Weglarz, Monika; Rosebrough, Robert W; McMurtry, John P; Angel, Roselina

    2010-12-01

    The embryo to neonate transition is a critical period of development that has significant impact on broiler production. During this time important genetic programs governing metabolism and growth are established. The goal of this work was to study the effects of early post-hatch (PH) development and the time of initiation of feeding on activation of the genetic program regulating hepatic lipogenesis. A comparison of liver total RNA samples at hatch and 7 days PH was performed using oligonucleotide-based (Affymetrix GeneChip®) chicken genome microarrays. During the first week PH there was significant up-regulation of key lipogenic genes including: ATP citrate lyase (ACL), malic enzyme (ME), fatty acid synthase (FAS), acetyl-CoA carboxylase alpha (ACCα), stearoyl-CoA desaturase-1 (SCD-1), sterol regulatory element binding protein-2 (SREBP-2) and thyroid hormone responsive spot 14α (Spot 14α) among others. These findings were confirmed using gene-specific RT-PCR assays. In a follow-up study, we investigated the effects of withholding feed for the first 48 h PH (delayed feeding, DF) on lipogenic gene expression through 8 days PH. Body weight gain was significantly depressed by DF. Plasma levels of the major metabolic hormones that regulate lipogenic gene expression (insulin, glucagon and T(3)) changed significantly during PH development, but were largely unaffected by DF. Plasma glucose was significantly lower in the DF group at 24h PH but recovered thereafter. In general, DF inhibited the up-regulation of lipogenic genes until feeding was initiated. Delayed up-regulation was also observed for the lipogenic transcription factor genes, SREBP-1, SREBP-2 and peroxisome proliferator-activated receptor gamma (PPARγ), but not for carbohydrate response element binding protein (ChREB) or liver X receptor (LXR). Our results offer additional insight into the transcriptional programming of hepatic lipogenesis in response to the transition from high fat (yolk) to high

  6. Transcriptome Analysis of Honeybee (Apis Mellifera) Haploid and Diploid Embryos Reveals Early Zygotic Transcription during Cleavage

    PubMed Central

    Pires, Camilla Valente; Freitas, Flávia Cristina de Paula; Cristino, Alexandre S.; Dearden, Peter K.; Simões, Zilá Luz Paulino

    2016-01-01

    In honeybees, the haplodiploid sex determination system promotes a unique embryogenesis process wherein females develop from fertilized eggs and males develop from unfertilized eggs. However, the developmental strategies of honeybees during early embryogenesis are virtually unknown. Similar to most animals, the honeybee oocytes are supplied with proteins and regulatory elements that support early embryogenesis. As the embryo develops, the zygotic genome is activated and zygotic products gradually replace the preloaded maternal material. The analysis of small RNA and mRNA libraries of mature oocytes and embryos originated from fertilized and unfertilized eggs has allowed us to explore the gene expression dynamics in the first steps of development and during the maternal-to-zygotic transition (MZT). We localized a short sequence motif identified as TAGteam motif and hypothesized to play a similar role in honeybees as in fruit flies, which includes the timing of early zygotic expression (MZT), a function sustained by the presence of the zelda ortholog, which is the main regulator of genome activation. Predicted microRNA (miRNA)-target interactions indicated that there were specific regulators of haploid and diploid embryonic development and an overlap of maternal and zygotic gene expression during the early steps of embryogenesis. Although a number of functions are highly conserved during the early steps of honeybee embryogenesis, the results showed that zygotic genome activation occurs earlier in honeybees than in Drosophila based on the presence of three primary miRNAs (pri-miRNAs) (ame-mir-375, ame-mir-34 and ame-mir-263b) during the cleavage stage in haploid and diploid embryonic development. PMID:26751956

  7. Effect of exogenous transforming growth factor β1 (TGF-β1) on early bovine embryo development.

    PubMed

    Barrera, Antonio D; García, Elina V; Miceli, Dora C

    2018-06-08

    SummaryDuring preimplantation development, embryos are exposed and have the capacity to respond to different growth factors present in the maternal environment. Among these factors, transforming growth factor β1 (TGF-β1) is a well known modulator of embryonic growth and development. However, its action during the first stages of development, when the embryo transits through the oviduct, has not been yet elucidated. The objective of the present study was to examine the effect of early exposure to exogenous TGF-β1 on embryo development and expression of pluripotency (OCT4, NANOG) and DNA methylation (DNMT1, DNMT3A, DNMT3B) genes in bovine embryos produced in vitro. First, gene expression analysis of TGF-β receptors confirmed a stage-specific expression pattern, showing greater mRNA abundance of TGFBR1 and TGFBR2 from the 2- to the 8-cell stage, before embryonic genome activation. Second, embryo culture for the first 48 h in serum-free CR1aa medium supplemented with 50 or 100 ng/ml recombinant TGF-β1 did not affect the cleavage and blastocyst rate (days 7 and 8). However, RT-qPCR analysis showed a significant increase in the relative abundance of NANOG and DNMT3A in the 8-cell stage embryos and expanded blastocysts (day 8) derived from TGF-β1 treated embryos. These results suggest an early action of exogenous TGF-β1 on the bovine embryo, highlighting the importance to provide a more comprehensive understanding of the role of TGF-β signalling during early embryogenesis.

  8. Bovine oocytes and early embryos express Staufen and ELAVL RNA-binding proteins.

    PubMed

    Calder, M D; Madan, P; Watson, A J

    2008-05-01

    RNA-binding proteins (RBP) influence RNA editing, localization, stability and translation and may contribute to oocyte developmental competence by regulating the stability and turnover of oogenetic mRNAs. The expression of Staufen 1 and 2 and ELAVL1, ELAVL2 RNA-binding proteins during cow early development was characterized. Cumulus-oocyte complexes were collected from slaughterhouse ovaries, matured, inseminated and subjected to embryo culture in vitro. Oocyte or preimplantation embryo pools were processed for RT-PCR and whole-mount immunofluorescence analysis of mRNA expression and protein distribution. STAU1 and STAU2 and ELAVL1 mRNAs and proteins were detected throughout cow preimplantation development from the germinal vesicle (GV) oocyte to the blastocyst stage. ELAVL2 mRNAs were detectable from the GV to the morula stage, whereas ELAVL2 protein was in all stages examined and localized to both cytoplasm and nuclei. The findings provide a foundation for investigating the role of RBPs during mammalian oocyte maturation and early embryogenesis.

  9. Dissection and Downstream Analysis of Zebra Finch Embryos at Early Stages of Development

    PubMed Central

    Murray, Jessica R.; Stanciauskas, Monika E.; Aralere, Tejas S.; Saha, Margaret S.

    2014-01-01

    The zebra finch (Taeniopygiaguttata) has become an increasingly important model organism in many areas of research including toxicology1,2, behavior3, and memory and learning4,5,6. As the only songbird with a sequenced genome, the zebra finch has great potential for use in developmental studies; however, the early stages of zebra finch development have not been well studied. Lack of research in zebra finch development can be attributed to the difficulty of dissecting the small egg and embryo. The following dissection method minimizes embryonic tissue damage, which allows for investigation of morphology and gene expression at all stages of embryonic development. This permits both bright field and fluorescence quality imaging of embryos, use in molecular procedures such as in situ hybridization (ISH), cell proliferation assays, and RNA extraction for quantitative assays such as quantitative real-time PCR (qtRT-PCR). This technique allows investigators to study early stages of development that were previously difficult to access. PMID:24999108

  10. Essential Role of Chromatin Remodeling Protein Bptf in Early Mouse Embryos and Embryonic Stem Cells

    PubMed Central

    Landry, Joseph; Sharov, Alexei A.; Piao, Yulan; Sharova, Lioudmila V.; Xiao, Hua; Southon, Eileen; Matta, Jennifer; Tessarollo, Lino; Zhang, Ying E.; Ko, Minoru S. H.; Kuehn, Michael R.; Yamaguchi, Terry P.; Wu, Carl

    2008-01-01

    We have characterized the biological functions of the chromatin remodeling protein Bptf (Bromodomain PHD-finger Transcription Factor), the largest subunit of NURF (Nucleosome Remodeling Factor) in a mammal. Bptf mutants manifest growth defects at the post-implantation stage and are reabsorbed by E8.5. Histological analyses of lineage markers show that Bptf−/− embryos implant but fail to establish a functional distal visceral endoderm. Microarray analysis at early stages of differentiation has identified Bptf-dependent gene targets including homeobox transcriptions factors and genes essential for the development of ectoderm, mesoderm, and both definitive and visceral endoderm. Differentiation of Bptf−/− embryonic stem cell lines into embryoid bodies revealed its requirement for development of mesoderm, endoderm, and ectoderm tissue lineages, and uncovered many genes whose activation or repression are Bptf-dependent. We also provide functional and physical links between the Bptf-containing NURF complex and the Smad transcription factors. These results suggest that Bptf may co-regulate some gene targets of this pathway, which is essential for establishment of the visceral endoderm. We conclude that Bptf likely regulates genes and signaling pathways essential for the development of key tissues of the early mouse embryo. PMID:18974875

  11. Cytoskeletal changes in oocytes and early embryos during in vitro fertilization process in mice.

    PubMed

    Gumus, E; Bulut, H E; Kaloglu, C

    2010-02-01

    The cytoskeleton plays crucial roles in the development and fertilization of germ cells and in the early embryo development. The growth, maturation and fertilization of oocytes require an active movement and a correct localization of cellular organelles. This is performed by the re-organization of microtubules and actin filaments. Therefore, the aim of the present study was to determine the changes in cytoskeleton during in vitro fertilization process using appropriate immunofluorescence techniques. While the chromatin content was found to be scattered throughout the nucleus during the oocyte maturation period, it was seen only around nucleolus following the completion of the maturation. Microtubules, during oocyte maturation, were regularly distributed throughout the ooplasm which was then localized in the subcortical region of oocytes. Similarly microfilaments were scattered throughout the ooplasm during the oocyte maturation period whereas they were seen in the subcortical region around the polar body and above the meiotic spindle throughout the late developmental stages. In conclusion, those changes occurred in microtubules and microfilaments might be closely related to the re-organization of the genetic material during the oocyte maturation and early embryo development.

  12. The mRNA-bound proteome of the early fly embryo

    PubMed Central

    Wessels, Hans-Hermann; Imami, Koshi; Baltz, Alexander G.; Kolinski, Marcin; Beldovskaya, Anastasia; Selbach, Matthias; Small, Stephen; Ohler, Uwe; Landthaler, Markus

    2016-01-01

    Early embryogenesis is characterized by the maternal to zygotic transition (MZT), in which maternally deposited messenger RNAs are degraded while zygotic transcription begins. Before the MZT, post-transcriptional gene regulation by RNA-binding proteins (RBPs) is the dominant force in embryo patterning. We used two mRNA interactome capture methods to identify RBPs bound to polyadenylated transcripts within the first 2 h of Drosophila melanogaster embryogenesis. We identified a high-confidence set of 476 putative RBPs and confirmed RNA-binding activities for most of 24 tested candidates. Most proteins in the interactome are known RBPs or harbor canonical RBP features, but 99 exhibited previously uncharacterized RNA-binding activity. mRNA-bound RBPs and TFs exhibit distinct expression dynamics, in which the newly identified RBPs dominate the first 2 h of embryonic development. Integrating our resource with in situ hybridization data from existing databases showed that mRNAs encoding RBPs are enriched in posterior regions of the early embryo, suggesting their general importance in posterior patterning and germ cell maturation. PMID:27197210

  13. cAMP and in vivo hypoxia induce tob, ifr1, and fos expression in erythroid cells of the chick embryo.

    PubMed

    Dragon, Stefanie; Offenhäuser, Nina; Baumann, Rosemarie

    2002-04-01

    During avian embryonic development, terminal erythroid differentiation occurs in the circulation. Some of the key events, such as the induction of erythroid 2,3-bisphosphoglycerate (2,3-BPG), carbonic anhydrase (CAII), and pyrimidine 5'-nucleotidase (P5N) synthesis are oxygen dependent (Baumann R, Haller EA, Schöning U, and Weber M, Dev Biol 116: 548-551, 1986; Dragon S and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 280: R870-R878, 2001; Dragon S, Carey C, Martin K, and Baumann R, J Exp Biol 202: 2787-2795, 1999; Dragon S, Glombitza S, Götz R, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996; Dragon S, Hille R, Götz R, and Baumann R, Blood 91: 3052-3058, 1998; Million D, Zillner P, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 261: R1188-R1196, 1991) in an indirect way: hypoxia stimulates the release of norepinephrine (NE)/adenosine into the circulation (Dragon et al., J Exp Biol 202: 2787-2795, 1999; Dragon et al., Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996). This leads via erythroid beta-adrenergic/adenosine A(2) receptor activation to a cAMP signal inducing several proteins in a transcription-dependent manner (Dragon et al., Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996; Dragon et al., Blood 91: 3052-3058, 1998; Glombitza S, Dragon S, Berghammer M, Pannermayr M, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 271: R973-R981, 1996). To understand how the cAMP-dependent processes are initiated, we screened an erythroid cDNA library for cAMP-regulated genes. We detected three genes that were strongly upregulated (>5-fold) by cAMP in definitive and primitive red blood cells. They are homologous to the mammalian Tob, Ifr1, and Fos proteins. In addition, the genes are induced in the intact embryo during short-term hypoxia. Because the genes are regulators of proliferation and differentiation in other cell types, we suggest that c

  14. A single center, open label study of intradermal administration of an inactivated purified chick embryo cell culture rabies virus vaccine in adults.

    PubMed

    Recuenco, Sergio; Warnock, Eli; Osinubi, Modupe O V; Rupprecht, Charles E

    2017-08-03

    In the USA, rabies vaccines (RVs) are licensed for intramuscular (IM) use only, although RVs are licensed for use by the intradermal (ID) route in many other countries. Recent limitations in supplies of RV in the USA reopened discussions on the more efficient use of available biologics, including utilization of more stringent risk assessments, and potential ID RV administration. A clinical trial was designed to compare the immunogenic and adverse effects of a purified chicken embryo cell (PCEC) RV administered ID or IM. Enrollment was designed in four arms, ID Pre-Exposure Prophylaxis (Pre-EP), IM Pre-EP, ID Booster, and IM Booster vaccination. Enrollment included 130 adult volunteers. The arms with IM administration received vaccine according to the current ACIP recommendations: Pre-EP, three 1mL (2.5 I.U.) RV doses, each on day 0, 7, and 21; or a routine Booster, one 1ml dose. The ID groups received the same schedule, but doses administered were in a volume of 0.1mL (0.25 I.U.). The rate of increase in rabies virus neutralizing antibody titers 14-21days after vaccination were similar in the ID and correspondent IM groups. The GMT values for ID vaccination were slightly lower than those for IM vaccination, for both naïve and booster groups, and these differences were statistically significant by t-test. Fourteen days after completing vaccination, all individuals developed RV neutralizing antibody titers over the minimum arbitrary value obtained with the rapid fluorescent focus inhibition test (RFFIT). Antibodies were over the set threshold until the end of the trial, 160days after completed vaccination. No serious adverse reactions were reported. Most frequent adverse reactions were erythema, induration and tenderness, localized at the site of injection. Multi use of 1mL rabies vaccine vials for ID doses of 0.1 was demonstrated to be both safe and inmunogenic. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Types of neural cells in the spinal ganglia of human embryos and early fetuses.

    PubMed

    Olszewska, B; Woźniak, W; Gardner, E; O'Rahilly, R

    1979-01-01

    Spinal ganglial of human embryos and fetuses ranging in C.-R. length from 15 to 74 mm and in age from 6 1/2 to 11 postovulatory weeks were studied by light and electron microscopy. A sequence of events in differentiation and maturation enabled five types of cells to be distinguished: 1. apolar, undifferentiated neuroblasts are the main cells at 6 1/2 to 7 1/2 weeks; 2. early bipolar neuroblasts (strictly speaking, types 2 to 5 are immature neurons) predominate at the end of the embryonic period proper (8 postovulatory weeks); 3. intermediate bipolar neuroblasts are characteristic of the early fetal period; 4. late bipolar neuroblasts, in which two proceses arise separately from one pole of the cell, appear at about 10 postovulatory weeks; 5. unipolar neuroblasts are found within another week and, by that time, cells of types 1 and 2 are no longer present.

  16. Patterns of protein synthesis in oocytes and early embryos of Rana esculenta complex.

    PubMed

    Chen, P S; Stumm-Zollinger, E

    1986-01-01

    We have used isotopic labelling and both one-and two-dimensional electrophoretic procedures to analyse the protien synthesis patterns in oocytes and early embryos of three phenotypes of the European green frogs. The results demonstrated that protein patterns of Rana ridibunda and R. esculenta are identical, but that they differ from those of R. lessonae. Progeny of the lethal cross R. esculenta × R. esculenta showed a distinct delay in the appearance of stage-specific proteins during early embryogenesis. The heat-shock response of R. ridibunda and R. esculenta oocytes was found to be identical, but different from that of Xenopus laevis. The implications of these findings, with respect to hybridogenesis in R. esculenta complex and variations in the regulations of heat shock genes in different amphibian species, are discussed.

  17. Comparative effects of in ovo versus subcutaneous administration of the Marek's disease vaccine and pre-placement holding time on the early post-hatch quality of Ross × Ross 708 broiler chicks.

    PubMed

    Peebles, E D; Barbosa, T M; Cummings, T S; Dickson, J; Womack, S K

    2016-09-01

    Effects of method of administration [moa; in ovo (i.o.) or s.c.] of the Marek's disease vaccine and pre-placement holding time (pht) on early post-hatch male broiler chick quality was investigated. Sixty-five Ross × Ross 708 broiler hatching eggs were randomly set in each of 15 replicate trays (blocks) belonging to each of 4 pre-assigned moa and pht treatment combinations (3,900 total eggs) in a single stage Jamesway incubator. Eggs that were i.o.-vaccinated received injections at 18 d of incubation and male chicks from eggs that were not i.o.-injected were vaccinated by s.c. injection at hatch. The i.o. injections (50 μL) were delivered by a commercial multi-egg injector and the s.c. injections (200 μL) were delivered by an automatic pneumatic s.c. injector. Male chicks from each moa group also were subjected to either a 4 or 18 h pht. At hatch and placement total and yolk-free BW; body length; body mass index; yolk sac weight; yolk-free body and yolk sac weights as percentages of total BW; and yolk-free body and yolk moisture concentrations were determined. Chick BW also was determined at 7 d of age. Hatchability of fertile eggs was not affected by i.o. injection. However, at hatch, body length was increased and body mass index was decreased in response to i.o. injection. No main effect of moa or an interactive effect with pht was observed for the above variables at placement. However, body length was longer and body mass was lower in the 18 h than in the 4 h pht chicks. Placement yolk sac and body weights, and the 7 d BW of 18 h pht chicks was also lower than that of 4 h pht chicks. In conclusion, prolonging pht for 14 h adversely affected early chick quality, whereas i.o. injection did not negatively affect the early post-hatch quality of Ross × Ross 708 broiler chicks whether or not they were held for 4 or 18 h prior to placement. © 2016 Poultry Science Association Inc.

  18. Alcohol exposure alters DNA methylation profiles in mouse embryos at early neurulation

    PubMed Central

    Liu, Yunlong; Balaraman, Yokesh; Wang, Guohua; Nephew, Kenneth P.; Zhou, Feng C.

    2009-01-01

    Alcohol exposure during development can cause variable neurofacial deficit and growth retardation known as fetal alcohol spectrum disorders (FASD). The mechanism underlying FASD is not fully understood. However, alcohol, which is known to affect methyl donor metabolism, may induce aberrant epigenetic changes contributing to FASD. Using a tightly controlled whole-embryo culture, we investigated the effect of alcohol exposure (88 mM) at early embryonic neurulation on genome-wide DNA methylation and gene expression in the C57BL/6 mouse. The DNA methylation landscape around promoter CpG islands at early mouse development was analyzed using MeDIP (methylated DNA immunoprecipitation) coupled with microarray (MeDIP-chip). At early neurulation, genes associated with high CpG promoters (HCP) had a lower ratio of methylation but a greater ratio of expression. Alcohol-induced alterations in DNA methylation were observed, particularly in genes on chromosomes 7, 10 and X; remarkably, a >10 fold increase in the number of genes with increased methylation on chromosomes 10 and X was observed in alcohol-exposed embryos with a neural tube defect phenotype compared to embryos without a neural tube defect. Significant changes in methylation were seen in imprinted genes, genes known to play roles in cell cycle, growth, apoptosis, cancer, and in a large number of genes associated with olfaction. Altered methylation was associated with significant (p < 0.01) changes in expression for 84 genes. Sequenom EpiTYPER DNA methylation analysis was used for validation of the MeDIP-chip data. Increased methylation of genes known to play a role in metabolism (Cyp4f13) and decreased methylation of genes associated with development (Nlgn3, Elavl2, Sox21 and Sim1), imprinting (Igf2r) and chromatin (Hist1h3d) was confirmed. In a mouse model for FASD, we show for the first time that alcohol exposure during early neurulation can induce aberrant changes in DNA methylation patterns with associated changes

  19. Alcohol exposure alters DNA methylation profiles in mouse embryos at early neurulation.

    PubMed

    Liu, Yunlong; Balaraman, Yokesh; Wang, Guohua; Nephew, Kenneth P; Zhou, Feng C

    2009-10-01

    Alcohol exposure during development can cause variable neurofacial deficit and growth retardation known as fetal alcohol spectrum disorders (FASD). The mechanism underlying FASD is not fully understood. However, alcohol, which is known to affect methyl donor metabolism, may induce aberrant epigenetic changes contributing to FASD. Using a tightly controlled whole-embryo culture, we investigated the effect of alcohol exposure (88mM) at early embryonic neurulation on genome-wide DNA methylation and gene expression in the C57BL/6 mouse. The DNA methylation landscape around promoter CpG islands at early mouse development was analyzed using MeDIP (methylated DNA immunoprecipitation) coupled with microarray (MeDIP-chip). At early neurulation, genes associated with high CpG promoters (HCP) had a lower ratio of methylation but a greater ratio of expression. Alcohol-induced alterations in DNA methylation were observed, particularly in genes on chromosomes 7, 10, and X; remarkably, a >10 fold increase in the number of genes with increased methylation on chromosomes 10 and X was observed in alcohol-exposed embryos with a neural tube defect phenotype compared to embryos without a neural tube defect. Significant changes in methylation were seen in imprinted genes, genes known to play roles in cell cycle, growth, apoptosis, cancer, and in a large number of genes associated with olfaction. Altered methylation was associated with significant (p<0.01) changes in expression for 84 genes. Sequenom EpiTYPER DNA methylation analysis was used for validation of the MeDIP-chip data. Increased methylation of genes known to play a role in metabolism (Cyp4f13) and decreased methylation of genes associated with development (Nlgn3, Elavl2, Sox21 and Sim1), imprinting (Igf2r) and chromatin (Hist1h3d) was confirmed. In a mouse model for FASD, we show for the first time that alcohol exposure during early neurulation can induce aberrant changes in DNA methylation patterns with associated changes in

  20. Making lineage decisions with biological noise: Lessons from the early mouse embryo.

    PubMed

    Simon, Claire S; Hadjantonakis, Anna-Katerina; Schröter, Christian

    2018-04-30

    Understanding how individual cells make fate decisions that lead to the faithful formation and homeostatic maintenance of tissues is a fundamental goal of contemporary developmental and stem cell biology. Seemingly uniform populations of stem cells and multipotent progenitors display a surprising degree of heterogeneity, primarily originating from the inherent stochastic nature of molecular processes underlying gene expression. Despite this heterogeneity, lineage decisions result in tissues of a defined size and with consistent proportions of differentiated cell types. Using the early mouse embryo as a model we review recent developments that have allowed the quantification of molecular intercellular heterogeneity during cell differentiation. We first discuss the relationship between these heterogeneities and developmental cellular potential. We then review recent theoretical approaches that formalize the mechanisms underlying fate decisions in the inner cell mass of the blastocyst stage embryo. These models build on our extensive knowledge of the genetic control of fate decisions in this system and will become essential tools for a rigorous understanding of the connection between noisy molecular processes and reproducible outcomes at the multicellular level. We conclude by suggesting that cell-to-cell communication provides a mechanism to exploit and buffer intercellular variability in a self-organized process that culminates in the reproducible formation of the mature mammalian blastocyst stage embryo that is ready for implantation into the maternal uterus. This article is categorized under: Gene Expression and Transcriptional Hierarchies > Cellular Differentiation Establishment of Spatial and Temporal Patterns > Regulation of Size, Proportion, and Timing Gene Expression and Transcriptional Hierarchies > Gene Networks and Genomics Gene Expression and Transcriptional Hierarchies > Quantitative Methods and Models. © 2018 Wiley Periodicals, Inc.

  1. Toxicity and cardiac effects of carbaryl in early developing zebrafish (Danio rerio) embryos

    SciT

    Lin, C.C.; Hui, Michelle N.Y.; Cheng, S.H. E-mail: bhcheng@cityu.edu.hk

    2007-07-15

    Carbaryl, an acetylcholinesterase inhibitor, is known to be moderately toxic to adult zebrafish and has been reported to cause heart malformations and irregular heartbeat in medaka. We performed experiments to study the toxicity of carbaryl, specifically its effects on the heart, in early developing zebrafish embryos. LC50 and EC50 values for carbaryl at 28 h post-fertilization were 44.66 {mu}g/ml and 7.52 {mu}g/ml, respectively, and 10 {mu}g/ml carbaryl was used in subsequent experiments. After confirming acetylcholinesterase inhibition by carbaryl using an enzymatic method, we observed red blood cell accumulation, delayed hatching and pericardial edema, but not heart malformation as described inmore » some previous reports. Our chronic exposure data also demonstrated carbaryl-induced bradycardia, which is a common effect of acetylcholinesterase inhibitors due to the accumulation of acetylcholine, in embryos from 1 day post-fertilization (dpf) to 5 dpf. The distance between the sinus venosus, the point where blood enters the atrium, and the bulbus arteriosus, the point where blood leaves the ventricle, indicated normal looping of the heart tube. Immunostaining of myosin heavy chains with the ventricle-specific antibody MF20 and the atrium-specific antibody S46 showed normal development of heart chambers. At the same time, acute exposure resulted in carbaryl-induced bradycardia. Heart rate dropped significantly after a 10-min exposure to 100 {mu}g/ml carbaryl but recovered when carbaryl was removed. The novel observation of carbaryl-induced bradycardia in 1- and 2-dpf embryos suggested that carbaryl affected cardiac function possibly through an alternative mechanism other than acetylcholinesterase inhibition such as inhibition of calcium ion channels, since acetylcholine receptors in zebrafish are not functional until 3 dpf. However, the exact nature of this mechanism is currently unknown, and thus further studies are required.« less

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

    PubMed

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

    2016-10-01

    Biological clock genes expressed in reproductive tissues play important roles in maintaining the normal functions of reproductive system. However, disruption of female circadian rhythm on oocyte fertilization, preimplantation embryo development and blastocyst implantation potential is still unclear. In this study, ovulation, in vivo and in vitro oocyte fertilization, embryo development, implantation and intracellular reactive oxygen species (ROS) levels in ovary and oviduct were studied in female Bmal1+/+ and Bmal1-/- mice. The number of naturally ovulated oocyte in Bmal1-/- mice decreased (5.2 ± 0.8 vs 7.8 ± 0.8, P < 0.001), with an increasing abnormal oocyte ratio (20.4 ± 3.5 vs 11.7 ± 2.0%, P = 0.001) after superovulation. Significantly lower fertilization rate and obtained blastocyst number were observed in Bmal1-/- female mice either mated with wild-type in vivo or fertilized by sperm from wild-type male mice in vitro (all P < 0.05). Interestingly, in vitro fertilization rate of oocytes derived from Bmal1-/- increased significantly compared with in vivo study (P < 0.01). After transferring blastocysts derived from Bmal1+/+ and Bmal1-/- female mice to pseudopregnant mice, the implantation sites of the latter decreased 5 days later (8.0 ± 0.8 vs 5.3 ± 1.0, P = 0.005). The intracellular ROS levels in the ovary on proestrus day and in the oviduct on metestrus day increased significantly in Bmal1-/- mice compared with that of Bmal1+/+ mice. Deletion of the core biological clock gene Bmal1 significantly decreases oocyte fertilization rate, early embryo development and implantation potential in female mice, and these may be possibly caused by excess ROS levels generated in ovary and oviduct.

  3. Effect of PMA-induced protein kinase C activation on development and apoptosis in early zebrafish embryos.

    PubMed

    Hrubik, Jelena; Glisic, Branka; Samardzija, Dragana; Stanic, Bojana; Pogrmic-Majkic, Kristina; Fa, Svetlana; Andric, Nebojsa

    2016-12-01

    Protein kinase C (PKC) isoforms have been implicated in several key steps during early development, but the consequences of xenobiotic-induced PKC activation during early embryogenesis are still unknown. In this study, zebrafish embryos were exposed to a range of phorbol 12-myristate 13-acetate (PMA) concentrations (0-200μg/L) at different time points after fertilization. Results showed that 200μgPMA/L caused development of yolk bags, cardiac edema, slow blood flow, pulsating blood flow, slow pulse, elongated heart, lack of tail fins, curved tail, and coagulation. PMA exposure decreased survival rate of the embryos starting within the first 24h and becoming more pronounced after prolonged exposure (96h). PMA increased the number of apoptotic cells in the brain region as demonstrated by acridine orange staining and caused up-regulation of caspase 9 (casp9) and p53 up-regulated modulator of apoptosis (puma) mRNA in whole embryos. PMA caused oxidative stress in the embryos as demonstrated by decreased mRNA expression of catalase and superoxide dismutase 2. Inhibition of Pkc with GF109203X improved overall survival rate, reduced apoptosis in the brain and decreased expression of casp9 and puma in the PMA-exposed embryos. However, Pkc inhibition neither prevented development of deformities nor reversed oxidative stress in the PMA-exposed embryos. These data suggest that direct over-activation of Pkc during early embryogenesis of zebrafish is associated with apoptosis and decreased survival rate of the embryos. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. Starch Turnover and Metabolism during Flower and Early Embryo Development1[CC-BY

    PubMed Central

    Pazmino, Diana; Gagliardini, Valeria

    2016-01-01

    The accumulation of starch within photosynthetic tissues and within dedicated storage organs has been characterized extensively in many species, and a function in buffering carbon availability or in fueling later growth phases, respectively, has been proposed. However, developmentally regulated starch turnover within heterotrophic tissues other than dedicated storage organs is poorly characterized, and its function is not well understood. Here, we report on the characterization of starch turnover during flower, early embryo, and silique development in Arabidopsis (Arabidopsis thaliana) using a combined clearing-staining technique on whole-mount tissue. Besides the two previously documented waves of transient starch accumulation in the stamen envelope, occurring during meiosis and pollen mitosis I, we identified a novel, third wave of starch amylogenesis/amylolysis during the last stages of stamen development. To gain insights into the underlying molecular mechanisms, we analyzed publicly available microarray data, which revealed a developmentally coordinated expression of carbohydrate transport and metabolism genes during these waves of transient starch accumulation. Based on this analysis, we characterized starch dynamics in mutants affecting hexose phosphate metabolism and translocation, and identified the Glc-6-phosphate/phosphate antiporter GPT1 as the putative translocator of Glc-6-phosphate for starch biosynthesis in reproductive tissues. Based on these results, we propose a model of starch synthesis within the pollen grain and discuss the nutrient transport route feeding the embryo within the developing seed. PMID:27794100

  5. Single-cell transcriptome of early embryos and cultured embryonic stem cells of cynomolgus monkeys

    PubMed Central

    Nakamura, Tomonori; Yabuta, Yukihiro; Okamoto, Ikuhiro; Sasaki, Kotaro; Iwatani, Chizuru; Tsuchiya, Hideaki; Saitou, Mitinori

    2017-01-01

    In mammals, the development of pluripotency and specification of primordial germ cells (PGCs) have been studied predominantly using mice as a model organism. However, divergences among mammalian species for such processes have begun to be recognized. Between humans and mice, pre-implantation development appears relatively similar, but the manner and morphology of post-implantation development are significantly different. Nevertheless, the embryogenesis just after implantation in primates, including the specification of PGCs, has been unexplored due to the difficulties in analyzing the embryos at relevant developmental stages. Here, we present a comprehensive single-cell transcriptome dataset of pre- and early post-implantation embryo cells, PGCs and embryonic stem cells (ESCs) of cynomolgus monkeys as a model of higher primates. The identities of each transcriptome were also validated rigorously by other way such as immunofluorescent analysis. The information reported here will serve as a foundation for our understanding of a wide range of processes in the developmental biology of primates, including humans. PMID:28649393

  6. Selection and dynamics of embryonic stem cell integration into early mouse embryos

    PubMed Central

    Alexandrova, Stoyana; Kalkan, Tuzer; Humphreys, Peter; Riddell, Andrew; Scognamiglio, Roberta; Trumpp, Andreas; Nichols, Jennifer

    2016-01-01

    The process by which pluripotent cells incorporate into host embryos is of interest to investigate cell potency and cell fate decisions. Previous studies suggest that only a minority of the embryonic stem cell (ESC) inoculum contributes to the adult chimaera. How incoming cells are chosen for integration or elimination remains unclear. By comparing a heterogeneous mix of undifferentiated and differentiating ESCs (serum/LIF) with more homogeneous undifferentiated culture (2i/LIF), we examine the role of cellular heterogeneity in this process. Time-lapse ex vivo imaging revealed a drastic elimination of serum/LIF ESCs during early development in comparison with 2i/LIF ESCs. Using a fluorescent reporter for naive pluripotency (Rex1-GFP), we established that the acutely eliminated serum/LIF ESCs had started to differentiate. The rejected cells were apparently killed by apoptosis. We conclude that a selection process exists by which unwanted differentiating cells are eliminated from the embryo. However, occasional Rex1− cells were able to integrate. Upregulation of Rex1 occurred in a proportion of these cells, reflecting the potential of the embryonic environment to expedite diversion from differentiation priming to enhance the developing embryonic epiblast. PMID:26586221

  7. Copper induces expression and methylation changes of early development genes in Crassostrea gigas embryos.

    PubMed

    Sussarellu, Rossana; Lebreton, Morgane; Rouxel, Julien; Akcha, Farida; Rivière, Guillaume

    2018-03-01

    Copper contamination is widespread along coastal areas and exerts adverse effects on marine organisms such as mollusks. In the Pacific oyster, copper induces severe developmental abnormalities during early life stages; however, the underlying molecular mechanisms are largely unknown. This study aims to better understand whether the embryotoxic effects of copper in Crassostrea gigas could be mediated by alterations in gene expression, and the putative role of DNA methylation, which is known to contribute to gene regulation in early embryo development. For that purpose, oyster embryos were exposed to 4 nominal copper concentrations (0.1, 1, 10 and 20 μg L -1 Cu 2+ ) during early development assays. Embryotoxicity was monitored through the oyster embryo-larval bioassay at the D-larva stage 24 h post fertilization (hpf) and genotoxicity at gastrulation 7 hpf. In parallel, the relative expression of 15 genes encoding putative homeotic, biomineralization and DNA methylation proteins was measured at three developmental stages (3 hpf morula stage, 7 hpf gastrula stage, 24 hpf D-larvae stage) using RT-qPCR. Global DNA content in methylcytosine and hydroxymethylcytosine were measured by HPLC and gene-specific DNA methylation levels were monitored using MeDIP-qPCR. A significant increase in larval abnormalities was observed from copper concentrations of 10 μg L -1 , while significant genotoxic effects were detected at 1 μg L -1 and above. All the selected genes presented a stage-dependent expression pattern, which was impaired for some homeobox and DNA methylation genes (Notochord, HOXA1, HOX2, Lox5, DNMT3b and CXXC-1) after copper exposure. While global DNA methylation (5-methylcytosine) at gastrula stage didn't show significant changes between experimental conditions, 5-hydroxymethylcytosine, its degradation product, decreased upon copper treatment. The DNA methylation of exons and the transcript levels were correlated in control samples for HOXA1 but such

  8. Disparities in reproductive outcomes according to the endometrial preparation protocol in frozen embryo transfer : The risk of early pregnancy loss in frozen embryo transfer cycles.

    PubMed

    Hatoum, I; Bellon, L; Swierkowski, N; Ouazana, M; Bouba, S; Fathallah, K; Paillusson, B; Bailly, M; Boitrelle, F; Alter, L; Bergère, M; Selva, J; Wainer, R

    2018-03-01

    The purpose of this study was to determine the effect of stimulated and artificial endometrial preparation protocols on reproductive outcomes in frozen embryo transfer (FET) cycles. We performed a retrospective study of 1926 FET cycles over a 3.5-year period in the Fertility Unit at a University Hospital. Stimulated and artificial protocols were used for endometrial preparation. The embryos for FET were obtained from either in vitro fertilization or intracytoplasmic sperm injection cycles. Live birth rate and early pregnancy loss rates were retrospectively compared. In artificial protocols, oral or vaginal administration of oestradiol 2 mg two or three times a day was followed by vaginal supplementation with progesterone 200 mg two or three times a day. In stimulated protocols, recombinant follicle-stimulating hormone was administered from day 4 onward. Vaginal ultrasound was used for endometrial and ovarian monitoring. A pregnancy test was performed 14 days after FET. If it was positive, oestradiol and progesterone were administered up until the 12th week of gestation in artificial cycles. We defined early pregnancy losses as biochemical pregnancies (preclinical losses) and miscarriages. Data on 865 artificial cycles (45% of the total) and 1061 stimulated cycles (55%) were collected. Early pregnancy loss rate was significantly lower for stimulated cycles (34.2%) than for artificial cycles (56.9%), and the live birth rate was significantly higher for stimulated cycles (59.7%) than for artificial cycles (29.1%). In frozen embryo transfer, artificial cycles were associated with more early pregnancy loss and lower live birth rate than stimulated cycles.

  9. [Establishment of sprouting embryoid body model mimicking early embryonic vasculogenesis in human embryo].

    PubMed

    Jiang, Hua; Feng, You-Ji; Xie, Yi; Han, Jin-Lan; Wang, Zack; Chen, Tong

    2008-10-14

    To establish a sprouting embryoid body model mimicking early embryonic vasculogenesis in human embryo. Human embryonic stem were (hESCs) were cultured on the mouse embryo fibroblasts and then were induced to differentiate to form three-dimensional EB. The hEBs were cultured in media containing various angiogenesis-related factors: vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), endostatin, angiostatin, and platelet factor (PF)-4 of different concentrations for 3 days to observe the sprouting of the hEBs. 3, 3, 3', 3'-tetramethylindo-carbocyanine perchlorate labeled acetylated low density lipoprotein (Dil-AcLDL) was added onto the hEBs foe 4 h Immunofluorescence assay was used to observe if Dil-AcLDL was absorbed and if CD31 was expressed so as to determine the existence of embryonic endothelial cells in the sprouting structures. The ideal culturing condition was analyzed. The differentiated EBs formed sprouting structures in the collagen I matrix containing VEGF and FGF. The sprouts among individual EBs were able to link to each other and form vascular network-like structures. In the presence of VEGF and FGF, the sprouts branching from the EBs assimilated Dil-AcLDL, expressed CD31 and formed a 3-dimensional cylindrical organization. The concentrations of growth factors ideally stimulating sprouting growth were 100 ng/ml of VEGF and 50 ng/ml of FGF. The networks among the EBs were abolished by the angiostatin, endostatin, and PF4. The sprouting from hEBs accumulates embryonic endothelial cells and the sprouting network-like structures are indeed endothelial in nature. Inducing of sprouting EBs is an ideal model that mimics early embryonic vasculogenesis in humans.

  10. Intracellular pH in early Xenopus embryos: its effect on current flow between blastomeres.

    PubMed Central

    Turin, L; Warner, A E

    1980-01-01

    1. Electrophysiological techniques were used to monitor the flow of electric current from one cell to the next in Xenopus laevis embryos between the 4-cell and early blastula stages of development. Intracellular pH and blastocoel pH were determined using pH-sensitive micro-electrodes. 2. The resting intracellular pH was 7.74+/-0.02 (S.E. of mean, n = 29); there were no systematic differences between developmental stages. Blastocoel cavity pH was 8.4+/-0.06 (S.E. of mean, n = 10). The intracellular buffer value was 18 m-equiv. H+/pH unit per litre. 3. In embryos treated with bicarbonate buffered Holtfreter solution equilibrated with 100% CO2 the intracellular pH fell to 6.3+/-0.17 (S.D., n = 8). The membrane potential fell and the input resistance increased. The size of the effect on membrane potential and input resistance varied. 4. From the 32-cell stage onwards current flow from one cell to the next was abolished when the intracellular pH fell to below 6.5; the effect was rapid in onset and completely reversible. At cleavage stages of development lowering intracellular pH with CO2 had no effect on current flow from cell to cell. 5. The relationship between intracellular pH and current flow from cell to cell was sigmoid and covered between 0.2 and 0.4 pH units. The pH at which current flow was completely abolished ranged from 6.85 to 6.4. 6. Alterations in extraembryonic pH over the range 5.8-7.5 had no effect on any parameter measured. 7. We conclude that lowering the intracellular pH increases the resistance of both non-junctional junctional membranes. The data do not allow us to extract the pH junctional conductance relationship. 8. Variations in intracellular pH may provide a useful tool for the study of the functional role of direct cell to cell communication in both adult organs and early embryos. PMID:6770084

  11. Characterization of the altered gene expression profile in early porcine embryos generated from parthenogenesis and somatic cell chromatin transfer.

    PubMed

    Zhou, Chi; Dobrinsky, John; Tsoi, Stephen; Foxcroft, George R; Dixon, Walter T; Stothard, Paul; Verstegen, John; Dyck, Michael K

    2014-01-01

    The in vitro production of early porcine embryos is of particular scientific and economic interest. In general, embryos produced from in vitro Assisted Reproductive Technologies (ART) manipulations, such as somatic cell chromatin transfer (CT) and parthenogenetic activation (PA), are less developmentally competent than in vivo-derived embryos. The mechanisms underlying the deficiencies of embryos generated from PA and CT have not been completely understood. To characterize the altered genes and gene networks in embryos generated from CT and PA, comparative transcriptomic analyses of in vivo (IVV) expanded blastocysts (XB), IVV hatched blastocyst (HB), PA XB, PA HB, and CT HB were performed using a custom microarray platform enriched for genes expressed during early embryonic development. Differential expressions of 1492 and 103 genes were identified in PA and CT HB, respectively, in comparison with IVV HB. The "eIF2 signalling", "mitochondrial dysfunction", "regulation of eIF4 and p70S6K signalling", "protein ubiquitination", and "mTOR signalling" pathways were down-regulated in PA HB. Dysregulation of notch signalling-associated genes were observed in both PA and CT HB. TP53 was predicted to be activated in both PA and CT HB, as 136 and 23 regulation targets of TP53 showed significant differential expression in PA and CT HB, respectively, in comparison with IVV HB. In addition, dysregulations of several critical pluripotency, trophoblast development, and implantation-associated genes (NANOG, GATA2, KRT8, LGMN, and DPP4) were observed in PA HB during the blastocyst hatching process. The critical genes that were observed to be dysregulated in CT and PA embryos could be indicative of underlying developmental deficiencies of embryos produced from these technologies.

  12. Maternal program of apoptosis activated shortly after midblastula transition by overexpression of S-adenosylmethionine decarboxylase in Xenopus early embryos.

    PubMed

    Shiokawa, K; Kai, M; Higo, T; Kaito, C; Yokoska, J; Yasuhiko, Y; Kajita, E; Nagano, M; Yamada, Y; Shibata, M; Muto, T; Shinga, J; Hara, H; Takayama, E; Fukamachi, H; Yaoita, Y; Igarashi, K

    2000-06-01

    When we studied polyamine metabolism in Xenopus embryos, we cloned the cDNA for Xenopus S-adenosylmethionine decarboxylase (SAMDC), which converts SAM (S-adenosylmethionine), the methyl donor, into decarboxylated SAM (dcSAM), the aminopropyl donor, and microinjected its in vitro transcribed mRNA into Xenopus fertilized eggs. We found here that the mRNA injection induces a SAM deficient state in early embryos due to over-function of the overexpressed SAMDC, which in turn induces inhibition of protein synthesis. Such embryos developed quite normally until blastula stage, but stopped development at the early gastrula stage, due to induction of massive cell dissociation and cell autolysis, irrespective of the dosage and stage of the mRNA injection. We found that the dissociated cells were TUNEL-positive, contained fragmented nuclei with ladder-forming DNA, and furthermore, rescued completely by coinjection of Bcl-2 mRNA. Thus, overexpression of SAMDC in Xenopus embryos appeared to switch on apoptotic program, probably via inhibition of protein synthesis. Here, we briefly review our results together with those reported from other laboratories. After discussing the general importance of this newly discovered apoptotic program, we propose that the maternal program of apoptosis serves as a surveillance mechanism to eliminate metabolically severely-damaged cells and functions as a 'fail-safe' mechanism for normal development in Xenopus embryos.

  13. Blastomere biopsy influences epigenetic reprogramming during early embryo development, which impacts neural development and function in resulting mice.

    PubMed

    Wu, Yibo; Lv, Zhuo; Yang, Yang; Dong, Guoying; Yu, Yang; Cui, Yiqiang; Tong, Man; Wang, Liu; Zhou, Zuomin; Zhu, Hui; Zhou, Qi; Sha, Jiahao

    2014-05-01

    Blastomere biopsy is used in preimplantation genetic diagnosis; however, the long-term implications on the offspring are poorly characterized. We previously reported a high risk of memory defects in adult biopsied mice. Here, we assessed nervous function of aged biopsied mice and further investigated the mechanism of neural impairment after biopsy. We found that aged biopsied mice had poorer spatial learning ability, increased neuron degeneration, and altered expression of proteins involved in neural degeneration or dysfunction in the brain compared to aged control mice. Furthermore, the MeDIP assay indicated a genome-wide low methylation in the brains of adult biopsied mice when compared to the controls, and most of the genes containing differentially methylated loci in promoter regions were associated with neural disorders. When we further compared the genomic DNA methylation profiles of 7.5-days postconception (dpc) embryos between the biopsy and control group, we found the whole genome low methylation in the biopsied group, suggesting that blastomere biopsy was an obstacle to de novo methylation during early embryo development. Further analysis on mRNA profiles of 4.5-dpc embryos indicated that reduced expression of de novo methylation genes in biopsied embryos may impact de novo methylation. In conclusion, we demonstrate an abnormal neural development and function in mice generated after blastomere biopsy. The impaired epigenetic reprogramming during early embryo development may be the latent mechanism contributing to the impairment of the nervous system in the biopsied mice, which results in a hypomethylation status in their brains.

  14. Ectodermal Wnt6 is an early negative regulator of limb chondrogenesis in the chicken embryo

    PubMed Central

    2010-01-01

    Background Pattern formation of the limb skeleton is regulated by a complex interplay of signaling centers located in the ectodermal sheath and mesenchymal core of the limb anlagen, which results, in the forelimb, in the coordinate array of humerus, radius, ulna, carpals, metacarpals and digits. Much less understood is why skeletal elements form only in the central mesenchyme of the limb, whereas muscle anlagen develop in the peripheral mesenchyme ensheathing the chondrogenic center. Classical studies have suggested a role of the limb ectoderm as a negative regulator of limb chondrogenesis. Results In this paper, we investigated the molecular nature of the inhibitory influence of the ectoderm on limb chondrogenesis in the avian embryo in vivo. We show that ectoderm ablation in the early limb bud leads to increased and ectopic expression of early chondrogenic marker genes like Sox9 and Collagen II, indicating that the limb ectoderm inhibits limb chondrogenesis at an early stage of the chondrogenic cascade. To investigate the molecular nature of the inhibitory influence of the ectoderm, we ectopically expressed Wnt6, which is presently the only known Wnt expressed throughout the avian limb ectoderm, and found that Wnt6 overexpression leads to reduced expression of the early chondrogenic marker genes Sox9 and Collagen II. Conclusion Our results suggest that the inhibitory influence of the ectoderm on limb chondrogenesis acts on an early stage of chondrogenesis upsteam of Sox9 and Collagen II. We identify Wnt6 as a candidate mediator of ectodermal chondrogenic inhibition in vivo. We propose a model of Wnt-mediated centripetal patterning of the limb by the surface ectoderm. PMID:20334703

  15. From soil mechanics to chick development.

    PubMed

    Wolpert, Lewis

    2018-01-01

    Here, I provide some recollections of my life, starting as a civil engineer in South Africa and how I gradually became interested in biology, particularly pattern formation. In retrospect, I think that my decision to work on chick embryos to study limb development back in 1966 turned out to be the right one. The principles discovered in these 50 years, both by my collaborators and by other colleagues, have established the principles of how the limb develops in higher vertebrates, including humans.

  16. Stress Induces AMP-Dependent Loss of Potency Factors Id2 and Cdx2 in Early Embryos and Stem Cells

    PubMed Central

    Xie, Yufen; Awonuga, Awoniyi; Liu, Jian; Rings, Edmond; Puscheck, Elizabeth Ella

    2013-01-01

    The AMP-activated protein kinase (AMPK) mediates rapid, stress-induced loss of the inhibitor of differentiation (Id)2 in blastocysts and trophoblast stem cells (TSC), and a lasting differentiation in TSC. However, it is not known if AMPK regulates other potency factors or regulates them before the blastocyst stage. The caudal-related homeodomain protein (Cdx)2 is a regulatory gene for determining TSC, the earliest placental lineage in the preimplantation mouse embryo, but is expressed in the oocyte and in early cleavage stage embryos before TSC arise. We assayed the expression of putative potency-maintaining phosphorylated Cdx2 ser60 in the oocyte, two-cell stage embryo, blastocyst, and in TSC. We studied the loss of Cdx2 phospho ser60 expression induced by hyperosmolar stress and its underlying mechanisms. Hyperosmolar stress caused rapid loss of nuclear Cdx2 phospho ser60 and Id2 in the two-cell stage embryo by 0.5 h. Stress-induced Cdx2 phospho ser60 and Id2 loss is reversed by the AMPK inhibitor compound C and is induced by the AMPK agonist 5-amino-1-β-d-ribofuranosyl-imidazole-4-carboxamide in the absence of stress. In the two-cell stage embryo and TSC hyperosmolar, stress caused AMPK-mediated loss of Cdx2 phospho ser60 as detected by immunofluorescence and immunoblot. We propose that AMPK may be the master regulatory enzyme for mediating stress-induced loss of potency as AMPK is also required for stress-induced loss of Id2 in blastocysts and TSC. Since AMPK mediates potency loss in embryos and stem cells it will be important to measure, test mechanisms for, and manage the AMPK function to optimize the stem cell and embryo quality in vitro and in vivo. PMID:23316940

  17. Characterisation of the dynamic behaviour of lipid droplets in the early mouse embryo using adaptive harmonic generation microscopy.

    PubMed

    Watanabe, Tomoko; Thayil, Anisha; Jesacher, Alexander; Grieve, Kate; Debarre, Delphine; Wilson, Tony; Booth, Martin; Srinivas, Shankar

    2010-06-03

    Lipid droplets (LD) are organelles with an important role in normal metabolism and disease. The lipid content of embryos has a major impact on viability and development. LD in Drosophila embryos and cultured cell lines have been shown to move and fuse in a microtubule dependent manner. Due to limitations in current imaging technology, little is known about the behaviour of LD in the mammalian embryo. Harmonic generation microscopy (HGM) allows one to image LD without the use of exogenous labels. Adaptive optics can be used to correct aberrations that would otherwise degrade the quality and information content of images. We have built a harmonic generation microscope with adaptive optics to characterise early mouse embryogenesis. At fertilization, LD are small and uniformly distributed, but in the implanting blastocyst, LD are larger and enriched in the invading giant cells of the trophectoderm. Time-lapse studies reveal that LD move continuously and collide but do not fuse, instead forming aggregates that subsequently behave as single units. Using specific inhibitors, we show that the velocity and dynamic behaviour of LD is dependent not only on microtubules as in other systems, but also on microfilaments. We explore the limits within which HGM can be used to study living embryos without compromising viability and make the counterintuitive finding that 16 J of energy delivered continuously over a period of minutes can be less deleterious than an order of magnitude lower energy delivered dis-continuously over a period of hours. LD in pre-implantation mouse embryos show a previously unappreciated complexity of behaviour that is dependent not only on microtubules, but also microfilaments. Unlike LD in other systems, LD in the mouse embryo do not fuse but form aggregates. This study establishes HGM with adaptive optics as a powerful tool for the study of LD biology and provides insights into the photo-toxic effects of imaging embryos.

  18. Misexpression of BRE gene in the developing chick neural tube affects neurulation and somitogenesis

    PubMed Central

    Wang, Guang; Li, Yan; Wang, Xiao-Yu; Chuai, Manli; Yeuk-Hon Chan, John; Lei, Jian; Münsterberg, Andrea; Lee, Kenneth Ka Ho; Yang, Xuesong

    2015-01-01

    The brain and reproductive expression (BRE) gene is expressed in numerous adult tissues and especially in the nervous and reproductive systems. However, little is known about BRE expression in the developing embryo or about its role in embryonic development. In this study, we used in situ hybridization to reveal the spatiotemporal expression pattern for BRE in chick embryo during development. To determine the importance of BRE in neurogenesis, we overexpressed BRE and also silenced BRE expression specifically in the neural tube. We established that overexpressing BRE in the neural tube indirectly accelerated Pax7+ somite development and directly increased HNK-1+ neural crest cell (NCC) migration and TuJ-1+ neurite outgrowth. These altered morphogenetic processes were associated with changes in the cell cycle of NCCs and neural tube cells. The inverse effect was obtained when BRE expression was silenced in the neural tube. We also determined that BMP4 and Shh expression in the neural tube was affected by misexpression of BRE. This provides a possible mechanism for how altering BRE expression was able to affect somitogenesis, neurogenesis, and NCC migration. In summary, our results demonstrate that BRE plays an important role in regulating neurogenesis and indirectly somite differentiation during early chick embryo development. PMID:25568339

  19. Single-cell multi-omics sequencing of mouse early embryos and embryonic stem cells.

    PubMed

    Guo, Fan; Li, Lin; Li, Jingyun; Wu, Xinglong; Hu, Boqiang; Zhu, Ping; Wen, Lu; Tang, Fuchou

    2017-08-01

    Single-cell epigenome sequencing techniques have recently been developed. However, the combination of different layers of epigenome sequencing in an individual cell has not yet been achieved. Here, we developed a single-cell multi-omics sequencing technology (single-cell COOL-seq) that can analyze the chromatin state/nucleosome positioning, DNA methylation, copy number variation and ploidy simultaneously from the same individual mammalian cell. We used this method to analyze the reprogramming of the chromatin state and DNA methylation in mouse preimplantation embryos. We found that within < 12 h of fertilization, each individual cell undergoes global genome demethylation together with the rapid and global reprogramming of both maternal and paternal genomes to a highly opened chromatin state. This was followed by decreased openness after the late zygote stage. Furthermore, from the late zygote to the 4-cell stage, the residual DNA methylation is preferentially preserved on intergenic regions of the paternal alleles and intragenic regions of maternal alleles in each individual blastomere. However, chromatin accessibility is similar between paternal and maternal alleles in each individual cell from the late zygote to the blastocyst stage. The binding motifs of several pluripotency regulators are enriched at distal nucleosome depleted regions from as early as the 2-cell stage. This indicates that the cis-regulatory elements of such target genes have been primed to an open state from the 2-cell stage onward, long before pluripotency is eventually established in the ICM of the blastocyst. Genes may be classified into homogeneously open, homogeneously closed and divergent states based on the chromatin accessibility of their promoter regions among individual cells. This can be traced to step-wise transitions during preimplantation development. Our study offers the first single-cell and parental allele-specific analysis of the genome-scale chromatin state and DNA

  20. Single-cell multi-omics sequencing of mouse early embryos and embryonic stem cells

    PubMed Central

    Guo, Fan; Li, Lin; Li, Jingyun; Wu, Xinglong; Hu, Boqiang; Zhu, Ping; Wen, Lu; Tang, Fuchou

    2017-01-01

    Single-cell epigenome sequencing techniques have recently been developed. However, the combination of different layers of epigenome sequencing in an individual cell has not yet been achieved. Here, we developed a single-cell multi-omics sequencing technology (single-cell COOL-seq) that can analyze the chromatin state/nucleosome positioning, DNA methylation, copy number variation and ploidy simultaneously from the same individual mammalian cell. We used this method to analyze the reprogramming of the chromatin state and DNA methylation in mouse preimplantation embryos. We found that within < 12 h of fertilization, each individual cell undergoes global genome demethylation together with the rapid and global reprogramming of both maternal and paternal genomes to a highly opened chromatin state. This was followed by decreased openness after the late zygote stage. Furthermore, from the late zygote to the 4-cell stage, the residual DNA methylation is preferentially preserved on intergenic regions of the paternal alleles and intragenic regions of maternal alleles in each individual blastomere. However, chromatin accessibility is similar between paternal and maternal alleles in each individual cell from the late zygote to the blastocyst stage. The binding motifs of several pluripotency regulators are enriched at distal nucleosome depleted regions from as early as the 2-cell stage. This indicates that the cis-regulatory elements of such target genes have been primed to an open state from the 2-cell stage onward, long before pluripotency is eventually established in the ICM of the blastocyst. Genes may be classified into homogeneously open, homogeneously closed and divergent states based on the chromatin accessibility of their promoter regions among individual cells. This can be traced to step-wise transitions during preimplantation development. Our study offers the first single-cell and parental allele-specific analysis of the genome-scale chromatin state and DNA

  1. Germ layer differentiation during early hindgut and cloaca formation in rabbit and pig embryos

    PubMed Central

    Hassoun, Romia; Schwartz, Peter; Rath, Detlef; Viebahn, Christoph; Männer, Jörg

    2010-01-01

    Relative to recent advances in understanding molecular requirements for endoderm differentiation, the dynamics of germ layer morphology and the topographical distribution of molecular factors involved in endoderm formation at the caudal pole of the embryonic disc are still poorly defined. To discover common principles of mammalian germ layer development, pig and rabbit embryos at late gastrulation and early neurulation stages were analysed as species with a human-like embryonic disc morphology, using correlative light and electron microscopy. Close intercellular contact but no direct structural evidence of endoderm formation such as mesenchymal–epithelial transition between posterior primitive streak mesoderm and the emerging posterior endoderm were found. However, a two-step process closely related to posterior germ layer differentiation emerged for the formation of the cloacal membrane: (i) a continuous mesoderm layer and numerous patches of electron-dense flocculent extracellular matrix mark the prospective region of cloacal membrane formation; and (ii) mesoderm cells and all extracellular matrix including the basement membrane are lost locally and close intercellular contact between the endoderm and ectoderm is established. The latter process involves single cells at first and then gradually spreads to form a longitudinally oriented seam-like cloacal membrane. These gradual changes were found from gastrulation to early somite stages in the pig, whereas they were found from early somite to mid-somite stages in the rabbit; in both species cloacal membrane formation is complete prior to secondary neurulation. The results highlight the structural requirements for endoderm formation during development of the hindgut and suggest new mechanisms for the pathogenesis of common urogenital and anorectal malformations. PMID:20874819

  2. Differential Expression of Metallothionein Isoforms in Terrestrial Snail Embryos Reflects Early Life Stage Adaptation to Metal Stress

    PubMed Central

    Baurand, Pierre-Emmanuel; Pedrini-Martha, Veronika; de Vaufleury, Annette; Niederwanger, Michael; Capelli, Nicolas; Scheifler, Renaud; Dallinger, Reinhard

    2015-01-01

    The aim of this study was to analyze the expression of three metallothionein (MT) isoform genes (CdMT, CuMT and Cd/CuMT), already known from adults, in the Early Life Stage (ELS) of Cantareus aspersus. This was accomplished by detection of the MT isoform-specific transcription adopting Polymerase Chain Reaction (PCR) amplification and quantitative Real Time (qRT)-PCR of the three MT genes. Freshly laid eggs were kept for 24 hours under control conditions or exposed to three cadmium (Cd) solutions of increasing concentration (5, 10, and 15 mg Cd/L). The transcription of the three MT isoform genes was detected via PCR in 1, 6 and 12-day-old control or Cd-exposed embryos. Moreover, the transcription of this isoform genes during development was followed by qRT-PCR in 6 and 12-day-old embryos. Our results showed that the CdMT and Cd/CuMT genes, but not the CuMT gene, are expressed in embryos at the first day of development. The transcription of the 3 MT genes in control embryos increased with development time, suggesting that the capacities of metal regulation and detoxification may have gradually increased throughout embryogenesis. However in control embryos, the most highly expressed MT gene was that of the Cd/CuMT isoform, whose transcription levels greatly exceeded those of the other two MT genes. This contrasts with the minor significance of this gene in adult snails and suggests that in embryos, this isoform may play a comparatively more important role in metal physiology compared to adult individuals. This function in adult snails appears not to be related to Cd detoxification. Instead, snail embryos responded to Cd exposure by over-expression of the CdMT gene in a concentration-dependent manner, whereas the expression of the Cd/CuMT gene remained unaffected. Moreover, our study demonstrates the ability of snail embryos to respond very early to Cd exposure by up-regulation of the CdMT gene. PMID:25706953

  3. Evaluation of a new five-injection, two-site,intradermal schedule for purified chick embryo cell rabies vaccine: A randomized, open-label, active-controlled trial in healthy adult volunteers in India.

    PubMed

    Sudarshan, M K; Madhusudana, S N; Mahendra, B J; Ashwath Narayana, D H; Ananda Giri, M S; Popova, O; Vakil, H B

    2005-07-01

    Human rabies is an ongoing significant public health problem inmany developing countries, with India reporting the highest incidence of rabies-related deaths (∼20,000 per year). Many people living in India cannot afford the standard IM postexposure prophylaxis (PEP) with cell-culture vaccines, which are administered using a 5-dose regimen developed in Essen, Germany. A potentially less expensive intradermal (ID) regimen, based on the Essen regimen, has been developed at the Kempegowda Institute of Medical Sciences (KIMS), Bangalore, India. The objective of this study was to compare the immunogenicity and local and systemic tolerability of the KIMS-1D regimen with those of the standard Essen IM regimen in healthy adult volunteers in India. This randomized, open-label, active-controlled trial was conductedat the Antirabies Clinic, Medical College, KIMS. Healthy adult volunteers were randomly assigned to receive purified chick embryo cell vaccine (PCECV) using the KIMS-1D regimen (0.1 mL injected ID at 2 body sites on days 0, 3, 7, 14, and 28 ["2-2-2-2-2"]) or the Essen IM regimen (1 mL injected IM at 1 body site on the same days Subjects were followed up for 365 days by the treating physician and encouraged to voluntarily report any adverse events (AEs). Serum rabies virus-neutralizing antibody (RVNA) concentrations were measured before the first injection on day 0 (baseline) and on days 14, 28, 90, 180, and 365, using the rapid fluorescent focus inhibition test. Ninety-one subjects were enrolled and included in the tolerabilityand immunogenicity analyses. The ID group comprised 45 subjects (26 men, 19 women; mean [SD] age, 20.84 [1.48] years); the IM group, 46 subjects (28 men, 18 women; mean [SD] age, 21.02 [1.16] years). The most common local AEs were pain at the injection site (2/225 [0.9%] in the ID group and 10/230 [4.3%] in the IM group; P < 0.006) and itching at the injection site (5/225 [2.2%] in the ID group and none in the IM group; P = 0.026). All of

  4. Evaluation of a new five-injection, two-site,intradermal schedule for purified chick embryo cell rabies vaccine: A randomized, open-label, active-controlled trial in healthy adult volunteers in India

    PubMed Central

    Sudarshan, M.K.; Madhusudana, S.N.; Mahendra, B.J.; Ashwath Narayana, D.H.; Ananda Giri, M.S.; Popova, O.; Vakil, H.B.

    2005-01-01

    Background: Human rabies is an ongoing significant public health problem inmany developing countries, with India reporting the highest incidence of rabies-related deaths (∼20,000 per year). Many people living in India cannot afford the standard IM postexposure prophylaxis (PEP) with cell-culture vaccines, which are administered using a 5-dose regimen developed in Essen, Germany. A potentially less expensive intradermal (ID) regimen, based on the Essen regimen, has been developed at the Kempegowda Institute of Medical Sciences (KIMS), Bangalore, India. Objective: The objective of this study was to compare the immunogenicity and local and systemic tolerability of the KIMS-1D regimen with those of the standard Essen IM regimen in healthy adult volunteers in India. Methods: This randomized, open-label, active-controlled trial was conductedat the Antirabies Clinic, Medical College, KIMS. Healthy adult volunteers were randomly assigned to receive purified chick embryo cell vaccine (PCECV) using the KIMS-1D regimen (0.1 mL injected ID at 2 body sites on days 0, 3, 7, 14, and 28 [“2-2-2-2-2”]) or the Essen IM regimen (1 mL injected IM at 1 body site on the same days Subjects were followed up for 365 days by the treating physician and encouraged to voluntarily report any adverse events (AEs). Serum rabies virus-neutralizing antibody (RVNA) concentrations were measured before the first injection on day 0 (baseline) and on days 14, 28, 90, 180, and 365, using the rapid fluorescent focus inhibition test. Results: Ninety-one subjects were enrolled and included in the tolerabilityand immunogenicity analyses. The ID group comprised 45 subjects (26 men, 19 women; mean [SD] age, 20.84 [1.48] years); the IM group, 46 subjects (28 men, 18 women; mean [SD] age, 21.02 [1.16] years). The most common local AEs were pain at the injection site (2/225 [0.9%] in the ID group and 10/230 [4.3%] in the IM group; P < 0.006) and itching at the injection site (5/225 [2.2%] in the ID group

  5. Biochemical defects of mutant nudel alleles causing early developmental arrest or dorsalization of the Drosophila embryo.

    PubMed Central

    LeMosy, E K; Leclerc, C L; Hashimoto, C

    2000-01-01

    The nudel gene of Drosophila is maternally required both for structural integrity of the egg and for dorsoventral patterning of the embryo. It encodes a structurally modular protein that is secreted by ovarian follicle cells. Genetic and molecular studies have suggested that the Nudel protein is also functionally modular, with a serine protease domain that is specifically required for ventral development. Here we describe biochemical and immunolocalization studies that provide insight into the molecular basis for the distinct phenotypes produced by nudel mutations and for the interactions between these alleles. Mutations causing loss of embryonic dorsoventral polarity result in a failure to activate the protease domain of Nudel. Our analyses support previous findings that catalytic activity of the protease domain is required for dorsoventral patterning and that the Nudel protease is auto-activated and reveal an important role for a region adjacent to the protease domain in Nudel protease function. Mutations causing egg fragility and early embryonic arrest result in a significant decrease in extracellular Nudel protein, due to defects in post-translational processing, stability, or secretion. On the basis of these and other studies of serine proteases, we suggest potential mechanisms for the complementary and antagonistic interactions between the nudel alleles. PMID:10628985

  6. Proteomic analysis of early-stage embryos: implications for egg quality in hapuku (Polyprion oxygeneios).

    PubMed

    Kohn, Yair Y; Symonds, Jane E; Kleffmann, Torsten; Nakagawa, Shinichi; Lagisz, Malgorzata; Lokman, P Mark

    2015-12-01

    In order to develop biomarkers that may help predict the egg quality of captive hapuku (Polyprion oxygeneios) and provide potential avenues for its manipulation, the present study (1) sequenced the proteome of early-stage embryos using isobaric tag for relative and absolute quantification analysis, and (2) aimed to establish the predictive value of the abundance of identified proteins with regard to egg quality through regression analysis. Egg quality was determined for eight different egg batches by blastomere symmetry scores. In total, 121 proteins were identified and assigned to one of nine major groups according to their function/pathway. A mixed-effects model analysis revealed a decrease in relative protein abundance that correlated with (decreasing) egg quality in one major group (heat-shock proteins). No differences were found in the other protein groups. Linear regression analysis, performed for each identified protein separately, revealed seven proteins that showed a significant decrease in relative abundance with reduced blastomere symmetry: two correlates that have been named in other studies (vitellogenin, heat-shock protein-70) and a further five new candidate proteins (78 kDa glucose-regulated protein, elongation factor-2, GTP-binding nuclear protein Ran, iduronate 2-sulfatase and 6-phosphogluconate dehydrogenase). Notwithstanding issues associated with multiple statistical testing, we conclude that these proteins, and especially iduronate 2-sulfatase and the generic heat-shock protein group, could serve as biomarkers of egg quality in hapuku.

  7. The methyltransferase Setdb1 is essential for meiosis and mitosis in mouse oocytes and early embryos.

    PubMed

    Eymery, Angeline; Liu, Zichuan; Ozonov, Evgeniy A; Stadler, Michael B; Peters, Antoine H F M

    2016-08-01

    Oocytes develop the competence for meiosis and early embryogenesis during their growth. Setdb1 is a histone H3 lysine 9 (H3K9) methyltransferase required for post-implantation development and has been implicated in the transcriptional silencing of genes and endogenous retroviral elements (ERVs). To address its role in oogenesis and pre-implantation development, we conditionally deleted Setdb1 in growing oocytes. Loss of Setdb1 expression greatly impaired meiosis. It delayed meiotic resumption, altered the dynamics of chromatin condensation, and impaired kinetochore-spindle interactions, bipolar spindle organization and chromosome segregation in more mature oocytes. The observed phenotypes related to changes in abundance of specific transcripts in mutant oocytes. Setdb1 maternally deficient embryos arrested during pre-implantation development and showed comparable defects during cell cycle progression and in chromosome segregation. Finally, transcriptional profiling data indicate that Setdb1 downregulates rather than silences expression of ERVK and ERVL-MaLR retrotransposons and associated chimearic transcripts during oogenesis. Our results identify Setdb1 as a newly discovered meiotic and embryonic competence factor safeguarding genome integrity at the onset of life. © 2016. Published by The Company of Biologists Ltd.

  8. Endometrium as an early sensor of in vitro embryo manipulation technologies

    PubMed Central

    Mansouri-Attia, Nadéra; Sandra, Olivier; Aubert, Julie; Degrelle, Séverine; Everts, Robin E.; Giraud-Delville, Corinne; Heyman, Yvan; Galio, Laurent; Hue, Isabelle; Yang, Xiangzhong; Tian, X. Cindy; Lewin, Harris A.; Renard, Jean-Paul

    2009-01-01

    Implantation is crucial for placental development that will subsequently impact fetal growth and pregnancy success with consequences on postnatal health. We postulated that the pattern of genes expressed by the endometrium when the embryo becomes attached to the mother uterus could account for the final outcome of a pregnancy. As a model, we used the bovine species where the embryo becomes progressively and permanently attached to the endometrium from day 20 of gestation onwards. At that stage, we compared the endometrial genes profiles in the presence of an in vivo fertilized embryo (AI) with the endometrial patterns obtained in the presence of nuclear transfer (SCNT) or in vitro fertilized embryos (IVF), both displaying lower and different potentials for term development. Our data provide evidence that the endometrium can be considered as a biological sensor able to fine-tune its physiology in response to the presence of embryos whose development will become altered much later after the implantation process. Compared with AI, numerous biological functions and several canonical pathways with a major impact on metabolism and immune function were found to be significantly altered in the endometrium of SCNT pregnancies at implantation, whereas the differences were less pronounced with IVF embryos. Determining the limits of the endometrial plasticity at the onset of implantation should bring new insights on the contribution of the maternal environment to the development of an embryo and the success of pregnancy. PMID:19297625

  9. No significant regulation of bicoid mRNA by Pumilio or Nanos in the early Drosophila embryo.

    PubMed

    Wharton, Tammy H; Nomie, Krystle J; Wharton, Robin P

    2018-01-01

    Drosophila Pumilio (Pum) is a founding member of the conserved Puf domain class of RNA-binding translational regulators. Pum binds with high specificity, contacting eight nucleotides, one with each of the repeats in its RNA-binding domain. In general, Pum is thought to block translation in collaboration with Nanos (Nos), which exhibits no binding specificity in isolation but is recruited jointly to regulatory sequences containing a Pum binding site in the 3'-UTRs of target mRNAs. Unlike Pum, which is ubiquitous in the early embryo, Nos is tightly restricted to the posterior, ensuring that repression of its best-characterized target, maternal hunchback (hb) mRNA, takes place exclusively in the posterior. An exceptional case of Nos-independent regulation by Pum has been described-repression of maternal bicoid (bcd) mRNA at the anterior pole of the early embryo, dependent on both Pum and conserved Pum binding sites in the 3'-UTR of the mRNA. We have re-investigated regulation of bcd in the early embryo; our experiments reveal no evidence of a role for Pum or its conserved binding sites in regulation of the perdurance of bcd mRNA or protein. Instead, we find that Pum and Nos control the accumulation of bcd mRNA in testes.

  10. Dynamic Properties of Electrotonic Coupling between Cells of Early Xenopus Embryos

    PubMed Central

    DiCaprio, R. A.; French, A. S.; Sanders, E. J.

    1974-01-01

    Frequency response functions were measured between the cells of Xenopus laevis embryos during the first two cleavage stages. Linear systems theory was then used to produce electronic models which account for the electrical behavior of the systems. Coupling between the cells may be explained by models which have simple resistive elements joining each cell to its neighbors. The vitelline, or fertilization, membrane which surrounds the embryos has no detectable resistance to the passage of electric current. The electrical properties of the four-cell embryo can only be explained by the existence of individual junctions linking each pair of cells. This arrangement suggests that electrotonic coupling is important in the development of the embryos, at least until the four-cell stage. ImagesFIGURE 5FIGURE 14FIGURE 15 PMID:19431351

  11. The effect of repeated light-dark shifts on uterine receptivity and early gestation in mice undergoing embryo transfer.

    PubMed

    Goldstein, Cathy A; O'Brien, Louise M; Bergin, Ingrid L; Saunders, Thomas L

    2018-04-01

    Female shift workers are at increased risk for negative reproductive outcomes, and animal evidence suggests that manipulation of the light-dark cycle is detrimental to early gestation in female mice. Specifically, failure of implantation may be responsible for these findings. The objective of this study was to better delineate which reproductive processes are vulnerable to detrimental effects of maternal circadian disturbance. We exposed mice undergoing embryo transfer to repetitive phase advances of the photoperiod. Embryos were derived from donor sperm and eggs from mice living in normal light-dark conditions to isolate the effects of photoperiod disruption on uterine receptivity and early gestation. Twenty-eight mice receiving embryo transfer underwent an experimental light-dark condition (advance of lights on and lights off by 6 hours every 4 days). Twenty-eight mice remained in a normal light-dark condition. Animals lived in their assigned light-dark condition beginning 2 weeks prior to embryo transfer and ending the day of uterine necropsy (post-coitus day 14.5). Wilcoxon-Mann-Whitney test demonstrated no significant differences between control and experimental light-dark conditions in pups (Z=0.10, p=.92), resorptions (Z=0.20, p=.84), or implantations (Z=-0.34, p=.73). Pup and placental weights were similar between groups. In this investigation, uterine receptivity and maintenance of early gestation were preserved despite recurrent phase advances in photoperiod. This finding, in the context of the current literature, suggests that the negative effects of circadian disruption are mediated by reproductive processes upstream of implantation.

  12. Improvement of porcine cloning efficiency by trichostain A through early-stage induction of embryo apoptosis.

    PubMed

    Ji, Qianqian; Zhu, Kongju; Liu, Zhiguo; Song, Zhenwei; Huang, Yuankai; Zhao, Haijing; Chen, Yaosheng; He, Zuyong; Mo, Delin; Cong, Peiqing

    2013-03-15

    Trichostain A (TSA), an inhibitor of histone deacetylases, improved developmental competence of SCNT embryos in many species, apparently by improved epigenetic reprogramming. The objective of the present study was to determine the effects of TSA-induced apoptosis in cloned porcine embryos. At various developmental stages, a comet assay and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining were used to detect apoptosis, and real-time polymerase chain reaction was used to assess expression of genes related to apoptosis and pluripotency. In this study, TSA significantly induced apoptosis (in a dose-dependent manner) at the one-, two-, and four-cell stages. However, in blastocyst stage embryos, TSA decreased the apoptotic index (P < 0.05). Expression levels of Caspase 3 were higher in TSA-treated versus control embryos at the two-cell stage (not statistically significant). The expression ratio of antiapoptotic Bcl-xl gene to proapoptotic Bax gene, an indicator of antiapoptotic potential, was higher in TSA-treated groups at the one-, two-, and four-cell and blastocyst stages. Furthermore, expression levels of pluripotency-related genes, namely, Oct4 and Nanog, were elevated at the morula stage (P < 0.05) in TSA treatment groups. We concluded that inducing apoptosis might be a mechanism by which TSA promotes development of reconstructed embryos. At the initial stage of apoptosis induction, abnormal cells were removed, thereby enhancing proliferation of healthy cells and improving embryo quality. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Role of nucleation-promoting factors in mouse early embryo development.

    PubMed

    Wang, Qiao-Chu; Liu, Jun; Wang, Fei; Duan, Xing; Dai, Xiao-Xin; Wang, Teng; Liu, Hong-Lin; Cui, Xiang-Shun; Sun, Shao-Chen; Kim, Nam-Hyung

    2013-06-01

    During mitosis nucleation-promoting factors (NPFs) bind to the Arp2/3 complex and activate actin assembly. JMY and WAVE2 are two critical members of the NPFs. Previous studies have demonstrated that NPFs promote multiple processes such as cell migration and cytokinesis. However, the role of NPFs in development of mammalian embryos is still unknown. Results of the present study show that the NPFs JMY and WAVE2 are critical for cytokinesis during development of mouse embryos. Both JMY and WAVE2 are expressed in mouse embryos. After injection of JMY or WAVE2 siRNA, all embryos failed to develop to the morula or blastocyst stages. Moreover, using fluorescence intensity analysis, we found that the expression of actin decreased, and multiple nuclei were observed within a single cell indicating that NPFs-induced actin reduction caused the failure of cell division. In addition, injection of JMY and WAVE2 siRNA also caused ARP2 degradation, indicating that involvement of NPFs in development of mouse embryos is mainly through regulation of ARP2/3-induced actin assembly. Taken together, these data suggested that WAVE2 and JMY are involved in development of mouse embryos, and their regulation may be through a NPFs-Arp2/3-actin pathway.

  14. Cannabinoid receptor 1 signaling in embryo neurodevelopment

    PubMed Central

    Psychoyos, Delphine; Vinod, K. Yaragudri; Cao, Jin; Hyson, Richard L.; Wlodarczyk, Bogdan; He, Weimin; Cooper, Thomas B.; Hungund, Basalingappa L.; Finnell, Richard H.

    2014-01-01

    In utero exposure to THC, the psychoactive component of marijuana, is associated with an increased risk for neurodevelopmental defects in the offspring by interfering with the functioning of the endocannabinoid (eCB) system. At the present time it is not clearly known whether the eCB system is present prior to neurogenesis. Using an array of biochemical techniques we analyzed the levels of CB1 receptors, eCBs (AEA and 2-AG), and the enzymes (NAPE-PLD, DAGLα, DAGLβ MAGL and FAAH) involved in the metabolism of the eCBs in chick and mouse models during development. The findings demonstrate the presence of eCB system in early embryo, prior to neurogenesis. The eCB system might play a critical role in early embryogenesis and there might be adverse developmental consequences of in utero exposure to marijuana and other drugs of abuse during this period. PMID:22311661

  15. Chick stem cells: Current progress and future prospects

    PubMed Central

    Intarapat, Sittipon; Stern, Claudio D.

    2013-01-01

    Chick embryonic stem cells (cESCs) can be derived from cells obtained from stage X embryos (blastoderm stage); these have the ability to contribute to all somatic lineages in chimaeras, but not to the germ line. However, lines of stem cells that are able to contribute to the germ line can be established from chick primordial germ cells (cPGCs) and embryonic germ cells (cEGCs). This review provides information on avian stem cells, emphasizing different sources of cells and current methods for derivation and culture of pluripotent cells from chick embryos. We also review technologies for isolation and derivation of chicken germ cells and the production of transgenic birds. PMID:24103496

  16. The immunologic and antioxidant effects of L-phenylalanine on the uterine implantation of mice embryos during early pregnancy.

    PubMed

    Dong, Yulan; Bai, Yongping; Liu, Guanhui; Wang, Zixu; Cao, Jing; Chen, Yaoxing; Yang, Hongliang

    2014-10-01

    L-phenylalanine (L-PHE) is a synthetic precursor of catecholamines. Because it cannot be synthesised by an organism, it must be absorbed from the environment. Despite the wide use of L-PHE, whether L-PHE has a negative impact on embryo implantation and development is poorly understood. This study attempted to determine the roles of L-PHE in embryo implantation and development and in the immune response and antioxidant status of the uterus in early pregnancy mice injected intraperitoneally with 320 mg/kg L-PHE. The embryo number of treated mice decreased by 57.6%, and the size of their embryos was reduced by 2.8% (P⟩0.05) along the long diameter and 11.9% (P⟨0.05) along the short diameter at E9 compared with control mice. In addition, L-PHE significantly suppressed B lymphocyte proliferation. L-PHE increased IL-2 secretion but decreased the IL-4 concentration, thereby up-regulating the ratio of IL-2/IL-4 to 1.37-8.45. An analysis of the oxidant and antioxidant status showed that, compared with the control mice, the level of superoxide dismutase activity decreased by 21.54-39.94% and the glutathione peroxidase activity decreased by 15.27-18.96% among the L-PHE-treated mice at E1-E9. However, the malonaldehyde content increased by 14.29%-90.11% among the L-PHE-treated mice. Therefore, L-PHE impaired embryo implantation by disrupting cytokine-based immunity and oxidative stress in the uterus.

  17. Early life sensory ability-ventilatory responses of thornback ray embryos (Raja clavata) to predator-type electric fields.

    PubMed

    Ball, Rachel Emma; Oliver, Matthew Kenneth; Gill, Andrew Bruce

    2016-07-01

    Predator avoidance is fundamental for survival and it can be particularly challenging for prey animals if physical movement away from a predatory threat is restricted. Many sharks and rays begin life within an egg capsule that is attached to the sea bed. The vulnerability of this sedentary life stage is exacerbated in skates (Rajidae) as the compulsory ventilatory activity of embryos makes them conspicuous to potential predators. Embryos can reduce this risk by mediating ventilatory activity if they detect the presence of a predator using an acute electrosense. To determine how early in embryonic life predator elicited behavioral responses can occur, the reactions of three different age groups (1/3 developed, 2/3 developed, and near hatching) of embryonic thornback rays Raja clavata were tested using predator-type electric field stimuli. Egg capsules were exposed to continuous or intermittent stimuli in order to assess varying predator-type encounter scenarios on the ventilatory behavior of different developmental stages. All embryos reacted with a "freeze response" following initial electric field (E-field) exposure, ceasing ventilatory behavior in response to predator presence, demonstrating electroreceptive functionality for the first time at the earliest possible stage in ontogeny. This ability coincided with the onset of egg ventilatory behavior and may represent an effective means to enhance survival. A continuous application of stimuli over time revealed that embryos can adapt their behavior and resume normal activity, whereas when presented intermittently, the E-field resulted in a significant reduction in overall ventilatory activity across all ages. Recovery from stimuli was significantly quicker in older embryos, potentially indicative of the trade-off between avoiding predation and adequate respiration. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 721-729, 2016. © 2015 Wiley Periodicals, Inc.

  18. HoxB2, HoxB4 and Alx4 genes are downregulated in the cadmium-induced omphalocele in the chick model.

    PubMed

    Doi, Takashi; Puri, Prem; Bannigan, John; Thompson, Jennifer

    2010-10-01

    In the chick embryo, administration of cadmium (Cd) induces omphalocele phenotype. HoxB2 and HoxB4, expressed in cell types that contribute to ventral body wall (VBW) formation, act together to mediate proper closure of the VBW, involving a key downstream transcription factor, Alx4. HoxB2 and HoxB4 knockout mice display VBW defects with specific downregulation of Alx4 gene expression, while homozygous Alx4 knockouts show omphalocele phenotype. Although the earliest histological changes in the Cd chick model occur commencing at 4H post treatment, the exact timing and molecular mechanism by which Cd acts is still unclear. We hypothesized that HoxB2, HoxB4 and Alx4 genes are downregulated during the critical timing of very early embryogenesis in the Cd-induced omphalocele chick model. After 60H incubation, chick embryos were harvested at 1H, 4H and 8H after treatment with saline or Cd and divided into controls and Cd group (n = 24 for each group). RT-PCR was performed to investigate the gene expression of HoxB2, HoxB4 and Alx4 and statistically analyzed (significance was accepted at p < 0.05). Immunohistochemical staining was also performed to evaluate the protein expression/distribution of HoxB2, HoxB4 and Alx4 in the chick embryo. The expression levels of HoxB2, HoxB4 and Alx4 gene at 4H were significantly downregulated in the Cd group as compared to controls, whereas there were no significant differences at the other time points. Immunoreactivity of HoxB2, HoxB4 and Alx4 at 4H is also markedly decreased in the ectoderm and the dermomyotome in the Cd chick model as compared to controls. Downregulation of HoxB2, HoxB4 and Alx4 expression during the narrow window of early embryogenesis may cause omphalocele in the Cd chick model by interfering with molecular signaling required for proper VBW formation. Furthermore, these results support the concept that HoxB2, HoxB4 and Alx4 genes work together to mediate proper VBW formation.

  19. Pregnancy rates for embryos transferred from early postpartum beef cows into recipients with normal estrous cycles.

    PubMed

    Schrick, F N; Inskeep, E K; Butcher, R L

    1993-09-01

    Survival rate of embryos from first ovulations of postpartum cows with SHORT (6.9 +/- 0.3 days; n = 35) or NORMAL (17.1 +/- 0.3 days; n = 42) luteal phases and quality of the embryos on Day 6 were compared. At 19 to 23 days postpartum, cows were allotted to receive a norgestomet implant for 9 days (normal luteal phase) or to serve as untreated controls (short luteal phase). Calves were weaned 7 days after initiation of treatment to induce behavioral estrus in cows for mating. In 25 cows, growth of the ovulatory follicle was monitored by ultrasonography. On Day 6 after estrus, embryos were recovered nonsurgically, and live embryos were transferred into recipient cows exhibiting normal estrous cycles. The medium used to flush the embryos from the uterus of each donor cow was assayed for prostaglandin F2 alpha (PGF2 alpha). Days from calf removal to estrus and size of ovulatory follicles at ovulation (4.1 +/- 0.3 days and 16.7 +/- 0.7 mm, respectively) did not differ between NORMAL and SHORT cows. Interval from detection of the ovulatory follicle to ovulation was longer in NORMAL (10 +/- 0.7 days) than in SHORT cows (8 +/- 0.6 days; p < 0.05). Rates of recovery of an embryo or ovum (64%), rates of fertilization (65%), and quality or stage of development of Day 6 embryos did not differ between SHORT and NORMAL cows. Overall pregnancy rate from recovered oocytes was 13% for SHORT and 32% for NORMAL cows (p = 0.06); survival of fertilized oocytes was 23% for SHORT and 47% for NORMAL cows (p = 0.08).(ABSTRACT TRUNCATED AT 250 WORDS)

  20. The Potential Role of As-sumo-1 in the Embryonic Diapause Process and Early Embryo Development of Artemia sinica

    PubMed Central

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

    2014-01-01

    During embryonic development of Artemia sinica, environmental stresses induce the embryo diapause phenomenon, required to resist apoptosis and regulate cell cycle activity. The small ubiquitin-related modifier-1 (SUMO), a reversible post-translational protein modifier, plays an important role in embryo development. SUMO regulates multiple cellular processes, including development and other biological processes. The molecular mechanism of diapause, diapause termination and the role of As-sumo-1 in this processes and in early embryo development of Artemia sinica still remains unknown. In this study, the complete cDNA sequences of the sumo-1 homolog, sumo ligase homolog, caspase-1 homolog and cyclin B homolog from Artemia sinica were cloned. The mRNA expression patterns of As-sumo-1, sumo ligase, caspase-1, cyclin B and the location of As-sumo-1 were investigated. SUMO-1, p53, Mdm2, Caspase-1, Cyclin B and Cyclin E proteins were analyzed during different developmental stages of the embryo of A. sinica. Small interfering RNA (siRNA) was used to verify the function of sumo-1 in A. sinica. The full-length cDNA of As-sumo-1 was 476 bp, encoding a 92 amino acid protein. The As-caspases-1 cDNA was 966 bp, encoding a 245 amino-acid protein. The As-sumo ligase cDNA was 1556 bp encoding, a 343 amino acid protein, and the cyclin B cDNA was 739 bp, encoding a 133 amino acid protein. The expressions of As-sumo-1, As-caspase-1 and As-cyclin B were highest at the 10 h stage of embryonic development, and As-sumo ligase showed its highest expression at 0 h. The expression of As-SUMO-1 showed no tissue or organ specificity. Western blotting showed high expression of As-SUMO-1, p53, Mdm2, Caspase-1, Cyclin B and Cyclin E at the 10 h stage. The siRNA caused abnormal development of the embryo, with increased malformation and mortality. As-SUMO-1 is a crucial regulation and modification protein resumption of embryonic diapause and early embryo development of A. sinica. PMID:24404204

  1. Use of the Coelomic Grafting Technique for Prolonged ex utero Cultivation of Late Preprimitive Streak-Stage Rabbit Embryos.

    PubMed

    Püschel, Bernd; Männer, Jörg

    2016-01-01

    Due to its morphological similarity with the early human embryo, the pregastrulation-stage rabbit may represent an appropriate mammalian model for studying processes involved in early human development. The usability of mammalian embryos for experimental studies depends on the availability of whole embryo culture methods facilitating prolonged ex utero development. While currently used culture methods yield high success rates for embryos from primitive streak stages onward, the success rate of extended cultivation of preprimitive streak-stage mammalian embryos is low for all previously established methods and for all studied species. This limits the usability of preprimitive streak-stage rabbit embryos in experimental embryology. We have tested whether the extraembryonic coelom of 4-day-old chick embryos may be used for prolonged ex utero culture of preprimitive streak-stage rabbit embryos (stage 2, 6.2 days post coitum). We found that, within this environment, stage 2 rabbit blastocysts can be cultured at decreasing success rates (55% after 1 day, 35% after 2 days, 15% after 3 days) up to a maximum of 72 h. Grafted blastocysts can continue development from the onset of gastrulation to early organogenesis and thereby form all structures characterizing age-matched controls (e.g. neural tube, somites, beating heart). Compared to normal controls, successfully cultured embryos developed at a slower rate and finally showed some structural and gross morphological anomalies. The method presented here was originally developed for whole embryo culture of mouse embryos by Gluecksohn-Schoenheimer in 1941. It is a simple and inexpensive method that may represent a useful extension to presently available ex utero culture systems for rabbit embryos. © 2016 S. Karger AG, Basel.

  2. Quantitative imaging of lipids in live mouse oocytes and early embryos using CARS microscopy

    PubMed Central

    Bradley, Josephine; Pope, Iestyn; Masia, Francesco; Sanusi, Randa; Langbein, Wolfgang; Borri, Paola

    2016-01-01

    Mammalian oocytes contain lipid droplets that are a store of fatty acids, whose metabolism plays a substantial role in pre-implantation development. Fluorescent staining has previously been used to image lipid droplets in mammalian oocytes and embryos, but this method is not quantitative and often incompatible with live cell imaging and subsequent development. Here we have applied chemically specific, label-free coherent anti-Stokes Raman scattering (CARS) microscopy to mouse oocytes and pre-implantation embryos. We show that CARS imaging can quantify the size, number and spatial distribution of lipid droplets in living mouse oocytes and embryos up to the blastocyst stage. Notably, it can be used in a way that does not compromise oocyte maturation or embryo development. We have also correlated CARS with two-photon fluorescence microscopy simultaneously acquired using fluorescent lipid probes on fixed samples, and found only a partial degree of correlation, depending on the lipid probe, clearly exemplifying the limitation of lipid labelling. In addition, we show that differences in the chemical composition of lipid droplets in living oocytes matured in media supplemented with different saturated and unsaturated fatty acids can be detected using CARS hyperspectral imaging. These results demonstrate that CARS microscopy provides a novel non-invasive method of quantifying lipid content, type and spatial distribution with sub-micron resolution in living mammalian oocytes and embryos. PMID:27151947

  3. Quantitative imaging of lipids in live mouse oocytes and early embryos using CARS microscopy.

    PubMed

    Bradley, Josephine; Pope, Iestyn; Masia, Francesco; Sanusi, Randa; Langbein, Wolfgang; Swann, Karl; Borri, Paola

    2016-06-15

    Mammalian oocytes contain lipid droplets that are a store of fatty acids, whose metabolism plays a substantial role in pre-implantation development. Fluorescent staining has previously been used to image lipid droplets in mammalian oocytes and embryos, but this method is not quantitative and often incompatible with live cell imaging and subsequent development. Here we have applied chemically specific, label-free coherent anti-Stokes Raman scattering (CARS) microscopy to mouse oocytes and pre-implantation embryos. We show that CARS imaging can quantify the size, number and spatial distribution of lipid droplets in living mouse oocytes and embryos up to the blastocyst stage. Notably, it can be used in a way that does not compromise oocyte maturation or embryo development. We have also correlated CARS with two-photon fluorescence microscopy simultaneously acquired using fluorescent lipid probes on fixed samples, and found only a partial degree of correlation, depending on the lipid probe, clearly exemplifying the limitation of lipid labelling. In addition, we show that differences in the chemical composition of lipid droplets in living oocytes matured in media supplemented with different saturated and unsaturated fatty acids can be detected using CARS hyperspectral imaging. These results demonstrate that CARS microscopy provides a novel non-invasive method of quantifying lipid content, type and spatial distribution with sub-micron resolution in living mammalian oocytes and embryos. © 2016. Published by The Company of Biologists Ltd.

  4. Maternal hCG concentrations in early IVF pregnancies: associations with number of cells in the Day 2 embryo and oocytes retrieved.

    PubMed

    Tanbo, T G; Eskild, A

    2015-12-01

    Do number of cells in the transferred cleavage stage embryo and number of oocytes retrieved for IVF influence maternal hCG concentrations in early pregnancies? Compared with transfer of a 2-cell embryo, transfer of a 4-cell embryo results in higher hCG concentrations on Day 12 after transfer, and more than 20 oocytes retrieved were associated with low hCG concentrations. Maternal hCG concentration in very early pregnancy varies considerably among women, but is likely to be an indicator of time since implantation of the embryo into the endometrium, in addition to number and function of trophoblast cells. We followed 1047 pregnancies after IVF/ICSI from oocyte retrieval until Day 12 after embryo transfer. Women were recruited in Norway during the years 2005-2013. Successful pregnancies after transfer of one single embryo that had been cultured for 2 days were included. Maternal hCG was quantified on Day 12 after embryo transfer by chemiluminescence immunoassay, which measures intact hCG and the free β-hCG chain. Information on a successful pregnancy, defined as birth after >16 weeks, was obtained by linkage to the Medical Birth Registry of Norway. Transfer of a 4-cell embryo resulted in higher maternal hCG concentrations compared with transfer of a 2-cell embryo (134.8 versus 87.8 IU/l, P < 0.05). A high number of oocytes retrieved (>20) was associated with low hCG concentrations (P < 0.05). The factors studied explain a limited part of the total variation of hCG concentrations in early pregnancy. Although embryo transfer was performed at the same time after fertilization, we do not know the exact time of implantation. A further limitation to our study is that the number of pregnancies after transfer of a 2-cell embryo was small (27 cases). Number of cells in the transferred embryo and number of oocytes retrieved may influence the conditions and timing for embryo implantation in different ways and thereby influence maternal hCG concentrations. Such knowledge may be

  5. Low dose trichloroethylene alters cytochrome P450 - 2C subfamily expression in the developing chick heart

    PubMed Central

    Makwana, Om; Ahles, Lauren; Lencinas, Alejandro; Selmin, Ornella I.; Runyan, Raymond B.

    2013-01-01

    Trichloroethylene (TCE) is an organic solvent and common environmental contaminant. TCE exposure is associated with heart defects in humans and animal models. Primary metabolism of TCE in adult rodent models is by specific hepatic cytochrome P450 enzymes (Lash et al., 2000). As association of TCE exposure with cardiac defects is in exposed embryos prior to normal liver development, we investigated metabolism of TCE in the early embryo. Developing chick embryos were dosed in ovo with environmentally relevant doses of TCE (8 ppb and 800 ppb) and RNA was extracted from cardiac and extra-cardiac tissue (whole embryo without heart). Real time PCR showed upregulation of CYP2H1 transcripts in response to TCE exposure in the heart. No detectable cytochrome expression was found in extra-cardiac tissue. As seen previously, the dose response was non-monotonic and 8ppb elicited stronger upregulation than 800 ppb. Immunostaining for CYP2C subfamily expression confirmed protein expression and showed localization in both myocardium and endothelium. TCE exposure increased protein expression in both tissues. These data demonstrate that the earliest embryonic expression of phase I detoxification enzymes is in the developing heart. Expression of these CYPs is likely to be relevant to the susceptibility of the developing heart to environmental teratogens. PMID:22855351

  6. 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.

  7. Comparative gene expression analysis of Dtg, a novel target gene of Dpp signaling pathway in the early Drosophila melanogaster embryo.

    PubMed

    Hodar, Christian; Zuñiga, Alejandro; Pulgar, Rodrigo; Travisany, Dante; Chacon, Carlos; Pino, Michael; Maass, Alejandro; Cambiazo, Verónica

    2014-02-10

    In the early Drosophila melanogaster embryo, Dpp, a secreted molecule that belongs to the TGF-β superfamily of growth factors, activates a set of downstream genes to subdivide the dorsal region into amnioserosa and dorsal epidermis. Here, we examined the expression pattern and transcriptional regulation of Dtg, a new target gene of Dpp signaling pathway that is required for proper amnioserosa differentiation. We showed that the expression of Dtg was controlled by Dpp and characterized a 524-bp enhancer that mediated expression in the dorsal midline, as well as, in the differentiated amnioserosa in transgenic reporter embryos. This enhancer contained a highly conserved region of 48-bp in which bioinformatic predictions and in vitro assays identified three Mad binding motifs. Mutational analysis revealed that these three motifs were necessary for proper expression of a reporter gene in transgenic embryos, suggesting that short and highly conserved genomic sequences may be indicative of functional regulatory regions in D. melanogaster genes. Dtg orthologs were not detected in basal lineages of Dipterans, which unlike D. melanogaster develop two extra-embryonic membranes, amnion and serosa, nevertheless Dtg orthologs were identified in the transcriptome of Musca domestica, in which dorsal ectoderm patterning leads to the formation of a single extra-embryonic membrane. These results suggest that Dtg was recruited as a new component of the network that controls dorsal ectoderm patterning in the lineage leading to higher Cyclorrhaphan flies, such as D. melanogaster and M. domestica. Copyright © 2013 Elsevier B.V. All rights reserved.

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

    PubMed

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

    2013-06-01

    The avian embryo provides a multifaceted model to study developmental mechanisms because of its accessibility to microsurgery, fluorescence cell labeling, in vivo imaging, and molecular manipulation. Early two-dimensional planar growth of the avian embryo mimics human development and provides unique access to complex cell migration patterns using light microscopy. Later developmental events continue to permit access to both light and other imaging modalities, making the avian embryo an excellent model for developmental imaging. For example, significant insights into cell and tissue behaviors within the primitive streak, craniofacial region, and cardiovascular and peripheral nervous systems have come from avian embryo studies. In this review, we provide an update to recent advances in embryo and tissue slice culture and imaging, fluorescence cell labeling, and gene profiling. We focus on how technical advances in the chick and quail provide a clearer understanding of how embryonic cell dynamics are beautifully choreographed in space and time to sculpt cells into functioning structures. We summarize how these technical advances help us to better understand basic developmental mechanisms that may lead to clinical research into human birth defects and tissue repair. Copyright © 2013 Wiley Periodicals, Inc.

  9. Inactivation of the Huntington's disease gene (Hdh) impairs anterior streak formation and early patterning of the mouse embryo

    PubMed Central

    Woda, Juliana M; Calzonetti, Teresa; Hilditch-Maguire, Paige; Duyao, Mabel P; Conlon, Ronald A; MacDonald, Marcy E

    2005-01-01

    Background Huntingtin, the HD gene encoded protein mutated by polyglutamine expansion in Huntington's disease, is required in extraembryonic tissues for proper gastrulation, implicating its activities in nutrition or patterning of the developing embryo. To test these possibilities, we have used whole mount in situ hybridization to examine embryonic patterning and morphogenesis in homozygous Hdhex4/5 huntingtin deficient embryos. Results In the absence of huntingtin, expression of nutritive genes appears normal but E7.0–7.5 embryos exhibit a unique combination of patterning defects. Notable are a shortened primitive streak, absence of a proper node and diminished production of anterior streak derivatives. Reduced Wnt3a, Tbx6 and Dll1 expression signify decreased paraxial mesoderm and reduced Otx2 expression and lack of headfolds denote a failure of head development. In addition, genes initially broadly expressed are not properly restricted to the posterior, as evidenced by the ectopic expression of Nodal, Fgf8 and Gsc in the epiblast and T (Brachyury) and Evx1 in proximal mesoderm derivatives. Despite impaired posterior restriction and anterior streak deficits, overall anterior/posterior polarity is established. A single primitive streak forms and marker expression shows that the anterior epiblast and anterior visceral endoderm (AVE) are specified. Conclusion Huntingtin is essential in the early patterning of the embryo for formation of the anterior region of the primitive streak, and for down-regulation of a subset of dynamic growth and transcription factor genes. These findings provide fundamental starting points for identifying the novel cellular and molecular activities of huntingtin in the extraembryonic tissues that govern normal anterior streak development. This knowledge may prove to be important for understanding the mechanism by which the dominant polyglutamine expansion in huntingtin determines the loss of neurons in Huntington's disease. PMID:16109169

  10. Inactivation of the Huntington's disease gene (Hdh) impairs anterior streak formation and early patterning of the mouse embryo.

    PubMed

    Woda, Juliana M; Calzonetti, Teresa; Hilditch-Maguire, Paige; Duyao, Mabel P; Conlon, Ronald A; MacDonald, Marcy E

    2005-08-18

    Huntingtin, the HD gene encoded protein mutated by polyglutamine expansion in Huntington's disease, is required in extraembryonic tissues for proper gastrulation, implicating its activities in nutrition or patterning of the developing embryo. To test these possibilities, we have used whole mount in situ hybridization to examine embryonic patterning and morphogenesis in homozygous Hdh(ex4/5) huntingtin deficient embryos. In the absence of huntingtin, expression of nutritive genes appears normal but E7.0-7.5 embryos exhibit a unique combination of patterning defects. Notable are a shortened primitive streak, absence of a proper node and diminished production of anterior streak derivatives. Reduced Wnt3a, Tbx6 and Dll1 expression signify decreased paraxial mesoderm and reduced Otx2 expression and lack of headfolds denote a failure of head development. In addition, genes initially broadly expressed are not properly restricted to the posterior, as evidenced by the ectopic expression of Nodal, Fgf8 and Gsc in the epiblast and T (Brachyury) and Evx1 in proximal mesoderm derivatives. Despite impaired posterior restriction and anterior streak deficits, overall anterior/posterior polarity is established. A single primitive streak forms and marker expression shows that the anterior epiblast and anterior visceral endoderm (AVE) are specified. Huntingtin is essential in the early patterning of the embryo for formation of the anterior region of the primitive streak, and for down-regulation of a subset of dynamic growth and transcription factor genes. These findings provide fundamental starting points for identifying the novel cellular and molecular activities of huntingtin in the extraembryonic tissues that govern normal anterior streak development. This knowledge may prove to be important for understanding the mechanism by which the dominant polyglutamine expansion in huntingtin determines the loss of neurons in Huntington's disease.

  11. Reduced heart rate and cardiac output differentially affect angiogenesis, growth, and development in early chicken embryos (Gallus domesticus).

    PubMed

    Branum, Sylvia R; Yamada-Fisher, Miho; Burggren, Warren

    2013-01-01

    An increase in both vascular circumferential tension and shear stress in the developing vasculature of the chicken embryo has been hypothesized to stimulate angiogenesis in the developing peripheral circulation chorioallantoic membrane (CAM). To test this hypothesis, angiogenesis in the CAM, development, and growth were measured in the early chicken embryo, following acute and chronic topical application of the purely bradycardic drug ZD7288. At hour 56, ZD7288 reduced heart rate (f(H)) by ~30% but had no significant effect on stroke volume (~0.19 ± 0.2 μL), collectively resulting in a significant fall in cardiac output (CO) from ~27 ± 3 to 18 ± 2 μL min(-1). Mean f(H) at 72 h of development was similarly significantly lowered by acute ZD7288 treatment (250 μM) to 128 ± 0.3 beats min(-1), compared with 174.5 ± 0.3 and 174.7 ± 0.8 beats min(-1) in control and Pannett-Compton (P-C) saline-treated embryos, respectively. Chronic dosing with ZD7288-and the attendant decreases in f(H) and CO-did not change eye diameter or cervical flexion (key indicators of development rate) at 120 h but significantly reduced overall growth (wet and dry body mass decreased by 20%). CAM vessel density index (reflecting angiogenesis) measured 200-400 μm from the umbilical stalk was not altered, but ZD7288 reduced vessel numbers-and therefore vessel density-by 13%-16% more distally (500-600 μm from umbilical stalk) in the CAM. In the ZD7288-treated embryos, a decrease in vessel length was found within the second branch order (~300-400 μm from the umbilical stock), while a decrease in vessel diameter was found closer to the umbilical stock, beginning in the first branch order (~200-300 μm). Paradoxically, chronic application of P-C saline also reduced peripheral CAM vessel density index at 500 and 600 μm by 13% and 7%, respectively, likely from washout of local angiogenic factors. In summary, decreased f(H) with reduced CO did not slow development rate but reduced embryonic

  12. Size-dependent regulation of dorsal-ventral patterning in the early Drosophila embryo

    PubMed Central

    Garcia, Mayra; Nahmad, Marcos; Reeves, Gregory T.; Stathopoulos, Angelike

    2013-01-01

    How natural variation in embryo size affects patterning of the Drosophila embryo dorsal-ventral (DV) axis is not known. Here we examined quantitatively the relationship between nuclear distribution of the Dorsal transcription factor, boundary positions for several target genes, and DV axis length. Data were obtained from embryos of a wild-type background as well as from mutant lines inbred to size select embryos of smaller or larger sizes. Our data show that the width of the nuclear Dorsal gradient correlates with DV axis length. In turn, for some genes expressed along the DV axis, the boundary positions correlate closely with nuclear Dorsal levels and with DV axis length; while the expression pattern of others is relatively constant and independent of the width of the Dorsal gradient. In particular, the patterns of snail (sna) and ventral nervous-system defective (vnd) correlate with nuclear Dorsal levels and exhibit scaling to DV length; while the pattern of intermediate neuroblasts defective (ind) remains relatively constant with respect to changes in Dorsal and DV length. However, in mutants that exhibit an abnormal expansion of the Dorsal gradient which fails to scale to DV length, only sna follows the Dorsal distribution and exhibits overexpansion; in contrast, vnd and ind do not overexpand suggesting some additional mechanism acts to refine the dorsal boundaries of these two genes. Thus, our results argue against the idea that the Dorsal gradient works as a global system of relative coordinates along the DV axis and suggest that individual targets respond to changes in embryo size in a gene-specific manner. PMID:23800450

  13. Recognition of the CDEI motif GTCACATG by mouse nuclear proteins and interference with the early development of the mouse embryo.

    PubMed Central

    Blangy, A; Léopold, P; Vidal, F; Rassoulzadegan, M; Cuzin, F

    1991-01-01

    We have reported previously (1) two unexpected consequences of the microinjection into fertilized mouse eggs of a recombinant plasmid designated p12B1, carrying a 343 bp insert of non-repetitive mouse DNA. Injected at very low concentrations, this plasmid could be established as an extrachromosomal genetic element. When injected in greater concentration, an early arrest of embryonic development resulted. In the present work, we have studied this toxic effect in more detail by microinjecting short synthetic oligonucleotides with sequences from the mouse insert. Lethality was associated with the nucleotide sequence GTCACATG, identical with the CDEl element of yeast centromeres. Development of injected embryos was arrested between the one-cell and the early morula stages, with abnormal structures and DNA contents. Electrophoretic mobility shift and DNAse foot-printing assays demonstrated the binding of mouse nuclear protein(s) to the CDEl-like box. Base changes within the CDEl sequence prevented both the toxic effects in embryos and the formation of protein complex in vitro, suggesting that protein binding at such sites in chromosomal DNA plays an important role in early development. Images PMID:1766880

  14. Heterogeneous nuclear ribonucleoprotein C1 may control miR-30d levels in endometrial exosomes affecting early embryo implantation.

    PubMed

    Balaguer, N; Moreno, I; Herrero, M; González, M; Simón, C; Vilella, F

    2018-05-29

    as a model of hEECs in silencing experiments due to the low survival rates of primary hEECs after transfection. The data show that hnRNPC1 may be involved in the internalization of miR-30d inside Exosomes. The decreased rates of embryo adhesion in endometrial epithelial-like cells transiently silenced with sihnRNPC1evidence that hnRNPC1 could be an important player in the maternal-embryo communication established in the early stages of implantation. This work was supported by the Miguel Servet Program Type I of Instituto de Salud Carlos III [CP13/00038]; FIS project [PI14/00545] to FV; the 'Atracció de Talent' Program from VLC-CAMPUS [UV-INV-PREDOC14-178329 to NB]; a Torres-Quevedo grant (PTQ-13-06133) by the Spanish Ministry of Economy and Competitiveness to IM; and MINECO/FEDER Grant [SAF2015-67154-R] to CS. The authors declare there is no conflict of interest.

  15. Effect of early addition of bone morphogenetic protein 5 (BMP5) to embryo culture medium on in vitro development and expression of developmentally important genes in bovine preimplantation embryos.

    PubMed

    García, Elina V; Miceli, Dora C; Rizo, Gabriela; Valdecantos, Pablo A; Barrera, Antonio D

    2015-09-01

    Previous studies have reported that bone morphogenetic protein 5 (BMP5) is differentially expressed in the isthmus of bovine oviducts and it is present in the oviductal fluid. However, the specific action of this factor is unknown. To evaluate whether BMP5 exerts some effect during early bovine embryo development, gene expression of BMP5, BMP receptors, and the effect of exogenous BMP5 on in vitro development and expression of developmentally important genes were assessed. In experiment 1, pools of embryos at two-cell, four-cell, eight-cell, and blastocyst stages, derived from in vitro fertilization, were collected for analysis of BMP5 and BMP receptors (BMPR1A, BMPR1B, and BMPR2) messenger RNA (mRNA) expression. On the basis of previous results, in experiment 2, presumptive zygotes were cultured for the first 48 hours after insemination in CR1aa medium assaying three different treatments: (1) control (CR1aa); (2) vehicle control (CR1aa + 0.04 mM HCl), and (3) BMP5 treatment (CR1aa + 100 ng/mL of BMP5). The cleavage rate was evaluated 48 hours after insemination (Day 2), and then, embryos were transferred to CR1aa + 10% fetal bovine serum. The blastocyst rate was determined on Day 7. In experiment 3, pools of embryos at two-cell, four-cell, eight-cell, and blastocyst stages, derived from control and BMP5-treated groups, were collected for analysis of ID2 (BMP target gene), OCT4, NANOG, and SOX2 (pluripotency genes) mRNA expression. BMP5 transcripts were not detectable in any of the embryonic stages examined, whereas the relative mRNA abundance of the three BMP receptors analyzed was greater in early embryo development stages before maternal-embryonic transition, raising the possibility of a direct effect of exogenous BMPs on the embryo during the first developmental period. Although early addition of 100 ng/mL of BMP5 to the embryo culture medium had no effect on the cleavage rate, a significantly higher proportion of cleaved embryos developed to the

  16. Production of somatic chimera chicks by injection of bone marrow cells into recipient blastoderms.

    PubMed

    Heo, Young Tae; Lee, Sung Ho; Kim, Teoan; Kim, Nam Hyung; Lee, Hoon Taek

    2012-01-01

    Several types of cells, including blastoderm cells, primordial germ cells, and embryonic germ cells were injected into early-stage recipient embryos to produce chimera avians and to gain insights into cell development. However, a limited number of studies of avian adult stem cells have also been conducted. This study is, to the best of our knowledge, the first to evaluate chicken bone marrow cells' (chBMC) ability to differentiate into multiple cell lineages and capability to generate chimera chicks. We induced random differentiation of chBMCs in vitro and injected immunologically selected pluripotent cells in chBMCs into the blastoderms of recipient eggs. The multipotency of BMCs from the barred Plymouth rock (BPR) was confirmed via AP staining, RT-PCR, immunocytochemistry, and FACS using specific markers, such as Oct-4 and SSEA-1, 3 and 4. Isolated chBMCs were found to be able to induce in vitro differentiation to multiple cell lineages. Approximately 5,000 chBMCs were injected into the blastoderms of white leghorn (WL) recipients and proved able to contribute to the generation of somatic chimera chicks with a frequency of 2.7% (2 of 73). Confirmation of chimerism in hatched chicks was achieved via PCR analysis using D-loop-specific primers of BPR and WL. Our study demonstrated the successful production of chimera chicks using chBMC. Therefore, we propose that the use of adult chBMCs may constitute a new possible approach to the production of chimera poultry, and may provide helpful studies in avian developmental biology.

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

    PubMed

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

    2016-01-01

    Does ulipristal acetate (UPA), a selective progesterone receptor modulator used for emergency contraception (EC), interfere with fertilization or early embryo development in vitro and in vivo? At doses similar to those used for EC, UPA does not affect mouse gamete transport, fertilization or embryo development. UPA acts as an emergency contraceptive mainly by inhibiting or delaying ovulation. However, there is little information regarding its effects on post-ovulatory events preceding implantation. This was an in vitro and in vivo experimental study involving the use of mouse gametes and embryos from at least three animals in each set of experiments. For in vitro fertilization experiments, mouse epididymal spermatozoa capacitated in the presence of different concentrations of UPA (0-1000 ng/ml) were used to inseminate cumulus-intact or cumulus-free eggs in the presence or absence of UPA during gamete co-incubation, and the percentage of fertilized eggs was determined. For in vivo fertilization experiments, superovulated females caged with proven fertile males were injected with UPA (40 mg/kg) or vehicle just before or just after mating and the percentage of fertilized eggs recovered from the ampulla was determined. To investigate the effect of UPA on embryo development, zygotes were recovered from mated females, cultured in the presence of UPA (1000 ng/ml) for 4 days and the progression of embryo development was monitored daily. In vitro studies revealed that the presence of UPA during capacitation and/or gamete co-incubation does not affect fertilization. Whereas the in vivo administration of UPA at the same time as hCG injection produced a decrease in the number of eggs ovulated compared with controls (vehicle injected animals, P < 0.05), no effects on fertilization were observed when UPA was administered shortly before or after mating. No differences were observed in either the percentage of cleaved embryos or the cleavage speed when UPA was present during in

  18. [Molecular cloning, expression of rat Msx-1 and Msx-2 during early embryo genesis and roles for mandibular chondrogenesis].

    PubMed

    Ishiguro, S

    1999-03-01

    Quail-chick chimera experiments have shown a contribution of carnial neural crest cells to the craniofacial skeletal elements. Moreover, tissue interactions between epithelial-mesenchymal interaction during early facial process development are required for both skeletal differentiation and morphogenesis. In this study, it was observed that Msx homeobox containing genes expressed in the facial process were important molecules of cartilage morphogenesis. Rat cDNAs were isolated and encoded by Msx-1 and -2, and then the expression patterns using in situ hybridization were investigated during early rat face development. These genes were correlatively expressed in the cranial neural crest forming area (E 9.5 dpc) and the facial process (E 12.5 dpc). Antisence inhibition of Msx genes in the E 12.5 mandibular process exhibited the alteration of their gene expression and cartilage patterns. Antisence inhibition of Msx-1 induced lack of the medial portion of cartilage, and antisence inhibition of Msx-2 enhanced chondrogenesis of mandibular process under the organ culture condition. Thus it was concluded that expression of Msx genes during mandibular process development comprises important signals of chondrogenesis.

  19. Development of the embryonic heat shock response and the impact of repeated thermal stress in early stage lake whitefish (Coregonus clupeaformis) embryos.

    PubMed

    Whitehouse, Lindy M; McDougall, Chance S; Stefanovic, Daniel I; Boreham, Douglas R; Somers, Christopher M; Wilson, Joanna Y; Manzon, Richard G

    2017-10-01

    Lake whitefish (Coregonus clupeaformis) embryos were exposed to thermal stress (TS) at different developmental stages to determine when the heat shock response (HSR) can be initiated and if it is altered by exposure to repeated TS. First, embryos were subject to one of three different TS temperatures (6, 9, or 12°C above control) at 4 points in development (21, 38, 60 and 70 days post-fertilisation (dpf)) for 2h followed by a 2h recovery to understand the ontogeny of the HSR. A second experiment explored the effects of repeated TS on the HSR in embryos from 15 to 75 dpf. Embryos were subjected to one of two TS regimes; +6°C TS for 1h every 6 days or +9°C TS for 1h every 6 days. Following a 2h recovery, a subset of embryos was sampled. Our results show that embryos could initiate a HSR via upregulation of heat shock protein 70 (hsp70) mRNA at all developmental ages studied, but that this response varied with age and was only observed with a TS of +9 or +12°C. In comparison, when embryos received multiple TS treatments, hsp70 was not induced in response to the 1h TS and 2h recovery, and a downregulation was observed at 39 dpf. Downregulation of hsp47 and hsp90α mRNA was also observed in early age embryos. Collectively, these data suggest that embryos are capable of initiating a HSR at early age and throughout embryogenesis, but that repeated TS can alter the HSR, and may result in either reduced responsiveness or a downregulation of inducible hsps. Our findings warrant further investigation into both the short- and long-term effects of repeated TS on lake whitefish development. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Overexpression of S-adenosylmethionine decarboxylase (SAMDC) in early Xenopus embryos induces cell dissociation and inhibits transition from the blastula to gastrula stage.

    PubMed

    Shibata, M; Shinga, J; Yasuhiko, Y; Kai, M; Miura, K; Shimogori, T; Kashiwagi, K; Igarashi, K; Shiokawa, K

    1998-07-01

    Xenopus early embryos contain relatively low levels of S-adenosyl-methionine decarboxylase (SAMDC) and its mRNA. When SAMDC mRNA was injected into Xenopus embryos, it was preserved until the blastula stage and induced a large increase in SAMDC activity. The SAMDC-overexpressed embryos developed normally until the blastula stage but at the early gastrula stage cells which received the mRNA, dissociated autonomously and stopped synthesizing protein. In a hypotonic medium, the dissociated cells, and hence whole embryos, autolyzed. However, in isotonic media dissociated cells did not autolyze, although they did not divide and their DNA and RNA synthesis activity was greatly inhibited. The effects of SAMDC overexpression were abolished by coinjection of ethylglyoxal-bis(guanylhydrazone) (EGBG), a specific inhibitor of SAMDC. In SAMDC-overexpressed embryos the level of putrescine decreased and that of spermidine increased, though to limited extents, resulting in a considerable decrease in the putrescine/spermidine ratio. However, direct injection of spermidine did not mimic the effect of SAMDC overexpression, and putrescine coinjected with SAMDC mRNA to maintain the normal putrescine/spermidine ratio did not rescue the embryos. Conversely, the level of S-adenosylmethionine (SAM) greatly decreased and coinjection of SAM, which restored the level of SAM, rescued the embryos. We concluded that in SAMDC-overexpressed embryos a SAM-deficient state was induced and this caused cell dissociation and inhibition of transition from the blastula to gastrula stage. We suggest that the SAM-deficient embryos obtained in the present study provide a unique system for studying the cellular control mechanism underlying the blastula-gastrula transition.

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

    PubMed

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

    2015-10-01

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

  2. Early molecular events involved in Pinus pinaster Ait. somatic embryo development under reduced water availability: transcriptomic and proteomic analyses.

    PubMed

    Morel, Alexandre; Teyssier, Caroline; Trontin, Jean-François; Eliášová, Kateřina; Pešek, Bedřich; Beaufour, Martine; Morabito, Domenico; Boizot, Nathalie; Le Metté, Claire; Belal-Bessai, Leila; Reymond, Isabelle; Harvengt, Luc; Cadene, Martine; Corbineau, Françoise; Vágner, Martin; Label, Philippe; Lelu-Walter, Marie-Anne

    2014-09-01

    Maritime pine somatic embryos (SEs) require a reduction in water availability (high gellan gum concentration in the maturation medium) to reach the cotyledonary stage. This key switch, reported specifically for pine species, is not yet well understood. To facilitate the use of somatic embryogenesis for mass propagation of conifers, we need a better understanding of embryo development. Comparison of both transcriptome (Illumina RNA sequencing) and proteome [two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis with mass spectrometry (MS) identification] of immature SEs, cultured on either high (9G) or low (4G) gellan gum concentration, was performed, together with analysis of water content, fresh and dry mass, endogenous abscisic acid (ABA; gas chromatography-MS), soluble sugars (high-pressure liquid chromatography), starch and confocal laser microscope observations. This multiscale, integrated analysis was used to unravel early molecular and physiological events involved in SE development. Under unfavorable conditions (4G), the glycolytic pathway was enhanced, possibly in relation to cell proliferation that may be antagonistic to SE development. Under favorable conditions (9G), SEs adapted to culture constraint by activating specific protective pathways, and ABA-mediated molecular and physiological responses promoting embryo development. Our results suggest that on 9G, germin-like protein and ubiquitin-protein ligase could be used as predictive markers of SE development, whereas protein phosphatase 2C could be a biomarker for culture adaptive responses. This is the first characterization of early molecular mechanisms involved in the development of pine SEs following an increase in gellan gum concentration in the maturation medium, and it is also the first report on somatic embryogenesis in conifers combining transcriptomic and proteomic datasets. © 2014 Scandinavian Plant Physiology Society.

  3. Selective loss of mouse embryos due to the expression of transgenic major histocompatibility class I molecules early in embryogenesis.

    PubMed

    Aït-Azzouzene, D; Langkopf, A; Cohen, J; Bleux, C; Gendron, M C; Kanellopoulos-Langevin, C

    1998-05-01

    Among the numerous hypotheses proposed to explain the absence of fetal rejection by the mother in mammals, it has been suggested that regulation of expression of the polymorphic major histocompatibility complex (MHC) at the fetal-maternal interface plays a major role. In addition to a lack of MHC gene expression in the placenta throughout gestation, the absence of polymorphic MHC molecules on the early embryo, as well as their low level of expression after midgestation, could contribute to this important biologic phenomenon. In order to test this hypothesis, we have produced transgenic mice able to express polymorphic MHC class I molecules early in embryogenesis. We have placed the MHC class la gene H-2Kb under the control of a housekeeping gene promoter, the hydroxy-methyl-glutaryl coenzyme A reductase (HMG) gene minimal promoter. This construct has been tested for functionality after transfection into mouse fibroblast L cells. The analysis of three founder transgenic mice and their progeny suggested that fetoplacental units that could express the H-2Kb heavy chains are unable to survive in utero beyond midgestation. We have shown further that a much higher resorption rate, on days 11 to 13 of embryonic development, is observed among transgenic embryos developing from eggs microinjected at the one-cell stage with the pHMG-Kb construct than in control embryos. This lethality is not due to immune phenomena, since it is observed in histocompatible combinations between mother and fetus. These results are discussed in the context of what is currently known about the regulation of MHC expression at the fetal-maternal interface and in various transgenic mouse models.

  4. Esrrb-Cre Excises loxP-Flanked Alleles in Early Four-Cell Embryos

    PubMed Central

    Kim, Suyeon; Shaffer, Benjamin; Simerly, Calvin R.; Richard Chaillet, J.; Barak, Yaacov

    2015-01-01

    Among transgenic mice with ubiquitous Cre recombinase activity, all strains to date excise loxP-flanked (floxed) alleles, either at or before the zygote stage or at nondescript stages of development. This manuscript describes a new mouse strain, in which Cre recombinase, integrated into the Esrrb locus, efficiently excises floxed alleles in pre-implantation embryos at the onset of the four-cell stage. By enabling inactivation of genes only after the embryo has undergone two cleavages, this strain should facilitate in vivo studies of genes with essential gametic or zygotic functions. In addition, this study describes a new, highly pluripotent hybrid C57BL/6J × 129S1/SvImJ mouse embryonic stem cell line, HYB12, in which this knock-in and additional targeted alleles have been generated. PMID:26663459

  5. The study of early human embryos using interactive 3-dimensional computer reconstructions.

    PubMed

    Scarborough, J; Aiton, J F; McLachlan, J C; Smart, S D; Whiten, S C

    1997-07-01

    Tracings of serial histological sections from 4 human embryos at different Carnegie stages were used to create 3-dimensional (3D) computer models of the developing heart. The models were constructed using commercially available software developed for graphic design and the production of computer generated virtual reality environments. They are available as interactive objects which can be downloaded via the World Wide Web. This simple method of 3D reconstruction offers significant advantages for understanding important events in morphological sciences.

  6. Ultrastructural dynamics of human reproduction, from ovulation to fertilization and early embryo development.

    PubMed

    Familiari, Giuseppe; Heyn, Rosemarie; Relucenti, Michela; Nottola, Stefania A; Sathananthan, A Henry

    2006-01-01

    This study describes the updated, fine structure of human gametes, the human fertilization process, and human embryos, mainly derived from assisted reproductive technology (ART). As clearly shown, the ultrastructure of human reproduction is a peculiar multistep process, which differs in part from that of other mammalian models, having some unique features. Particular attention has been devoted to the (1) sperm ultrastructure, likely "Tygerberg (Kruger) strict morphology criteria"; (2) mature oocyte, in which the MII spindle is barrel shaped, anastral, and lacking centrioles; (3) three-dimensional microarchitecture of the zona pellucida with its unique supramolecular filamentous organization; (4) sperm-egg interactions with the peculiarity of the sperm centrosome that activates the egg and organizes the sperm aster and mitotic spindles of the embryo; and (5) presence of viable cumulus cells whose metabolic activity is closely related to egg and embryo behavior in in vitro as well as in vivo conditions, in a sort of extraovarian "microfollicular unit." Even if the ultrastructural morphodynamic features of human fertilization are well understood, our knowledge about in vivo fertilization is still very limited and the complex sequence of in vivo biological steps involved in human reproduction is only partially reproduced in current ART procedures.

  7. Early somatic embryo induction events in alfalfa callus cultures. [Medicago sativa

    SciT

    El-Bakry, A.A.; Hildebrand, D.F.

    1987-04-01

    High and low regenerating alfalfa Medicago sativa L. cv Regen S full sibs were isolated from a callus culture screen on modified Blaydes medium. The average number of embryos per ovary were thirty and zero for the high and low genotypes respectively after six weeks in culture. Proembryonic cell masses (4-8 celled) were observed after 4-5 days in culture and maximum meristematic activity was at 6-7 days in culture, for the high regenerating genotypes. Well formed globular embryos, both epidermal and subepidermal in origin, were observed after 2 weeks is culture. Samples in culture for 3, 6 and 14 daysmore » from the high and low regenerating genotypes were radiolabeled in vivo with /sup 35/S-methionine and run both on one and two dimension gels. The results will be discussed in relation to differences in proteins between the high and low regenerating genotypes at the stage of maximum meristematic activity (day 6) and differences occurring relative to the appearance of globular stage embryos (day 14) will be presented.« less

  8. Cortical geometry may influence placement of interface between Par protein domains in early Caenorhabditis elegans embryos.

    PubMed

    Dawes, Adriana T; Iron, David

    2013-09-21

    During polarization, proteins and other polarity determinants segregate to the opposite ends of the cell (the poles) creating biochemically and dynamically distinct regions. Embryos of the nematode worm Caenorhabditis elegans (C. elegans) polarize shortly after fertilization, creating distinct regions of Par protein family members. These regions are maintained through to first cleavage when the embryo divides along the plane specified by the interface between regions, creating daughter cells with different protein content. In wild type single cell embryos the interface between these Par protein regions is reliably positioned at approximately 60% egg length, however, it is not known what mechanisms are responsible for specifying the position of the interface. In this investigation, we use two mathematical models to investigate the movement and positioning of the interface: a biologically based reaction-diffusion model of Par protein dynamics, and the analytically tractable perturbed Allen-Cahn equation. When we numerically simulate the models on a static 2D domain with constant thickness, both models exhibit a persistently moving interface that specifies the boundary between distinct regions. When we modify the simulation domain geometry, movement halts and the interface is stably positioned where the domain thickness increases. Using asymptotic analysis with the perturbed Allen-Cahn equation, we show that interface movement depends explicitly on domain geometry. Using a combination of analytic and numeric techniques, we demonstrate that domain geometry, a historically overlooked aspect of cellular simulations, may play a significant role in spatial protein patterning during polarization. Copyright © 2013 Elsevier Ltd. All rights reserved.

  9. Establishment of left–right asymmetry in vertebrate development: the node in mouse embryos

    PubMed Central

    Komatsu, Yoshihiro

    2014-01-01

    Establishment of vertebrate left–right asymmetry is a critical process for normal embryonic development. After the discovery of genes expressed asymmetrically along the left–right axis in chick embryos in the mid 1990s, the molecular mechanisms responsible for left–right patterning in vertebrate embryos have been studied extensively. In this review article, we discuss the mechanisms by which the initial symmetry along the left–right axis is broken in the mouse embryo. We focus on the role of primary cilia and molecular mechanisms of ciliogenesis at the node when symmetry is broken and left–right asymmetry is established. The node is considered a signaling center for early mouse embryonic development, and the results we review here have led to a better understanding of how the node functions and establishes left–right asymmetry. PMID:23771646

  10. Establishment of left-right asymmetry in vertebrate development: the node in mouse embryos.

    PubMed

    Komatsu, Yoshihiro; Mishina, Yuji

    2013-12-01

    Establishment of vertebrate left-right asymmetry is a critical process for normal embryonic development. After the discovery of genes expressed asymmetrically along the left-right axis in chick embryos in the mid 1990s, the molecular mechanisms responsible for left-right patterning in vertebrate embryos have been studied extensively. In this review article, we discuss the mechanisms by which the initial symmetry along the left-right axis is broken in the mouse embryo. We focus on the role of primary cilia and molecular mechanisms of ciliogenesis at the node when symmetry is broken and left-right asymmetry is established. The node is considered a signaling center for early mouse embryonic development, and the results we review here have led to a better understanding of how the node functions and establishes left-right asymmetry.

  11. Early miscarriage rate in lean polycystic ovary syndrome women after euploid embryo transfer - a matched-pair study.

    PubMed

    Luo, Lu; Gu, Fang; Jie, Huying; Ding, Chenhui; Zhao, Qiang; Wang, Qiong; Zhou, Canquan

    2017-11-01

    The early miscarriage rate is reported to be higher in patients with polycystic ovary syndrome (PCOS) compared with non-PCOS patients. However, whether PCOS is an independent risk factor for early miscarriage is still controversial; to what extent embryonic aneuploidy accounts for miscarriages of PCOS is still unknown. In this 1:3 matched-pair study, 67 lean PCOS patients and 201 controls matched for age, body mass index (BMI) and embryo scores undergoing a single euploid blastocyst transfer in vitrified-warmed cycles were analysed. Clinical pregnancy, early miscarriage and live birth rates were compared. Logistic regression analysis was performed to further evaluate the factors associated with early miscarriage and live birth. Clinical pregnancy rates were 50.7% in PCOS and 55.2% in control groups. Early miscarriage rate was significantly (P = 0.029) increased in the PCOS group compared with controls; non-PCOS patients had a significantly higher live birth rate than PCOS patients, P < 0.001. Further regression analyses showed that PCOS was significantly associated with a higher risk of early miscarriage and decreased chance of live birth. In conclusion, PCOS in women undergoing pre-implantation genetic diagnosis may, independently from BMI and karyotype, increase the risk of miscarriage. Copyright © 2017 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  12. Gene expression analysis in zebrafish embryos: a potential approach to predict effect concentrations in the fish early life stage test.

    PubMed

    Weil, Mirco; Scholz, Stefan; Zimmer, Michaela; Sacher, Frank; Duis, Karen

    2009-09-01

    Based on the hypothesis that analysis of gene expression could be used to predict chronic fish toxicity, the zebrafish (Danio rerio) embryo test (DarT), developed as a replacement method for the acute fish test, was expanded to a gene expression D. rerio embryo test (Gene-DarT). The effects of 14 substances on lethal and sublethal endpoints of the DarT and on expression of potential marker genes were investigated: the aryl hydrocarbon receptor 2, cytochrome P450 1A (cypla), heat shock protein 70, fizzy-related protein 1, the transcription factors v-maf musculoaponeurotic fibrosarcoma oncogene family protein g (avian) 1 and NF-E2-p45-related factor, and heme oxygenase 1 (hmox1). After exposure of zebrafish embryos for 48 h, differential gene expression was evaluated using reverse transcriptase-polymerase chain reaction, gel electrophoresis, and densitometric analysis of the gels. All tested compounds significantly affected the expression of at least one potential marker gene, with cyp1a and hmox1 being most sensitive. Lowest-observed-effect concentrations (LOECs) for gene expression were below concentrations resulting in 10% lethal effects in the DarT. For 10 (3,4- and 3,5-dichloroaniline, 1,4-dichlorobenzene, 2,4-dinitrophenol, atrazine, parathion-ethyl, chlorotoluron, genistein, 4-nitroquinoline-1-oxide, and cadmium) out of the 14 tested substances, LOEC values derived with the Gene-DarT differ by a factor of less than 10 from LOEC values of fish early life stage tests with zebrafish. For pentachloroaniline and pentachlorobenzene, the Gene-DarT showed a 23- and 153-fold higher sensitivity, respectively, while for lindane, it showed a 13-fold lower sensitivity. For ivermectin, the Gene-DarT was by a factor of more than 1,000 less sensitive than the acute fish test. The results of the present study indicate that gene expression analysis in zebrafish embryos could principally be used to predict effect concentrations in the fish early life stage test.

  13. Integrating toxicity risk in bird eggs and chicks: using chick down feathers to estimate mercury concentrations in eggs.

    PubMed

    Ackerman, Joshua T; Eagles-Smith, Collin A

    2009-03-15

    The concentration of mercury (Hg) in eggs that causes reduced hatching success is regarded as a critical end point for Hg toxicity in birds. However, incorporating effects of in ovo mercury exposure on chick health and survival could improve risk assessment. We developed equations to predict Hg in eggs using Hg in chick down feathers, and vice versa, by assessing the relationship between Hg in feathers (0.5-32.4 microg g(-1) fw) and eggs (0.04-2.79 microg g(-1) fww) for three waterbird species in San Francisco Bay, CA. Feather Hg sampled from embryos of pipping eggs was highly correlated with fresh whole-egg Hg (n=94, r2 = 0.96). Additionally, using an egg microsampling technique, albumen Hg was correlated with feather Hg sampled from chicks in the same nest (n=28, r2 = 0.79). Down feather Hg in recaptured chicks (< or =10 days old) was correlated with down feather Hg at hatching (< or =3 days old; n=88, r2 = 0.74). Our results demonstrate the utility of using down feathers of chicks < or =10 days of age to nonlethally predict Hg in eggs and thus provide the ability to develop exposure thresholds for eggs that incorporate in ovo Hg's effects on both egg hatchability and subsequent chick mortality.

  14. Integrating toxicity risk in bird eggs and chicks: Using chick down feathers to estimate mercury concentrations in eggs

    Ackerman, Joshua T.; Eagles-Smith, Collin A.

    2009-01-01

    The concentration of mercury (Hg) in eggs that causes reduced hatching success is regarded as a critical end point for Hg toxicity in birds. However, incorporating effects of in ovo mercury exposure on chick health and survival could improve risk assessment. We developed equations to predict Hg in eggs using Hg in chick down feathers, and vice versa, by assessing the relationship between Hg in feathers (0.5−32.4 μg g−1 fw) and eggs (0.04−2.79 μg g−1 fww) for three waterbird species in San Francisco Bay, CA. Feather Hg sampled from embryos of pipping eggs was highly correlated with fresh whole-egg Hg (n = 94, r2 = 0.96). Additionally, using an egg microsampling technique, albumen Hg was correlated with feather Hg sampled from chicks in the same nest (n = 28, r2 = 0.79). Down feather Hg in recaptured chicks (≤10 days old) was correlated with down feather Hg at hatching (≤3 days old; n = 88, r2 = 0.74). Our results demonstrate the utility of using down feathers of chicks ≤10 days of age to nonlethally predict Hg in eggs and thus provide the ability to develop exposure thresholds for eggs that incorporate in ovo Hg’s effects on both egg hatchability and subsequent chick mortality.

  15. The First Human Cloned Embryo.

    ERIC Educational Resources Information Center

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

    2002-01-01

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

  16. Conference lecture: influence of stress on estrus, gametes and early embryo development in the sow.

    PubMed

    Einarsson, S; Brandt, Y; Rodriguez-Martinez, H; Madej, A

    2008-11-01

    Systems with loose-housed sows have become common. Regrouping, which is commonly done after weaning and may coincide with many important reproductive events, causes stressful situations with elevated blood cortisol concentrations. Depending on group size, approximately 2-7 d are required for a new group of sows to become relatively stable. In a series of studies, the social stress after regrouping was simulated with repeated adrenocorticotrophic hormone (ACTH) treatments for approximately 48h. Sows were allocated into control and experimental groups, fitted with jugular catheters, and blood samples were collected every 2 or 4h. Follicular development and ovulation were monitored by transrectal ultrasonography every 4h. Simulated stress during pro-estrus prolonged estrus and disturbed the follicular growth and ovulation. Giving ACTH during estrus elevated concentrations of cortisol and progesterone, and changed the intraluminal environment, including exaggerated amounts of mucus in the UTJ and isthmus. Although ACTH had no effect on the time of ovulation (relative to onset of standing estrus), or on embryo development, fewer oocytes/embryos were retrieved from the ACTH group than from the control group (51% vs. 81%, P<0.05), and there was a tendency towards faster embryo transportation to the uterus. Short-term fasting after ovulation had an unfavourable effect on sperm numbers in UTJ/isthmus, cleavage rate of fertilized ova, as well as ova transport through the isthmic part of the oviduct. Treatment with ACTH after ovulation reduced numbers of spermatozoa at the zona pellucida and retarded cleavage rate of fertilized ova. Therefore, the timing of stress seemed to be an important factor regarding effects on reproductive events.

  17. Properties of Barley Seed Chitinases and Release of Embryo-Associated Isoforms during Early Stages of Imbibition 1

    PubMed Central

    Swegle, Mark; Kramer, Karl J.; Muthukrishnan, Subbaratnam

    1992-01-01

    Barley (Hordeum vulgare L.) seeds contain at least five proteins with chitinase (CH) activity. Two of these (CH1 and CH2) are found primarily in the aleurone and endosperm tissues, and the other three (CH3, CH4, and CH5) are enriched in the embryo. From the bran fraction, three of these CHs (CH1, CH2, and CH3) were purified to apparent homogeneity. These three CHs have apparent molecular masses of 27, 34, and 35 kilodaltons and isoelectric points of 9.3, 9.2, and 8.7, respectively. CH2 and CH3 have amino terminal sequences resembling a portion of the chitin-binding domain of lectins and other plant defense proteins. CH1 lacks this domain. All three CHs exhibit antifungal activity and inhibit the mycelial growth of some species of trichoderma and Fusarium in vitro. During the early period of imbibition by seeds, two of the embryo-associated CHs are selectively released into the surrounding aqueous medium. ImagesFigure 1Figure 2Figure 3Figure 4 PMID:16668964

  18. The role of eggshell and underlying vitelline membrane for normal pattern formation in the early C. elegans embryo.

    PubMed

    Schierenberg, Einhard; Junkersdorf, Bernd

    1992-12-01

    The embryo of the nematode Caenorhabditis elegans is surrounded by an inconspicuous inner vitelline membrane and a prominent outer chitinous eggshell proper. We demonstrate that the complete removal of the chitinous eggshell does not interfere with successful development to yield a normal worm. The same result can be obtained when the vitelline membrane is penetrated with laser microbeam irradiation of only the eggshell proper, gently enough to permit its resealing after a while. However, when large holes are made into the eggshell the concomitantly penetrated vitelline membrane does not reseal. While early development is quite normal under these conditions, gastrulation is defective in that gut precursor cells do not migrate in properly, eventually leading to embryonic arrest. This suggests a crucial role for pattern formation of the "micro-environment" around the embryo preserved by the intact vitelline membrane. Removing both eggshell and vitelline membrane results in a string-like arrangement of founder cells and subsequent grossly abnormal cell patterns. Our experiments support the idea that the prominent eggshell proper just functions as a mechanical protection while the thin vitelline membrane directly or indirectly serves as a necessary control element affecting the positions of cells which to begin with are determined by the orientation of the cleavage spindle.

  19. Teen Chick Lit

    ERIC Educational Resources Information Center

    Meloni, Christine

    2006-01-01

    For young teen girls, reading has become hot again. With their appealing covers, witty heroines and humorous plots, teen chick lit books are bringing girls out of the malls and into local libraries and bookstores in search of the next must-have title. These fun books are about boys, friendship, family, fitting in, and growing up. What makes the…

  20. Effect of a 1800 MHz electromagnetic field emitted during embryogenesis on chick development and hatchability.

    PubMed

    Pawlak, K; Nieckarz, Z; Sechman, A; Wojtysiak, D; Bojarski, B; Tombarkiewicz, B

    2018-06-01

    The level of artificial electromagnetic field (EMF) has steadily increased with the development of human civilization. The developing chicken embryo has been considered a good model to study the effects of EMF on living organisms. The aim of the study was to determine the effect of a 1800 MHz electromagnetic field during embryogenesis on the frequency of chick embryo malformations, morphometric parameters of the heart and liver and concentration of corticosterone in blood plasma, lipid and glycogen content in the liver of newly hatched chicks. A 1800 MHz EMF was found to shorten the duration of embryogenesis (earlier pipping and hatching of chicks) while having no effect on the quantity and quality of chicks and on increasing the incidence of embryo malformations. Exposure of chick embryos to EMF caused decreases in relative heart weight and right ventricle wall thickness. The pipping and hatching of chicks can be accelerated by stressful impact of EMF, which is confirmed by a significant increase in plasma corticosterone concentrations and decrease in fat and glycogen in the liver of chicks exposed during embryogenesis on the electromagnetic field with a frequency of 1800 MHz. © 2018 Blackwell Verlag GmbH.

  1. EVALUATING THE EFFECTS OF FLY ASH EXPOSURE ON FISH EARLY LIFE STAGES: FATHEAD MINNOW EMBRYO-LARVAL TESTS

    SciT

    Greeley Jr, Mark Stephen; Elmore, Logan R; McCracken, Kitty

    2012-05-01

    On December 22, 2008, a dike containing fly ash and bottom ash in an 84-acre complex of the Tennessee Valley Authority's (TVA) Kingston Steam Plant in East Tennessee failed and released a large quantity of ash into the adjacent Emory River. Ash deposits extended as far as 4 miles upstream (Emory River mile 6) of the Plant, and some ash was carried as far downstream as Tennessee River mile 564 ({approx}4 miles downstream of the Tennessee River confluence with the Clinch River). A byproduct of coal burning power plants, fly ash contains a variety of metals and other elements which,more » at sufficient concentrations and in specific forms, can be toxic to biological systems. The effects of fly ash contamination on exposed fish populations depend on the magnitude and duration of exposure, with the most significant risk considered to be the effects of specific ash constituents, especially selenium, on fish early life stages. Uptake by adult female fish of fly ash constituents through the food chain and subsequent maternal transfer of contaminants to the developing eggs is thought to be the primary route of selenium exposure to larval fish (Woock and others 1987, Coyle and others 1993, Lemly 1999, Moscatello and others 2006), but direct contact of the fertilized eggs and developing embryos to ash constituents in river water and sediments is also a potential risk factor (Woock and others 1987, Coyle and others 1993, Jezierska and others 2009). To address the risk of fly ash from the Kingston spill to the reproductive health of downstream fish populations, ORNL has undertaken a series of studies in collaboration with TVA including: (1) a field study of the bioaccumulation of fly ash constituents in fish ovaries and the reproductive condition of sentinel fish species in reaches of the Emory and Clinch Rivers affected by the fly ash spill; (2) laboratory tests of the potential toxicity of fly ash from the spill area on fish embryonic and larval development (reported in the

  2. Adaptive capacity of the habitat modifying sea urchin Centrostephanus rodgersii to ocean warming and ocean acidification: performance of early embryos.

    PubMed

    Foo, Shawna A; Dworjanyn, Symon A; Poore, Alistair G B; Byrne, Maria

    2012-01-01

    Predicting effects of rapid climate change on populations depends on measuring the effects of climate stressors on performance, and potential for adaptation. Adaptation to stressful climatic conditions requires heritable genetic variance for stress tolerance present in populations. We quantified genetic variation in tolerance of early development of the ecologically important sea urchin Centrostephanus rodgersii to near-future (2100) ocean conditions projected for the southeast Australian global change hot spot. Multiple dam-sire crosses were used to quantify the interactive effects of warming (+2-4 °C) and acidification (-0.3-0.5 pH units) across twenty-seven family lines. Acidification, but not temperature, decreased the percentage of cleavage stage embryos. In contrast, temperature, but not acidification decreased the percentage of gastrulation. Cleavage success in response to both stressors was strongly affected by sire identity. Sire and dam identity significantly affected gastrulation and both interacted with temperature to determine developmental success. Positive genetic correlations for gastrulation indicated that genotypes that did well at lower pH also did well in higher temperatures. Significant genotype (sire) by environment interactions for both stressors at gastrulation indicated the presence of heritable variation in thermal tolerance and the ability of embryos to respond to changing environments. The significant influence of dam may be due to maternal provisioning (maternal genotype or environment) and/or offspring genotype. It appears that early development in this ecologically important sea urchin is not constrained in adapting to the multiple stressors of ocean warming and acidification. The presence of tolerant genotypes indicates the potential to adapt to concurrent warming and acidification, contributing to the resilience of C. rodgersii in a changing ocean.

  3. Adaptive Capacity of the Habitat Modifying Sea Urchin Centrostephanus rodgersii to Ocean Warming and Ocean Acidification: Performance of Early Embryos

    PubMed Central

    Foo, Shawna A.; Dworjanyn, Symon A.; Poore, Alistair G. B.; Byrne, Maria

    2012-01-01

    Background Predicting effects of rapid climate change on populations depends on measuring the effects of climate stressors on performance, and potential for adaptation. Adaptation to stressful climatic conditions requires heritable genetic variance for stress tolerance present in populations. Methodology/Principal Findings We quantified genetic variation in tolerance of early development of the ecologically important sea urchin Centrostephanus rodgersii to near-future (2100) ocean conditions projected for the southeast Australian global change hot spot. Multiple dam-sire crosses were used to quantify the interactive effects of warming (+2–4°C) and acidification (−0.3−0.5 pH units) across twenty-seven family lines. Acidification, but not temperature, decreased the percentage of cleavage stage embryos. In contrast, temperature, but not acidification decreased the percentage of gastrulation. Cleavage success in response to both stressors was strongly affected by sire identity. Sire and dam identity significantly affected gastrulation and both interacted with temperature to determine developmental success. Positive genetic correlations for gastrulation indicated that genotypes that did well at lower pH also did well in higher temperatures. Conclusions/Significance Significant genotype (sire) by environment interactions for both stressors at gastrulation indicated the presence of heritable variation in thermal tolerance and the ability of embryos to respond to changing environments. The significant influence of dam may be due to maternal provisioning (maternal genotype or environment) and/or offspring genotype. It appears that early development in this ecologically important sea urchin is not constrained in adapting to the multiple stressors of ocean warming and acidification. The presence of tolerant genotypes indicates the potential to adapt to concurrent warming and acidification, contributing to the resilience of C. rodgersii in a changing ocean. PMID

  4. 5Alpha-Reduced Steroids Are Major Metabolites in the Early Equine Embryo Proper and Its Membranes.

    PubMed

    Raeside, James I; Christie, Heather L; Betteridge, Keith J

    2015-09-01

    Steroid production and metabolism by early conceptuses are very important for the establishment and maintenance of pregnancy in horses. Our earlier work suggested the possible formation of 5alpha-reduced steroids in equine conceptuses. We have now demonstrated the formation of 5alpha-reduced metabolites of androstenedione, testosterone, and progesterone by the embryo and its membranes. A total of 44 conceptuses were collected from 26 mares between 20 and 31 days of pregnancy. Tissues from the embryo proper and from the separated components of the conceptus (bilaminar and trilaminar trophoblast, allantois) were incubated with tritium-labeled substrates. 5Alpha-reduced metabolites (5alpha-dihydro- and 3beta,5alpha-tetrahydro- steroids) as radiolabeled products were identified from a series of chromatographic steps using four solvent systems for high-performance liquid chromatography. Use of a 5alpha-reductase inhibitor confirmed the metabolites were indeed 5alpha-reduced steroids. For the embryo, the only products from androstenedione were 5alpha-dihydroandrostenedione and 3beta,5alpha-tetrahydroandrostenedione, with no evidence of more polar metabolites; there was some 3beta,5alpha-tetrahydrotestosterone but no 5alpha-dihydrotestosterone from testosterone, and formation of androstenedione was followed by the production of 5alpha-dihydroandrostenedione and 3beta,5alpha-tetrahydroandrostenedione. The major 5alpha-reduced product from progesterone was 3beta,5alpha-tetrahydroprogesterone, with lesser amounts of 5alpha-dihydroprogesterone. For the membranes, reductions to tetrahydro, 5alpha-reduced steroids were prominent in most instances, but also present were considerable amounts of products more polar than the substrates. The well-recognized activity of some 5alpha-reduced steroids--for example, 5alpha-dihydrotestosterone in male sexual differentiation--provokes interest in their even earlier appearance, as seen in this study, and suggests a possible role for them in

  5. Sensory regulation of spontaneous limb movements in the midstage embryonic chick.

    PubMed

    Sharp, Andrew A

    2015-05-01

    It is becoming increasingly apparent that somatosensation plays an important role in regulating prenatal movement and developmental plasticity. Numerous studies performed on embryonic chicks and perinatal rats are beginning to implicate proprioception to be particularly important in modulating motility very soon after afferent connections are made in the spinal cord. In this report, we demonstrate new approaches in the chick embryo to explore the role of sensation in modulating embryonic movement. Force recordings from the legs of chick embryos on E9 and E11, during spontaneous motility, demonstrate changes in sensory regulation consistent with the concept that sensory regulation is functioning one day after sensory synapse formation and that the complexity of this regulation increases by E11. Additionally, we present new video data showing activation of embryonic motility on E5 and E9 in embryos expressing channelrhodopsin in the spinal cord as a novel way to approach the issues of sensorimotor development. © 2015 Wiley Periodicals, Inc.

  6. Yolk hormones influence in ovo chemosensory learning, growth, and feeding behavior in domestic chicks.

    PubMed

    Bertin, Aline; Meurisse, Maryse; Arnould, Cécile; Leterrier, Christine; Constantin, Paul; Cornilleau, Fabien; Vaudin, Pascal; Burlot, Thierry; Delaveau, Joel; Rat, Christophe; Calandreau, Ludovic

    2016-03-01

    In this study, we assessed whether prenatal exposure to elevated yolk steroid hormones can influence in ovo chemosensory learning and the behavior of domestic chicks. We simulated a maternal environmental challenge by experimentally enhancing yolk progesterone, testosterone, and estradiol concentrations in hen eggs prior to incubation. The embryos from these hormones-treated eggs (HO) as well as sham embryos (O) that had received the vehicle-only were exposed to the odor of fish oil (menhaden) between embryonic Days 11 and 20. An additional group of control embryos (C) was not exposed to the odor. All chicks were tested following hatching for their feeding preferences between foods that were or were not odorized with the menhaden odor. In the 3-min choice tests, the behavior of O chicks differed significantly according to the type of food whereas C and HO chicks showed no preference between odorized and non-odorized food. Our result suggests weaker response in HO chicks. In addition, HO chicks showed impaired growth and reduced intake of an unfamiliar food on the 24-h time scale compared to controls. Our data suggest that embryonic exposure to increased yolk hormone levels can alter growth, chemosensory learning, and the development of feeding behaviors. © 2015 Wiley Periodicals, Inc.

  7. Time-lapse culture with morphokinetic embryo selection improves pregnancy and live birth chances and reduces early pregnancy loss: a meta-analysis.

    PubMed

    Pribenszky, Csaba; Nilselid, Anna-Maria; Montag, Markus

    2017-11-01

    Embryo evaluation and selection is fundamental in clinical IVF. Time-lapse follow-up of embryo development comprises undisturbed culture and the application of the visual information to support embryo evaluation. A meta-analysis of randomized controlled trials was carried out to study whether time-lapse monitoring with the prospective use of a morphokinetic algorithm for selection of embryos improves overall clinical outcome (pregnancy, early pregnancy loss, stillbirth and live birth rate) compared with embryo selection based on single time-point morphology in IVF cycles. The meta-analysis of five randomized controlled trials (n = 1637) showed that the application of time-lapse monitoring was associated with a significantly higher ongoing clinical pregnancy rate (51.0% versus 39.9%), with a pooled odds ratio of 1.542 (P < 0.001), significantly lower early pregnancy loss (15.3% versus 21.3%; OR: 0.662; P = 0.019) and a significantly increased live birth rate (44.2% versus 31.3%; OR 1.668; P = 0.009). Difference in stillbirth was not significant between groups (4.7% versus 2.4%). Quality of the evidence was moderate to low owing to inconsistencies across the studies. Selective application and variability were also limitations. Although time-lapse is shown to significantly improve overall clinical outcome, further high-quality evidence is needed before universal conclusions can be drawn. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  8. Green Light-emitting Diodes Light Stimuli during Incubation Enhances Posthatch Growth without Disrupting Normal Eye Development of Broiler Embryos and Hatchlings.

    PubMed

    Zhang, L; Zhu, X D; Wang, X F; Li, J L; Gao, F; Zhou, G H

    2016-11-01

    Monochromatic green light-emitting diodes (LED) light stimuli influences the posthatch growth performance of chicks. This study was undertaken with the following objectives: i) to examine whether the green LED light stimuli induces an overheating effect by determining weight loss rate of fertile eggs during incubation period; ii) to look for the development of eyes and other primary organs at different ages of embryos and newly hatched chicks. Arbor Acres fertile broiler eggs (n = 480) were randomly assigned to 3 incubation groups and exposed to continuous white light, green light, or a dark environment (control) from the first day to 19 d of incubation. The light sourced from LED lamps with the intensity of 30 lx at eggshell level. The results showed that either green or white light stimuli during incubation did not significantly affect the weight loss rate of fertile eggs, hatching time, hatchability, chick embryo, or body weight (BW), the weight percentage of heart, liver, and eyes, as well as obvious systematic abnormalities in eye weight, side-to-side, back-to-front, or corneal diameter from 15 d of embryogenesis to 6 d of posthatch (p>0.05). Compared with the dark condition, green light stimuli during incubation tended to increase feed intake (p = 0.080), improved the BW gain of chicks during 0 to 6 day posthatch (p<0.05), and increased the percentage of pectoral muscle to the BW on 3- and 6-day-old chicks. In addition, embryos or chicks in green light had lower weight percentage of yolk retention on 19 d of embryogenesis and 1 d of posthatch in comparison to those in dark or white group (p<0.05). These results suggest that providing 30 lx green LED light stimuli during incubation has no detrimental effect on the development of eyes, heart and liver of embryos and hatchlings, but does have potential benefits in terms of enhancement of the chick growth during the early posthatch stages. In addition, the fertile broiler eggs stimulated with 30 lx green LED

  9. Insulin-like growth factor (IGF) signalling is required for early dorso-anterior development of the zebrafish embryo.

    PubMed

    Eivers, Edward; McCarthy, Karena; Glynn, Catherine; Nolan, Catherine M; Byrnes, Lucy

    2004-12-01

    The insulin-like growth factor (IGF) signalling pathway has been highly conserved in animal evolution and, in mammals and Xenopus, plays a key role in embryonic growth and development, with the IGF-1 receptor (IGF-1R) being a crucial regulator of the signalling cascade. Here we report the first functional role for the IGF pathway in zebrafish. Expression of mRNA coding for a dominant negative IGF-1R resulted in embryos that were small in size compared to controls and had disrupted head and CNS development. At its most extreme, this phenotype was characterized by a complete loss of head and eye structures, an absence of notochord and the presence of abnormal somites. In contrast, up-regulation of IGF signalling following injection of IGF-1 mRNA, resulted in a greatly expanded development of anterior structures at the expense of trunk and tail. IGF-1R knockdown caused a significant decrease in the expression of Otx2, Rx3, FGF8, Pax6.2 and Ntl, while excess IGF signalling expanded Otx2 expression in presumptive forebrain tissue and widened the Ntl expression domain in the developing notochord. The observation that IGF-1R knockdown reduced expression of two key organizer genes (chordin and goosecoid) suggests that IGF signalling plays a role in regulating zebrafish organizer activity. This is supported by the expression of IGF-1, IGF-2 and IGF-1R in shield-stage zebrafish embryos and the demonstration that IGF signalling influences expression of BMP2b, a gene that plays an important role in zebrafish pattern formation. Our data is consistent with a common pathway for integration of IGF, FGF8 and anti-BMPs in early vertebrate development.

  10. Asymmetric distribution of hypoxia-inducible factor α regulates dorsoventral axis establishment in the early sea urchin embryo.

    PubMed

    Chang, Wei-Lun; Chang, Yi-Cheng; Lin, Kuan-Ting; Li, Han-Ru; Pai, Chih-Yu; Chen, Jen-Hao; Su, Yi-Hsien

    2017-08-15

    Hypoxia signaling is an ancient pathway by which animals can respond to low oxygen. Malfunction of this pathway disturbs hypoxic acclimation and can result in various diseases, including cancers. The role of hypoxia signaling in early embryogenesis remains unclear. Here, we show that in the blastula of the sea urchin Strongylocentrotus purpuratus , hypoxia-inducible factor α (HIFα), the downstream transcription factor of the hypoxia pathway, is localized and transcriptionally active on the future dorsal side. This asymmetric distribution is attributable to its oxygen-sensing ability. Manipulations of the HIFα level entrained the dorsoventral axis, as the side with the higher level of HIFα tends to develop into the dorsal side. Gene expression analyses revealed that HIFα restricts the expression of nodal to the ventral side and activates several genes encoding transcription factors on the dorsal side. We also observed that intrinsic hypoxic signals in the early embryos formed a gradient, which was disrupted under hypoxic conditions. Our results reveal an unprecedented role of the hypoxia pathway in animal development. © 2017. Published by The Company of Biologists Ltd.

  11. Gastrointestinal obstruction in penguin chicks.

    PubMed

    Perpiñán, David; Curro, Thomas G

    2009-12-01

    A 7-day-old gentoo penguin (Pygoscelis papua) was found dead and postmortem examination revealed impaction of the ventriculus with feathers. A review of mortality in gentoo penguin chicks from 1997 to 2007 at that institution revealed another case of feather impaction of the ventriculus in a 4-week-old chick, a sibling of the previous chick. A third case of gastrointestinal impaction occurred in a 24-day-old king penguin (Aptenodytes patagonicus) with omphallitis and enteritis. In this chick, a fibrin mat produced a complete obstruction of the intestine at the level of Meckel's diverticulum.

  12. Embryo-endometrial interactions during early development after embryonic diapause in the marsupial tammar wallaby.

    PubMed

    Renfree, Marilyn B; Shaw, Geoff

    2014-01-01

    The marsupial tammar wallaby has the longest period of embryonic diapause of any mammal. Reproduction in the tammar is seasonal, regulated by photoperiod and also lactation. Reactivation is triggered by falling daylength after the austral summer solstice in December. Young are born late January and commence a 9-10-month lactation. Females mate immediately after birth. The resulting conceptus develops over 6- 7 days to form a unilaminar blastocyst of 80-100 cells and enters lactationally, and later seasonally, controlled diapause. The proximate endocrine signal for reactivation is an increase in progesterone which alters uterine secretions. Since the diapausing blastocyst is surrounded by the zona and 2 other acellular coats, the mucoid layer and shell coat, the uterine signals that maintain or terminate diapause must involve soluble factors in the secretions rather than any direct cellular interaction between uterus and embryo. Our studies suggest involvement of a number of cytokines in the regulation of diapause in tammars. The endometrium secretes platelet activating factor (PAF) and leukaemia inhibitory factor, which increase after reactivation. Receptors for PAF are low on the blastocyst during diapause but are upregulated at reactivation. Conversely, there is endometrial expression of the muscle segment homeobox gene MSX2 throughout diapause, but it is rapidly downregulated at reactivation. These patterns are consistent with those observed in diapausing mice and mink after reactivation, despite the very different patterns of endocrine control of diapause in these 3 divergent species. These common patterns suggest a similar underlying mechanism for diapause, perhaps common to all mammals, but which is activated in only a few.

  13. Effects of lipopolysaccharide (LPS) induced inflammatory response on early embryo survival in ewes

    Early pregnant ewes were used to determine the effects of endogenous (through LPS activation) and exogenous TNF-alpha tumor necrosis factor-alpha (TNF-alpha) on embryonic loss. Thirty-eight Dorset x Texel ewes were synchronized for estrus and bred to fertile rams (d0). On d5/6, ewes were assigned t...

  14. Impact of bisphenol A (BPA) on early embryo development in the marine mussel Mytilus galloprovincialis: Effects on gene transcription.

    PubMed

    Balbi, Teresa; Franzellitti, Silvia; Fabbri, Rita; Montagna, Michele; Fabbri, Elena; Canesi, Laura

    2016-11-01

    Bisphenol A (BPA), a monomer used in plastic manufacturing, is weakly estrogenic and a potential endocrine disruptor in mammals. Although it degrades quickly, it is pseudo-persistent in the environment because of continual inputs, with reported concentrations in aquatic environments between 0.0005 and 12 μg/L. BPA represents a potential concern for aquatic ecosystems, as shown by its reproductive and developmental effects in aquatic vertebrates. In invertebrates, endocrine-related effects of BPA were observed in different species and experimental conditions, with often conflicting results, indicating that the sensitivity to this compound can vary considerably among related taxa. In the marine mussel Mytilus galloprovincialis BPA was recently shown to affect early development at environmental concentrations. In this work, the possible effects of BPA on mussel embryos were investigated at the molecular level by evaluating transcription of 13 genes, selected on the basis of their biological functions in adult mussels. Gene expression was first evaluated in trocophorae and D-veligers (24 and 48 h post fertilization) grown in physiological conditions, in comparison with unfertilized eggs. Basal expressions showed a general up-regulation during development, with distinct transcript levels in trocophorae and D-veligers. Exposure of fertilized eggs to BPA (10 μg/L) induced a general upregulation at 24 h pf, followed by down regulation at 48 h pf. Mytilus Estrogen Receptors, serotonin receptor and genes involved in biomineralization (Carbonic Anydrase and Extrapallial Protein) were the most affected by BPA exposure. At 48 h pf, changes in gene expression were associated with irregularities in shell formation, as shown by scanning electron microscopy (SEM), indicating that the formation of the first shelled embryo, a key step in mussel development, represents a sensitive target for BPA. Similar results were obtained with the natural estrogen 17β-estradiol. The

  15. Insulin resistance does not affect early embryo development but lowers implantation rate in in vitro maturation-in vitro fertilization-embryo transfer cycle.

    PubMed

    Chang, Eun M; Han, Ji E; Seok, Hyun H; Lee, Dong R; Yoon, Tae K; Lee, Woo S

    2013-07-01

    Although extensive evidence indicates the hyperinsulinemia directly contributes to reproductive dysfunction in polycystic ovarian syndrome (PCOS), influence of insulin resistance (IR) on assisted reproductive technology outcomes is poorly understood. In this study we aimed to evaluate the effects of IR on in vitro maturation-in vitro fertilization-embryo transfer (IVM-IVF-ET) in patients with PCOS. Prospective observational study. Women with PCOS (n = 115) commencing IVM. IR (n = 51) and non-IR (n = 64) women with PCOS ready to commence an IVM cycle were recruited. IR was diagnosed using the glucose tolerance test (GTT) and homeostasis model assessment (HOMA) index. Patients with an abnormal GTT and/or HOMA index >2·4 were considered IR. Patients underwent 115 cycles of unstimulated hCG-primed IVM. Maturation, fertilization, cleavage rates, the number of good-quality embryo, and blastocyst formation rates were not significantly different between groups. However, implantation (11·6% vs 28·7%, P = 0·001, respectively), clinical pregnancy (23·5% vs 53·1%, P = 0·002, respectively), and ongoing pregnancy rates (21·6% vs 46·9%. P = 0·006, respectively) were significantly decreased in the IR group. The negative effect of IR on pregnancy outcomes remained after controlling for age, body mass index (BMI) and lipid profiles (OR 4·928, 95% CI 1·735-13·991, P = 0·003). Pregnancy rate after IVM is impaired in IR patients with PCOS. Oocyte development and embryo quality are not affected, suggesting that the effects of hyperinsulinemia on endometrial function and implantation process underlie the decreased pregnancy rate. © 2012 John Wiley & Sons Ltd.

  16. DNA repair efficiency in germ cells and early mouse embryos and consequences for radiation-induced transgenerational genomic damage

    SciT

    Marchetti, Francesco; Wyrobek, Andrew J.

    Exposure to ionizing radiation and other environmental agents can affect the genomic integrity of germ cells and induce adverse health effects in the progeny. Efficient DNA repair during gametogenesis and the early embryonic cycles after fertilization is critical for preventing transmission of DNA damage to the progeny and relies on maternal factors stored in the egg before fertilization. The ability of the maternal repair machinery to repair DNA damage in both parental genomes in the fertilizing egg is especially crucial for the fertilizing male genome that has not experienced a DNA repair-competent cellular environment for several weeks prior to fertilization.more » During the DNA repair-deficient period of spermatogenesis, DNA lesions may accumulate in sperm and be carried into the egg where, if not properly repaired, could result in the formation of heritable chromosomal aberrations or mutations and associated birth defects. Studies with female mice deficient in specific DNA repair genes have shown that: (i) cell cycle checkpoints are activated in the fertilized egg by DNA damage carried by the sperm; and (ii) the maternal genotype plays a major role in determining the efficiency of repairing genomic lesions in the fertilizing sperm and directly affect the risk for abnormal reproductive outcomes. There is also growing evidence that implicates DNA damage carried by the fertilizing gamete as a mediator of postfertilization processes that contribute to genomic instability in subsequent generations. Transgenerational genomic instability most likely involves epigenetic mechanisms or error-prone DNA repair processes in the early embryo. Maternal and embryonic DNA repair processes during the early phases of mammalian embryonic development can have far reaching consequences for the genomic integrity and health of subsequent generations.« less

  17. The cell cycle of early mammalian embryos: lessons from genetic mouse models.

    PubMed

    Artus, Jérôme; Babinet, Charles; Cohen-Tannoudji, Michel

    2006-03-01

    Genes coding for cell cycle components predicted to be essential for its regulation have been shown to be dispensable in mice, at the whole organism level. Such studies have highlighted the extraordinary plasticity of the embryonic cell cycle and suggest that many aspects of in vivo cell cycle regulation remain to be discovered. Here, we discuss the particularities of the mouse early embryonic cell cycle and review the mutations that result in cell cycle defects during mouse early embryogenesis, including deficiencies for genes of the cyclin family (cyclin A2 and B1), genes involved in cell cycle checkpoints (Mad2, Bub3, Chk1, Atr), genes involved in ubiquitin and ubiquitin-like pathways (Uba3, Ubc9, Cul1, Cul3, Apc2, Apc10, Csn2) as well as genes the function of which had not been previously ascribed to cell cycle regulation (Cdc2P1, E4F and Omcg1).

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

    NASA Astrophysics Data System (ADS)

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

    2017-03-01

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

  19. Parental contributions to early embryo development: influences of urinary phthalate and phthalate alternatives among couples undergoing IVF treatment.

    PubMed

    Wu, Haotian; Ashcraft, Lisa; Whitcomb, Brian W; Rahil, Tayyab; Tougias, Ellen; Sites, Cynthia K; Pilsner, J Richard

    2017-01-01

    Are preconception urinary concentrations of phthalates and phthalate alternatives associated with diminished early stage embryo quality in couples undergoing IVF? Male, but not female, urinary concentrations of select metabolites of phthalates and phthalate alternatives are associated with diminished blastocyst quality. Although phthalates are endocrine disrupting compounds associated with adverse reproductive health, they are in widespread use across the world. Male and female preconception exposures to select phthalates have been previously associated with adverse reproductive outcomes in both the general population and in those undergoing IVF. This prospective cohort included 50 subfertile couples undergoing IVF in western Massachusetts. This study includes the first 50 couples recruited from the Baystate Medical Center's Fertility Center in Springfield, MA, as part of the Sperm Environmental Epigenetics and Development Study (SEEDS). Relevant data from both partners, including embryo quality at the cleavage (Day 3) and blastocyst (Day 5) stages, were collected by clinic personnel during the normal course of an IVF cycle. A spot urine sample was collected from both male and female partners on the same day as semen sample procurement and oocyte retrieval. Concentrations of 17 urinary metabolite were quantified by liquid chromatography mass spectrometry and normalized via specific gravity. Generalized estimating equations were used to estimate odds ratios (OR) and 95% CI, with urinary phthalates and phthalate alternatives fitted as continuous variables and embryo quality as a binary variable. The 50 couples contributed 761 oocytes, of which 423 progressed to the cleavage stage, 261 were high-quality cleavage stage embryos, 137 were transferrable quality blastocysts and 47 were high-quality blastocysts. At the cleavage stage, male urinary monoethyl phthalate concentrations were positively associated with high-quality cleavage stage embryos (OR = 1.20, 95% CI 1

  20. Notch and Delta mRNAs in early-stage and mid-stage Drosophila embryos exhibit complementary patterns of protein producing potentials

    PubMed Central

    Shepherd, Andrew; Wesley, Uma; Wesley, Cedric

    2010-01-01

    Notch and Delta proteins generate Notch signaling that specifies cell fates during animal development. There is an intriguing phenomenon in Drosophila embryogenesis that has not received much attention and whose significance to embryogenesis is unknown. Notch and Delta mRNAs expressed in early-stage embryos are shorter than their counterparts in mid-stage embryos. We show here that the difference in sizes is due to mRNA 3′ processing at alternate polyadenylation sites. While the early-stage Notch mRNA has a lower protein-producing potential than the mid-stage Notch mRNA, the early-stage Delta mRNA has a higher protein-producing potential than the mid-stage Delta mRNA. Our data can explain the complementary patterns of Notch and Delta protein levels in early-stage and mid-stage embryos. Our data also raise the possibility that the manner and regulation of Notch signaling change in the course of embryogenesis and that this change is effected by 3′ UTR and mRNA 3′ processing factors. PMID:20201103

  1. Forskolin improves the cryosurvival of in vivo-derived porcine embryos at very early stages using two vitrification methods.

    PubMed

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

    2013-04-01

    This study was aimed to determine the effect of forskolin on the viability of in vivo-derived porcine embryos vitrified by the superfine open pulled straw (SOPS) or solid surface vitrification (SSV) methods at the 2-cell, 4-cell, and blastocyst stages. Zygotes, 2- to 4-cell embryos, and morulae were obtained from superovulated sows. After collection, embryos were cultured for 24h with 0 or 10 μM forskolin and then vitrified using the SOPS and SSV method, or not vitrified (fresh controls). Fresh and vitrified-warmed 2-cells, 4-cells, and blastocysts were cultured for additional 96 h, 72 h and 24 h, respectively. At the end of the culture, embryos were evaluated for progression to the blastocyst stage and total cell number. The vitrification method did not affect any of the parameters evaluated for any embryo stage. Forskolin increased (P<0.01) the blastocyst formation and the final developmental stage of vitrified 2- and 4-cell embryos. However, these embryos exhibited lower (P<0.003) blastocyst formation rates than their fresh counterparts. The total cell number and hatching rate were similar in both groups (vitrified and fresh) of 2- and 4-cell embryos. Vitrified blastocysts exhibited viabilities, final developmental stages, hatching rates, and total cell numbers that were similar to those of their fresh counterparts, regardless of the addition of forskolin. In conclusion, the SOPS and SSV methods are suitable for the cryopreservation of in vivo-derived 2- to 4-cell porcine embryos. Pre-treatment with forskolin for 24h before vitrification improves the cryotolerance of 2- and 4-cell porcine embryos. Copyright © 2013 Elsevier Inc. All rights reserved.

  2. Immunogenicity and safety of purified chick-embryo cell rabies vaccine under Zagreb 2-1-1 or 5-dose Essen regimen in Chinese children 6 to 17 years old and adults over 50 years: A randomized open-label study

    PubMed Central

    Li, RongCheng; Li, YanPing; Wen, ShuQing; Wen, HuiChun; Nong, Yi; Mo, Zhaojun; Xie, Fang; Pellegrini, Michele

    2015-01-01

    The aim of this Phase IIIb, open-label, randomized study was to demonstrate the non-inferiority of immune responses and to assess the safety of a purified chick-embryo cell rabies vaccine (PCECV) in healthy Chinese children (6 to 17 years) and older adults (≥51 years) following 2 alternative intramuscular (IM) simulated post-exposure prophylaxis (PEP) regimens: 4-dose Zagreb or 5-dose Essen regimen. Serum samples were collected prior to vaccination on Days 1 and 15 and on day 43 to assess immune response by rabies virus neutralizing antibody (RVNA) concentrations. Solicited adverse events (AEs) were recorded for up to 7 days following each vaccine dose, and unsolicited AEs throughout the entire study period. PCECV vaccination induced a strong immune response at Day 15, and the non-inferiority in immune response of the Zagreb vs. the Essen regimen was demonstrated in children and older adults. At Day 15,100% of children (N = 224), and 99% of subjects ≥51 years of age (N = 376) developed adequate RVNA concentrations (≥0.5 IU/mL); at Day 43 all subjects achieved RVNA concentrations ≥0.5 IU/mL, for both PEP regimens. The well-known tolerability and safety profile of the PCECV was again observed in this study following either Zagreb or Essen regimens. Rabies PEP vaccination with PCECV following a Zagreb regimen induced immune responses non-inferior to those of the Essen regimen, and had a similar safety and tolerability profile to the Essen regimen in Chinese children, adolescents, and adults over 51 years. ClinicalTrials.gov identifier: NCT01680016. PMID:25692350

  3. Immunogenicity and safety of purified chick-embryo cell rabies vaccine under Zagreb 2-1-1 or 5-dose Essen regimen in Chinese children 6 to 17 years old and adults over 50 years: a randomized open-label study.

    PubMed

    Li, RongCheng; Li, YanPing; Wen, ShuQing; Wen, HuiChun; Nong, Yi; Mo, Zhaojun; Xie, Fang; Pellegrini, Michele

    2015-01-01

    The aim of this Phase IIIb, open-label, randomized study was to demonstrate the non-inferiority of immune responses and to assess the safety of a purified chick-embryo cell rabies vaccine (PCECV) in healthy Chinese children (6 to 17 years) and older adults (≥51 years) following 2 alternative intramuscular (IM) simulated post-exposure prophylaxis (PEP) regimens: 4-dose Zagreb or 5-dose Essen regimen. Serum samples were collected prior to vaccination on Days 1 and 15 and on day 43 to assess immune response by rabies virus neutralizing antibody (RVNA) concentrations. Solicited adverse events (AEs) were recorded for up to 7 days following each vaccine dose, and unsolicited AEs throughout the entire study period. PCECV vaccination induced a strong immune response at Day 15, and the non-inferiority in immune response of the Zagreb vs. the Essen regimen was demonstrated in children and older adults. At Day 15,100% of children (N = 224), and 99% of subjects ≥51 years of age (N = 376) developed adequate RVNA concentrations (≥0.5 IU/mL); at Day 43 all subjects achieved RVNA concentrations ≥0.5 IU/mL, for both PEP regimens. The well-known tolerability and safety profile of the PCECV was again observed in this study following either Zagreb or Essen regimens. Rabies PEP vaccination with PCECV following a Zagreb regimen induced immune responses non-inferior to those of the Essen regimen, and had a similar safety and tolerability profile to the Essen regimen in Chinese children, adolescents, and adults over 51 years. ClinicalTrials.gov identifier: NCT01680016.

  4. P-TEFb, the Super Elongation Complex and Mediator Regulate a Subset of Non-paused Genes during Early Drosophila Embryo Development

    PubMed Central

    Dahlberg, Olle; Shilkova, Olga; Tang, Min; Holmqvist, Per-Henrik; Mannervik, Mattias

    2015-01-01

    Positive Transcription Elongation Factor b (P-TEFb) is a kinase consisting of Cdk9 and Cyclin T that releases RNA Polymerase II (Pol II) into active elongation. It can assemble into a larger Super Elongation Complex (SEC) consisting of additional elongation factors. Here, we use a miRNA-based approach to knock down the maternal contribution of P-TEFb and SEC components in early Drosophila embryos. P-TEFb or SEC depletion results in loss of cells from the embryo posterior and in cellularization defects. Interestingly, the expression of many patterning genes containing promoter-proximal paused Pol II is relatively normal in P-TEFb embryos. Instead, P-TEFb and SEC are required for expression of some non-paused, rapidly transcribed genes in pre-cellular embryos, including the cellularization gene Serendipity-α. We also demonstrate that another P-TEFb regulated gene, terminus, has an essential function in embryo development. Similar morphological and gene expression phenotypes were observed upon knock down of Mediator subunits, providing in vivo evidence that P-TEFb, the SEC and Mediator collaborate in transcription control. Surprisingly, P-TEFb depletion does not affect the ratio of Pol II at the promoter versus the 3’ end, despite affecting global Pol II Ser2 phosphorylation levels. Instead, Pol II occupancy is reduced at P-TEFb down-regulated genes. We conclude that a subset of non-paused, pre-cellular genes are among the most susceptible to reduced P-TEFb, SEC and Mediator levels in Drosophila embryos. PMID:25679530

  5. Spatial Reorganization of the Endoplasmic Reticulum during Mitosis Relies on Mitotic Kinase Cyclin A in the Early Drosophila Embryo

    PubMed Central

    Bergman, Zane J.; Mclaurin, Justin D.; Eritano, Anthony S.; Johnson, Brittany M.; Sims, Amanda Q.; Riggs, Blake

    2015-01-01

    Mitotic cyclin-dependent kinase with their cyclin partners (cyclin:Cdks) are the master regulators of cell cycle progression responsible for regulating a host of activities during mitosis. Nuclear mitotic events, including chromosome condensation and segregation have been directly linked to Cdk activity. However, the regulation and timing of cytoplasmic mitotic events by cyclin:Cdks is poorly understood. In order to examine these mitotic cytoplasmic events, we looked at the dramatic changes in the endoplasmic reticulum (ER) during mitosis in the early Drosophila embryo. The dynamic changes of the ER can be arrested in an interphase state by inhibition of either DNA or protein synthesis. Here we show that this block can be alleviated by micro-injection of Cyclin A (CycA) in which defined mitotic ER clusters gathered at the spindle poles. Conversely, micro-injection of Cyclin B (CycB) did not affect spatial reorganization of the ER, suggesting CycA possesses the ability to initiate mitotic ER events in the cytoplasm. Additionally, RNAi-mediated simultaneous inhibition of all 3 mitotic cyclins (A, B and B3) blocked spatial reorganization of the ER. Our results suggest that mitotic ER reorganization events rely on CycA and that control and timing of nuclear and cytoplasmic events during mitosis may be defined by release of CycA from the nucleus as a consequence of breakdown of the nuclear envelope. PMID:25689737

  6. Imaging of Norway spruce early somatic embryos with the ESEM, Cryo-SEM and laser scanning microscope.

    PubMed

    Neděla, Vilém; Hřib, Jiří; Havel, Ladislav; Hudec, Jiří; Runštuk, Jiří

    2016-05-01

    This article describes the surface structure of Norway spruce early somatic embryos (ESEs) as a typical culture with asynchronous development. The microstructure of extracellular matrix covering ESEs were observed using the environmental scanning electron microscope as a primary tool and using the scanning electron microscope with cryo attachment and laser electron microscope as a complementary tool allowing our results to be proven independently. The fresh samples were observed in conditions of the air environment of the environmental scanning electron microscope (ESEM) with the pressure from 550Pa to 690Pa and the low temperature of the sample from -18°C to -22°C. The samples were studied using two different types of detector to allow studying either the thin surface structure or material composition. The scanning electron microscope with cryo attachment was used for imaging frozen extracellular matrix microstructure with higher resolution. The combination of both electron microscopy methods was suitable for observation of "native" plant samples, allowing correct evaluation of our results, free of error and artifacts. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Developmental insights from early mammalian embryos and core signaling pathways that influence human pluripotent cell growth and differentiation.

    PubMed

    Chen, Kevin G; Mallon, Barbara S; Johnson, Kory R; Hamilton, Rebecca S; McKay, Ronald D G; Robey, Pamela G

    2014-05-01

    Human pluripotent stem cells (hPSCs) have two potentially attractive applications: cell replacement-based therapies and drug discovery. Both require the efficient generation of large quantities of clinical-grade stem cells that are free from harmful genomic alterations. The currently employed colony-type culture methods often result in low cell yields, unavoidably heterogeneous cell populations, and substantial chromosomal abnormalities. Here, we shed light on the structural relationship between hPSC colonies/embryoid bodies and early-stage embryos in order to optimize current culture methods based on the insights from developmental biology. We further highlight core signaling pathways that underlie multiple epithelial-to-mesenchymal transitions (EMTs), cellular heterogeneity, and chromosomal instability in hPSCs. We also analyze emerging methods such as non-colony type monolayer (NCM) and suspension culture, which provide alternative growth models for hPSC expansion and differentiation. Furthermore, based on the influence of cell-cell interactions and signaling pathways, we propose concepts, strategies, and solutions for production of clinical-grade hPSCs, stem cell precursors, and miniorganoids, which are pivotal steps needed for future clinical applications. Published by Elsevier B.V.

  8. Global regulation of mRNA translation and stability in the early Drosophila embryo by the Smaug RNA-binding protein

    PubMed Central

    2014-01-01

    Background Smaug is an RNA-binding protein that induces the degradation and represses the translation of mRNAs in the early Drosophila embryo. Smaug has two identified direct target mRNAs that it differentially regulates: nanos and Hsp83. Smaug represses the translation of nanos mRNA but has only a modest effect on its stability, whereas it destabilizes Hsp83 mRNA but has no detectable effect on Hsp83 translation. Smaug is required to destabilize more than one thousand mRNAs in the early embryo, but whether these transcripts represent direct targets of Smaug is unclear and the extent of Smaug-mediated translational repression is unknown. Results To gain a panoramic view of Smaug function in the early embryo, we identified mRNAs that are bound to Smaug using RNA co-immunoprecipitation followed by hybridization to DNA microarrays. We also identified mRNAs that are translationally repressed by Smaug using polysome gradients and microarrays. Comparison of the bound mRNAs to those that are translationally repressed by Smaug and those that require Smaug for their degradation suggests that a large fraction of Smaug’s target mRNAs are both translationally repressed and degraded by Smaug. Smaug directly regulates components of the TRiC/CCT chaperonin, the proteasome regulatory particle and lipid droplets, as well as many metabolic enzymes, including several glycolytic enzymes. Conclusions Smaug plays a direct and global role in regulating the translation and stability of a large fraction of the mRNAs in the early Drosophila embryo, and has unanticipated functions in control of protein folding and degradation, lipid droplet function and metabolism. PMID:24393533

  9. Global regulation of mRNA translation and stability in the early Drosophila embryo by the Smaug RNA-binding protein.

    PubMed

    Chen, Linan; Dumelie, Jason G; Li, Xiao; Cheng, Matthew Hk; Yang, Zhiyong; Laver, John D; Siddiqui, Najeeb U; Westwood, J Timothy; Morris, Quaid; Lipshitz, Howard D; Smibert, Craig A

    2014-01-07

    Smaug is an RNA-binding protein that induces the degradation and represses the translation of mRNAs in the early Drosophila embryo. Smaug has two identified direct target mRNAs that it differentially regulates: nanos and Hsp83. Smaug represses the translation of nanos mRNA but has only a modest effect on its stability, whereas it destabilizes Hsp83 mRNA but has no detectable effect on Hsp83 translation. Smaug is required to destabilize more than one thousand mRNAs in the early embryo, but whether these transcripts represent direct targets of Smaug is unclear and the extent of Smaug-mediated translational repression is unknown. To gain a panoramic view of Smaug function in the early embryo, we identified mRNAs that are bound to Smaug using RNA co-immunoprecipitation followed by hybridization to DNA microarrays. We also identified mRNAs that are translationally repressed by Smaug using polysome gradients and microarrays. Comparison of the bound mRNAs to those that are translationally repressed by Smaug and those that require Smaug for their degradation suggests that a large fraction of Smaug's target mRNAs are both translationally repressed and degraded by Smaug. Smaug directly regulates components of the TRiC/CCT chaperonin, the proteasome regulatory particle and lipid droplets, as well as many metabolic enzymes, including several glycolytic enzymes. Smaug plays a direct and global role in regulating the translation and stability of a large fraction of the mRNAs in the early Drosophila embryo, and has unanticipated functions in control of protein folding and degradation, lipid droplet function and metabolism.

  10. Oxidative Damage to Rhesus Macaque Spermatozoa Results in Mitotic Arrest and Transcript Abundance Changes in Early Embryos1

    PubMed Central

    Burruel, Victoria; Klooster, Katie L.; Chitwood, James; Ross, Pablo J.; Meyers, Stuart A.

    2013-01-01

    ABSTRACT Our objective was to determine whether oxidative damage of rhesus macaque sperm induced by reactive oxygen species (ROS) in vitro would affect embryo development following intracytoplasmic sperm injection (ICSI) of metaphase II (MII) oocytes. Fresh rhesus macaque spermatozoa were treated with ROS as follows: 1 mM xanthine and 0.1 U/ml xanthine oxidase (XXO) at 37°C and 5% CO2 in air for 2.25 h. Sperm were then assessed for motility, viability, and lipid peroxidation. Motile ROS-treated and control sperm were used for ICSI of MII oocytes. Embryo culture was evaluated for 3 days for development to the eight-cell stage. Embryos were fixed and stained for signs of cytoplasmic and nuclear abnormalities. Gene expression was analyzed by RNA-Seq in two-cell embryos from control and treated groups. Exposure of sperm to XXO resulted in increased lipid peroxidation and decreased sperm motility. ICSI of MII oocytes with motile sperm induced similar rates of fertilization and cleavage between treatments. Development to four- and eight-cell stage was significantly lower for embryos generated with ROS-treated sperm than for controls. All embryos produced from ROS-treated sperm demonstrated permanent embryonic arrest and varying degrees of degeneration and nuclear fragmentation, changes that are suggestive of prolonged senescence or apoptotic cell death. RNA-Seq analysis of two-cell embryos showed changes in transcript abundance resulting from sperm treatment with ROS. Differentially expressed genes were enriched for processes associated with cytoskeletal organization, cell adhesion, and protein phosphorylation. ROS-induced damage to sperm adversely affects embryo development by contributing to mitotic arrest after ICSI of MII rhesus oocytes. Changes in transcript abundance in embryos destined for mitotic arrest is evident at the two-cell stage of development. PMID:23904511

  11. Identification and functional analysis of long non-coding RNAs in human and mouse early embryos based on single-cell transcriptome data

    PubMed Central

    Qiu, Jia-jun; Ren, Zhao-rui; Yan, Jing-bin

    2016-01-01

    Epigenetics regulations have an important role in fertilization and proper embryonic development, and several human diseases are associated with epigenetic modification disorders, such as Rett syndrome, Beckwith-Wiedemann syndrome and Angelman syndrome. However, the dynamics and functions of long non-coding RNAs (lncRNAs), one type of epigenetic regulators, in human pre-implantation development have not yet been demonstrated. In this study, a comprehensive analysis of human and mouse early-stage embryonic lncRNAs was performed based on public single-cell RNA sequencing data. Expression profile analysis revealed that lncRNAs are expressed in a developmental stage–specific manner during human early-stage embryonic development, whereas a more temporal-specific expression pattern was identified in mouse embryos. Weighted gene co-expression network analysis suggested that lncRNAs involved in human early-stage embryonic development are associated with several important functions and processes, such as oocyte maturation, zygotic genome activation and mitochondrial functions. We also found that the network of lncRNAs involved in zygotic genome activation was highly preservative between human and mouse embryos, whereas in other stages no strong correlation between human and mouse embryo was observed. This study provides insight into the molecular mechanism underlying lncRNA involvement in human pre-implantation embryonic development. PMID:27542205

  12. Characterization of cDNAs encoding the chick retinoic acid receptor gamma 2 and preferential distribution of retinoic acid receptor gamma transcripts during chick skin development.

    PubMed

    Michaille, J J; Blanchet, S; Kanzler, B; Garnier, J M; Dhouailly, D

    1994-12-01

    Retinoic acid receptors alpha, beta and gamma (RAR alpha, beta and gamma) are ligand-inductible transcriptional activators which belong to the steroid/thyroid hormone receptor superfamily. At least two major isoforms (1 and 2) of each RAR arise by differential use of two promoters and alternative splicing. In mouse, the three RAR genes are expressed in stage- and tissue-specific patterns during embryonic development. In order to understand the role of the different RARs in chick, RAR gamma 2 cDNAs were isolated from an 8.5-day (stage 35 of Hamburger and Hamilton) chick embryo skin library. The deduced chick RAR gamma 2 amino acid sequence displays uncommon features such as 21 specific amino acid replacements, 12 of them being clustered in the amino-terminal region (domains A2 and B), and a truncated acidic carboxy-terminal region (F domain). However, the pattern of RAR gamma expression in chick embryo resembles that reported in mouse, particularly in skin where RAR gamma expression occurs in both the dermal and epidermal layers at the beginning of feather formation, and is subsequently restricted to the differentiating epidermal cells. Northern blot analysis suggests that different RAR gamma isoforms could be successively required during chick development.

  13. Development of the genital ducts and external genitalia in the early human embryo.

    PubMed

    Sajjad, Yasmin

    2010-10-01

    The course of development of the human genital tract is undifferentiated to the 9th week of development. At this time two symmetrical paired ducts known as the mesonephric (MD) and paramesonephric ducts (PMD) are present, which together with the urogenital sinus provide the tissue sources for internal and external genital development. Normal differentiation of the bipotential external genitalia and reproductive ducts are dependent upon the presence or absence of certain hormones. Masculinization of the internal and external genitalia during fetal development depends on the existence of two discrete testicular hormones. Testosterone secreted from Leydig cells induces the differentiation of the mesonephric ducts into the epididymis, vasa deferentia and seminal vesicles, whereas anti-Müllerian hormone (AMH) produced by Sertoli cells induces the regression of the paramesonephric ducts. The absence of AMH action in early fetal life results in the formation of the fallopian tubes, uterus and upper third of the vagina. In some target tissues, testosterone is converted to dihydrotestosterone, which is responsible for the masculinization of the urogenital sinus and external genitalia. © 2010 The Author. Journal of Obstetrics and Gynaecology Research © 2010 Japan Society of Obstetrics and Gynecology.

  14. Position- and Hippo signaling-dependent plasticity during lineage segregation in the early mouse embryo

    PubMed Central

    Posfai, Eszter; Petropoulos, Sophie; de Barros, Flavia Regina Oliveira; Schell, John Paul; Jurisica, Igor; Sandberg, Rickard; Lanner, Fredrik; Rossant, Janet

    2017-01-01

    The segregation of the trophectoderm (TE) from the inner cell mass (ICM) in the mouse blastocyst is determined by position-dependent Hippo signaling. However, the window of responsiveness to Hippo signaling, the exact timing of lineage commitment and the overall relationship between cell commitment and global gene expression changes are still unclear. Single-cell RNA sequencing during lineage segregation revealed that the TE transcriptional profile stabilizes earlier than the ICM and prior to blastocyst formation. Using quantitative Cdx2-eGFP expression as a readout of Hippo signaling activity, we assessed the experimental potential of individual blastomeres based on their level of Cdx2-eGFP expression and correlated potential with gene expression dynamics. We find that TE specification and commitment coincide and occur at the time of transcriptional stabilization, whereas ICM cells still retain the ability to regenerate TE up to the early blastocyst stage. Plasticity of both lineages is coincident with their window of sensitivity to Hippo signaling. DOI: http://dx.doi.org/10.7554/eLife.22906.001 PMID:28226240

  15. Organogenesis of the Musculoskeletal System in Horse Embryos and Early Fetuses.

    PubMed

    Barreto, Rodrigo da Silva Nunes; Rodrigues, Márcio Nogueira; Carvalho, Rafael Cardoso; De Oliveira E Silva, Fernanda Menezes; Rigoglio, Náthia Nathaly; Jacob, Júlio César Ferraz; Gastal, Eduardo Leite; Miglino, Maria Angélica

    2016-06-01

    Musculoskeletal system development involves heterotypical inductive interactions between tendons, muscles, and cartilage and knowledge on organogenesis is required for clarification of its function. The aim of this study was to describe the organogenesis of horse musculoskeletal system between 21 and 105 days of gestation, using detailed macroscopic and histological analyses focusing on essential developmental steps. At day 21 of gestation the skin was translucid, but epithelial condensation and fibrocartilaginous tissues were observed on day 25 of pregnancy. Smooth muscle was seen in lymphatic and blood vessel walls and the beginning of cartilaginous chondrocranium was detected at day 30 of gestation. At day 45, typical chondroblasts and chondrocytes were observed and at day 55, mandibular processes expanded toward the ventral midline of the pharynx. At day 75, muscles became thicker and muscle fibers were seen developing in carpal and metacarpal joints with the beginning of the ossification process. At day 105, major muscle groups, similar to those seen in an adult equine, were observed. The caudal area of the nasal capsule and trabecular cartilages increased in size and became ossified, developing into the ethmoid bone. The presence of nasal, frontal, parietal, and occipital bones was observed. In conclusion, novel features of equine musculoskeletal system development have been described here and each process was linked with an early musculoskeletal event. Data presented herein will facilitate a better understanding of the equine muscular system organogenesis and aid in the detection of congenital deformities. Anat Rec, 299:722-729, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Towards a CRISPR view of early human development: applications, limitations and ethical concerns of genome editing in human embryos.

    PubMed

    Plaza Reyes, Alvaro; Lanner, Fredrik

    2017-01-01

    Developmental biologists have become increasingly aware that the wealth of knowledge generated through genetic studies of pre-implantation mouse development might not easily be translated to the human embryo. Comparative studies have been fueled by recent technological advances in single-cell analysis, allowing in-depth analysis of the human embryo. This field could shortly gain more momentum as novel genome editing technologies might, for the first time, also allow functional genetic studies in the human embryo. In this Spotlight article, we summarize the CRISPR-Cas9 genome editing system and discuss its potential applications and limitations in human pre-implantation embryos, and the ethical considerations thereof. © 2017. Published by The Company of Biologists Ltd.

  17. Chorio-Allantoic Membrane Grafting of Chick Limb Buds as a Class Practical.

    ERIC Educational Resources Information Center

    McLachlan, John C.

    1981-01-01

    A new method of carrying out grafts of early embryonic chick limb buds to the chick chorio-allantoic membrane and a processing schedule which renders cartilage elements visible in whole mount are discussed, including implications for the procedures and their results. (Author/DC)

  18. Olfactory lateralization in the chick.

    PubMed

    Vallortigara, G; Andrew, R J

    1994-04-01

    Chicks using their right nostril (and so with direct olfactory input to the right hemisphere), and presented simultaneously with two objects identical in visual appearance with the rearing object, and differing only in odour, chose that which smelled like the rearing object. Chicks using the left nostril chose equally readily but at random. Earlier work, using similar tests, has shown special interest of the right hemisphere in change in visual properties of familiar stimuli, suggesting that analysis of a wide range of properties of a familiar stimulus may be an important function of the right hemisphere in the chick, with consequent detection of novelty.

  19. Localization of basic fibroblast growth factor binding sites in the chick embryonic neural retina.

    PubMed

    Cirillo, A; Arruti, C; Courtois, Y; Jeanny, J C

    1990-12-01

    We have investigated the localization of basic fibroblast growth factor (bFGF) binding sites during the development of the neural retina in the chick embryo. The specificity of the affinity of bFGF for its receptors was assessed by competition experiments with unlabelled growth factor or with heparin, as well as by heparitinase treatment of the samples. Two different types of binding sites were observed in the neural retina by light-microscopic autoradiography. The first type, localized mainly to basement membranes, was highly sensitive to heparitinase digestion and to competition with heparin. It was not developmentally regulated. The second type of binding site, resistant to heparin competition, appeared to be associated with retinal cells from the earliest stages studied (3-day-old embryo, stages 21-22 of Hamburger and Hamilton). Its distribution was found to vary during embryonic development, paralleling layering of the neural retina. Binding of bFGF to the latter sites was observed throughout the retinal neuroepithelium at early stages but displayed a distinct pattern at the time when the inner and outer plexiform layers were formed. During the development of the inner plexiform layer, a banded pattern of bFGF binding was observed. These bands, lying parallel to the vitreal surface, seemed to codistribute with the synaptic bands existing in the inner plexiform layer. The presence of intra-retinal bFGF binding sites whose distribution varies with embryonic development suggests a regulatory mechanism involving differential actions of bFGF on neural retinal cells.

  20. Prenatal sensory experience affects hatching behavior in domestic chicks (Gallus gallus) and Japanese quail chicks (Coturnix coturnix japonica).

    PubMed

    Sleigh, Merry J; Casey, Michael B

    2014-07-01

    Species-typical developmental outcomes result from organismic and environmental constraints and experiences shared by members of a species. We examined the effects of enhanced prenatal sensory experience on hatching behaviors by exposing domestic chicks (n = 95) and Japanese quail (n = 125) to one of four prenatal conditions: enhanced visual stimulation, enhanced auditory stimulation, enhanced auditory and visual stimulation, or no enhanced sensory experience (control condition). In general, across species, control embryos had slower hatching behaviors than all other embryos. Embryos in the auditory condition had faster hatching behaviors than embryos in the visual and control conditions. Auditory-visual condition embryos showed similarities to embryos exposed to either auditory or visual stimulation. These results suggest that prenatal sensory experience can influence hatching behavior of precocial birds, with the type of stimulation being a critical variable. These results also provide further evidence that species-typical outcomes are the result of species-typical prenatal experiences. © 2013 Wiley Periodicals, Inc.

  1. Cloning, expression pattern, and potential role of apoptosis inhibitor 5 in the termination of embryonic diapause and early embryo development of Artemia sinica.

    PubMed

    Zhang, Shuang; Yao, Feng; Jing, Ting; Zhang, Mengchen; Zhao, Wei; Zou, Xiangyang; Sui, Linlin; Hou, Lin

    2017-09-10

    During the embryonic development of Artemia sinica, the diapause phenomenon can be induced by high salinity or low temperature conditions. The diapause embryo at the gastrula stage is maintained under the threat of apoptosis to guarantee the embryo's normal development. In this process, apoptosis inhibitor proteins play vital roles in protecting embryos against apoptosis. Apoptosis inhibitor5 (API5) plays a pivotal role in regulating the cell cycle and preventing programmed cell death after growth factor starvation. In the present study, we cloned the full-length cDNA representing the api5 gene from A. sinica (As-api5), which encodes a 372-amino acid protein. In situ hybridization experiments revealed that As-api5 expression is not tissue or organ specific. Quantitative real-time PCR analyses of the developmental expression of As-api5 showed that it reached its highest level at 10h, after which its expression decreased. High salinity and low temperature treatments increased the expression of As-api5. Western blotting was used to assess the abundance of As-API5 and related proteins (As-CyclinA, As-CyclinE, As-E2F1, As-CDK2, As-APAF1, and As-Caspase9). Downregulation of As-api5 expression using a short interfering RNA resulted in increased mortality and embryo malformation of A. sinica. Taken together, the results indicated that API5 plays a crucial role in embryonic diapause termination and early embryo development of A. sinica. Copyright © 2017. Published by Elsevier B.V.

  2. The C. elegans homolog of Drosophila Lethal giant larvae functions redundantly with PAR-2 to maintain polarity in the early embryo.

    PubMed

    Beatty, Alexander; Morton, Diane; Kemphues, Kenneth

    2010-12-01

    Polarity is essential for generating cell diversity. The one-cell C. elegans embryo serves as a model for studying the establishment and maintenance of polarity. In the early embryo, a myosin II-dependent contraction of the cortical meshwork asymmetrically distributes the highly conserved PDZ proteins PAR-3 and PAR-6, as well as an atypical protein kinase C (PKC-3), to the anterior. The RING-finger protein PAR-2 becomes enriched on the posterior cortex and prevents these three proteins from returning to the posterior. In addition to the PAR proteins, other proteins are required for polarity in many metazoans. One example is the conserved Drosophila tumor-suppressor protein Lethal giant larvae (Lgl). In Drosophila and mammals, Lgl contributes to the maintenance of cell polarity and plays a role in asymmetric cell division. We have found that the C. elegans homolog of Lgl, LGL-1, has a role in polarity but is not essential. It localizes asymmetrically to the posterior of the early embryo in a PKC-3-dependent manner, and functions redundantly with PAR-2 to maintain polarity. Furthermore, overexpression of LGL-1 is sufficient to rescue loss of PAR-2 function. LGL-1 negatively regulates the accumulation of myosin (NMY-2) on the posterior cortex, representing a possible mechanism by which LGL-1 might contribute to polarity maintenance.

  3. Myosin-1 inhibition by PClP affects membrane shape, cortical actin distribution and lipid droplet dynamics in early Zebrafish embryos

    PubMed Central

    Gupta, Prabuddha; Martin, René; Knölker, Hans-Joachim; Nihalani, Deepak; Kumar Sinha, Deepak

    2017-01-01

    Myosin-1 (Myo1) represents a mechanical link between the membrane and actin-cytoskeleton in animal cells. We have studied the effect of Myo1 inhibitor PClP in 1–8 cell Zebrafish embryos. Our results indicate a unique involvement of Myo1 in early development of Zebrafish embryos. Inhibition of Myo1 (by PClP) and Myo2 (by Blebbistatin) lead to arrest in cell division. While Myo1 isoforms appears to be important for both the formation and the maintenance of cleavage furrows, Myo2 is required only for the formation of furrows. We found that the blastodisc of the embryo, which contains a thick actin cortex (~13 μm), is loaded with cortical Myo1. Myo1 appears to be crucial for maintaining the blastodisc morphology and the actin cortex thickness. In addition to cell division and furrow formation, inhibition of Myo1 has a drastic effect on the dynamics and distribution of lipid droplets (LDs) in the blastodisc near the cleavage furrow. All these results above are effects of Myo1 inhibition exclusively; Myo2 inhibition by blebbistatin does not show such phenotypes. Therefore, our results demonstrate a potential role for Myo1 in the maintenance and formation of furrow, blastodisc morphology, cell-division and LD organization within the blastodisc during early embryogenesis. PMID:28678859

  4. The Armadillo Repeat Gene ZAK IXIK Promotes Arabidopsis Early Embryo and Endosperm Development through a Distinctive Gametophytic Maternal Effect[C][W][OA

    PubMed Central

    Ngo, Quy A.; Baroux, Celia; Guthörl, Daniela; Mozerov, Peter; Collinge, Margaret A.; Sundaresan, Venkatesan; Grossniklaus, Ueli

    2012-01-01

    The proper balance of parental genomic contributions to the fertilized embryo and endosperm is essential for their normal growth and development. The characterization of many gametophytic maternal effect (GME) mutants affecting seed development indicates that there are certain classes of genes with a predominant maternal contribution. We present a detailed analysis of the GME mutant zak ixik (zix), which displays delayed and arrested growth at the earliest stages of embryo and endosperm development. ZIX encodes an Armadillo repeat (Arm) protein highly conserved across eukaryotes. Expression studies revealed that ZIX manifests a GME through preferential maternal expression in the early embryo and endosperm. This parent-of-origin–dependent expression is regulated by neither the histone and DNA methylation nor the DNA demethylation pathways known to regulate some other GME mutants. The ZIX protein is localized in the cytoplasm and nucleus of cells in reproductive tissues and actively dividing root zones. The maternal ZIX allele is required for the maternal expression of MINISEED3. Collectively, our results reveal a reproductive function of plant Arm proteins in promoting early seed growth, which is achieved through a distinct GME of ZIX that involves mechanisms for maternal allele-specific expression that are independent of the well-established pathways. PMID:23064319

  5. The specification and global reprogramming of histone epigenetic marks during gamete formation and early embryo development in C. elegans.

    PubMed

    Samson, Mark; Jow, Margaret M; Wong, Catherine C L; Fitzpatrick, Colin; Aslanian, Aaron; Saucedo, Israel; Estrada, Rodrigo; Ito, Takashi; Park, Sung-kyu Robin; Yates, John R; Chu, Diana S

    2014-10-01

    In addition to the DNA contributed by sperm and oocytes, embryos receive parent-specific epigenetic information that can include histone variants, histone post-translational modifications (PTMs), and DNA methylation. However, a global view of how such marks are erased or retained during gamete formation and reprogrammed after fertilization is lacking. To focus on features conveyed by histones, we conducted a large-scale proteomic identification of histone variants and PTMs in sperm and mixed-stage embryo chromatin from C. elegans, a species that lacks conserved DNA methylation pathways. The fate of these histone marks was then tracked using immunostaining. Proteomic analysis found that sperm harbor ∼2.4 fold lower levels of histone PTMs than embryos and revealed differences in classes of PTMs between sperm and embryos. Sperm chromatin repackaging involves the incorporation of the sperm-specific histone H2A variant HTAS-1, a widespread erasure of histone acetylation, and the retention of histone methylation at sites that mark the transcriptional history of chromatin domains during spermatogenesis. After fertilization, we show HTAS-1 and 6 histone PTM marks distinguish sperm and oocyte chromatin in the new embryo and characterize distinct paternal and maternal histone remodeling events during the oocyte-to-embryo transition. These include the exchange of histone H2A that is marked by ubiquitination, retention of HTAS-1, removal of the H2A variant HTZ-1, and differential reprogramming of histone PTMs. This work identifies novel and conserved features of paternal chromatin that are specified during spermatogenesis and processed in the embryo. Furthermore, our results show that different species, even those with diverged DNA packaging and imprinting strategies, use conserved histone modification and removal mechanisms to reprogram epigenetic information.

  6. A potential determinant role of adiponectin and receptors for the early embryo development in PCOS patients with obesity hinted by quantitative profiling.

    PubMed

    Zhang, Ning; Hao, Cuifang; Liu, Xiaoyan; Zhang, Shouxin; Zhang, Fengrong; Zhuang, Lili; Zhao, Dongmei

    2017-02-01

    To identify the quantitative profiling of adiponectin and its receptors (AdipoR1, AdipoR2, and T-cadherin) in cumulus cells (CCs) and to evaluate their roles in the early embryo development of polycystic ovary syndrome (PCOS) patients, in part, with obesity. Fifty-five subjects were divided into two groups according to the body mass index. Oocytes were further inseminated and only mature and normal fertilized oocytes (2PN) were included in this research. Real-time PCR and western blot were performed to identify adiponectin and its receptors in CCs. Adiponectin and receptors were ubiquitously expressed in CCs of PCOS and non-PCOS patients. The level of AdipoR2 in CCs from the oocytes yielding blastocyst after 5/6 days in vitro culture was markedly higher than in those from oocytes could not develop to blastocyst stage after Day 6, for non-obese or obese PCOS patients (0.1647 ± 0.0161 versus 0.0783 ± 0.0385, 0.1948 ± 0.0307 versus 0.1057 ± 0.0236, respectively, p < 0.05). In addition, only in patients with PCOS and concurrent obesity the AdipoR1 in CCs was considerably increased in CC-B + compared with CC-B - subgroup (0.5162 ± 0.0371 versus 0.2448 ± 0.0333, p < 0.01). The development of early embryo was associated with the up-regulation of AdipoR1 and AdipoR2 in PCOS patients. Our results suggested that adiponectin could positively modulate embryo development in humans. Further investigations should be carried out to unlock the crucial role that adiponectin plays in embryo development.

  7. Effects of prenatal exposure to antithyroid drugs on imprinting behavior in chicks.

    PubMed

    Kagami, Keisuke; Nishigori, Hidekazu; Nishigori, Hideo

    2010-09-01

    Thyroid hormones play important roles in vertebrate brain development. However, there is little understanding of the direct effects of fetal thyroid dysfunction, i.e., not acquired through the mother, on learning ability. In the present study, we use a chick embryo as a fetal model to investigate the effects of prenatal exposure to antithyroid drugs on imprinting behavior in hatched chicks. Methimazole (MMI) at 20micromol/egg or 5micromol/egg of propylthiouracil (PTU) was administered to eggs on day 14 while the control was given only a vehicle. An imprinting test was conducted after the chicks hatched. Day-old chicks were exposed to a rotating training object for 150min. The next day, the trained chicks were exposed to the training object and a novel object. The imprinting preference was represented as a preference score (PS) calculated as the rate of following the training object to following the training and novel objects. In the MMI-treated chicks, the PS was 0.68+/-0.06 (range, 0.38-0.88), which was significantly lower than that in the control chicks (0.86+/-0.04, p<0.01). In the PTU-treated chicks, the PS was 0.69+/-0.04 (range, 0.52-0.89), which was also significantly lower than that in the control (0.88+/-0.02, p<0.001). The present findings suggested that fetal thyroid dysfunction inhibited brain development, leading to impaired learning and memory. Our chick model can be considered useful for investigating the direct effects of prenatal exposure to antithyroid drugs or substances in the environment on learning ability after birth. Copyright 2010 Elsevier Inc. All rights reserved.

  8. Uterine responses to early pre-attachment embryos in the domestic dog and comparisons with other domestic animal species.

    PubMed

    Graubner, Felix R; Gram, Aykut; Kautz, Ewa; Bauersachs, Stefan; Aslan, Selim; Agaoglu, Ali R; Boos, Alois; Kowalewski, Mariusz P

    2017-08-01

    In the dog, there is no luteolysis in the absence of pregnancy. Thus, this species lacks any anti-luteolytic endocrine signal as found in other species that modulate uterine function during the critical period of pregnancy establishment. Nevertheless, in the dog an embryo-maternal communication must occur in order to prevent rejection of embryos. Based on this hypothesis, we performed microarray analysis of canine uterine samples collected during pre-attachment phase (days 10-12) and in corresponding non-pregnant controls, in order to elucidate the embryo attachment signal. An additional goal was to identify differences in uterine responses to pre-attachment embryos between dogs and other mammalian species exhibiting different reproductive patterns with regard to luteolysis, implantation, and preparation for placentation. Therefore, the canine microarray data were compared with gene sets from pigs, cattle, horses, and humans. We found 412 genes differentially regulated between the two experimental groups. The functional terms most strongly enriched in response to pre-attachment embryos related to extracellular matrix function and remodeling, and to immune and inflammatory responses. Several candidate genes were validated by semi-quantitative PCR. When compared with other species, best matches were found with human and equine counterparts. Especially for the pig, the majority of overlapping genes showed opposite expression patterns. Interestingly, 1926 genes did not pair with any of the other gene sets. Using a microarray approach, we report the uterine changes in the dog driven by the presence of embryos and compare these results with datasets from other mammalian species, finding common-, contrary-, and exclusively canine-regulated genes. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction.

  9. Uterine responses to early pre-attachment embryos in the domestic dog and comparisons with other domestic animal species†

    PubMed Central

    Graubner, Felix R.; Gram, Aykut; Kautz, Ewa; Bauersachs, Stefan; Aslan, Selim; Agaoglu, Ali R.; Boos, Alois

    2017-01-01

    Abstract In the dog, there is no luteolysis in the absence of pregnancy. Thus, this species lacks any anti-luteolytic endocrine signal as found in other species that modulate uterine function during the critical period of pregnancy establishment. Nevertheless, in the dog an embryo-maternal communication must occur in order to prevent rejection of embryos. Based on this hypothesis, we performed microarray analysis of canine uterine samples collected during pre-attachment phase (days 10-12) and in corresponding non-pregnant controls, in order to elucidate the embryo attachment signal. An additional goal was to identify differences in uterine responses to pre-attachment embryos between dogs and other mammalian species exhibiting different reproductive patterns with regard to luteolysis, implantation, and preparation for placentation. Therefore, the canine microarray data were compared with gene sets from pigs, cattle, horses, and humans. We found 412 genes differentially regulated between the two experimental groups. The functional terms most strongly enriched in response to pre-attachment embryos related to extracellular matrix function and remodeling, and to immune and inflammatory responses. Several candidate genes were validated by semi-quantitative PCR. When compared with other species, best matches were found with human and equine counterparts. Especially for the pig, the majority of overlapping genes showed opposite expression patterns. Interestingly, 1926 genes did not pair with any of the other gene sets. Using a microarray approach, we report the uterine changes in the dog driven by the presence of embryos and compare these results with datasets from other mammalian species, finding common-, contrary-, and exclusively canine-regulated genes. PMID:28651344

  10. Development of otoconia in the embryonic chick (Gallus domesticus)

    NASA Technical Reports Server (NTRS)

    Fermin, C. D.; Igarashi, M.

    1985-01-01

    In the chick (Gallus domesticus) embryo, otoconium formation started first over the macula sacculi around the 4th day of incubation, and a day later over the macula utriculi. It was determined that each otoconium formed as a result of the segmentation of the immature otolithic membrane, and that the calcium responsible for otoconium calcification was incorporated into the organic matrix of each otoconium in the form of small electron-dense granules (20-150 nm in. diameter). The presence of calcium in these granules was confirmed by histochemical staining with osmic-potassium pyroantimonate, by EDTA chelation, and by X-ray micronanalysis under the electron microscope.

  11. Evaluating the interaction between progesterone, TNF alpha and cortisol on early loss of transferred embryos in beef cows

    Fifty-eight non-lactating cows previously synchronized for estrus were assigned to two treatments to assess the effects of progesterone supplementation and its correlation with TNF-a and cortisol on the survival of the transferred embryos. On day 7 after exhibiting estrus (day 0), cows in both group...

  12. Spontaneous generalization of abstract multimodal patterns in young domestic chicks.

    PubMed

    Versace, Elisabetta; Spierings, Michelle J; Caffini, Matteo; Ten Cate, Carel; Vallortigara, Giorgio

    2017-05-01

    From the early stages of life, learning the regularities associated with specific objects is crucial for making sense of experiences. Through filial imprinting, young precocial birds quickly learn the features of their social partners by mere exposure. It is not clear though to what extent chicks can extract abstract patterns of the visual and acoustic stimuli present in the imprinting object, and how they combine them. To investigate this issue, we exposed chicks (Gallus gallus) to three days of visual and acoustic imprinting, using either patterns with two identical items or patterns with two different items, presented visually, acoustically or in both modalities. Next, chicks were given a choice between the familiar and the unfamiliar pattern, present in either the multimodal, visual or acoustic modality. The responses to the novel stimuli were affected by their imprinting experience, and the effect was stronger for chicks imprinted with multimodal patterns than for the other groups. Interestingly, males and females adopted a different strategy, with males more attracted by unfamiliar patterns and females more attracted by familiar patterns. Our data show that chicks can generalize abstract patterns by mere exposure through filial imprinting and that multimodal stimulation is more effective than unimodal stimulation for pattern learning.

  13. Insulin and branched-chain amino acid depletion during mouse preimplantation embryo culture programmes body weight gain and raised blood pressure during early postnatal life.

    PubMed

    Velazquez, Miguel A; Sheth, Bhavwanti; Smith, Stephanie J; Eckert, Judith J; Osmond, Clive; Fleming, Tom P

    2018-02-01

    Mouse maternal low protein diet exclusively during preimplantation development (Emb-LPD) is sufficient to programme altered growth and cardiovascular dysfunction in offspring. Here, we use an in vitro model comprising preimplantation culture in medium depleted in insulin and branched-chain amino acids (BCAA), two proposed embryo programming inductive factors from Emb-LPD studies, to examine the consequences for blastocyst organisation and, after embryo transfer (ET), postnatal disease origin. Two-cell embryos were cultured to blastocyst stage in defined KSOM medium supplemented with four combinations of insulin and BCAA concentrations. Control medium contained serum insulin and uterine luminal fluid amino acid concentrations (including BCAA) found in control mothers from the maternal diet model (N-insulin+N-bcaa). Experimental medium (three groups) contained 50% reduction in insulin and/or BCAA (L-insulin+N-bcaa, N-insulin+L-bcaa, and L-insulin+N-bcaa). Lineage-specific cell numbers of resultant blastocysts were not affected by treatment. Following ET, a combined depletion of insulin and BCAA during embryo culture induced a non sex-specific increase in birth weight and weight gain during early postnatal life. Furthermore, male offspring displayed relative hypertension and female offspring reduced heart/body weight, both characteristics of Emb-LPD offspring. Combined depletion of metabolites also resulted in a strong positive correlation between body weight and glucose metabolism that was absent in the control group. Our results support the notion that composition of preimplantation culture medium can programme development and associate with disease origin affecting postnatal growth and cardiovascular phenotypes and implicate two important nutritional mediators in the inductive mechanism. Our data also have implications for human assisted reproductive treatment (ART) practice. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  14. Influence of pre-storage incubation on hatchability traits, thyroid hormones, antioxidative status and immunity of newly hatched chicks at two chicken breeder flock ages.

    PubMed

    Ebeid, T A; Twfeek, F A; Assar, M H; Bealish, A M; Abd El-Karim, R E; Ragab, M

    2017-11-01

    Egg storage longer than 7 days is associated with negative effects on hatchability traits. Pre-storage incubation has been a suggested method to reduce the negative effects of long-term storage times by enhancing the developmental stage of the embryo and probably reducing the embryonic stress. The objective of the present study was to investigate the effects of pre-storage incubation and storage time on hatchability characteristics, chick quality and serum thyroid hormones, antioxidative properties and immunoglobulin Y (IgY) concentrations of newly hatched chicks at two breeder flock ages. A total of 8000 fertile eggs were obtained from two different ages of chicken breeder hens (Egyptian local cross, Inshas). Half of the eggs were collected from young breeder hens (28 weeks old) and the other half from old breeder hens (50 weeks old). In each breeder flock age, eggs were distributed in a completely randomized experimental design in a 2×4 factorial arrangement, with two storage periods (4 or 14 days) and four pre-storage incubation durations (0, 4, 6 or 8 h at 37.5°C). At 28 and 50 weeks of age, pre-storage incubation and its interaction with storage period influenced significantly the apparent fertility, hatchability of set eggs and hatchability of fertile eggs and this improvement in hatchability is attributed to the reduction in embryonic mortality (early, intermediate and late). Pre-storage incubation for 6 or 8 h elevated significantly the grade A chicks and reduced the grade B chicks in comparison with non-heated controls. Interestingly, for eggs stored for 14 days, pre-storage incubation for 6 or 8 h enhanced serum triiodothyronine, thyroxine, glutathione peroxidase activity, total antioxidant capacity and IgY concentrations significantly and decreased serum malondialdehyde concentration significantly in the newly hatched chicks. It could be concluded that pre-storage incubation enhanced the hatching results, improved the antioxidative properties, reduced

  15. Mouse Embryo Compaction.

    PubMed

    White, M D; Bissiere, S; Alvarez, Y D; Plachta, N

    2016-01-01

    Compaction is a critical first morphological event in the preimplantation development of the mammalian embryo. Characterized by the transformation of the embryo from a loose cluster of spherical cells into a tightly packed mass, compaction is a key step in the establishment of the first tissue-like structures of the embryo. Although early investigation of the mechanisms driving compaction implicated changes in cell-cell adhesion, recent work has identified essential roles for cortical tension and a compaction-specific class of filopodia. During the transition from 8 to 16 cells, as the embryo is compacting, it must also make fundamental decisions regarding cell position, polarity, and fate. Understanding how these and other processes are integrated with compaction requires further investigation. Emerging imaging-based techniques that enable quantitative analysis from the level of cell-cell interactions down to the level of individual regulatory molecules will provide a greater understanding of how compaction shapes the early mammalian embryo. © 2016 Elsevier Inc. All rights reserved.

  16. Immunolocalization and expression of Na(+)/K(+) -ATPase in embryos, early larval stages and adults of the freshwater shrimp Palaemonetes argentinus (Decapoda, Caridea, Palaemonidae).

    PubMed

    Ituarte, Romina Belén; Lignot, Jehan-Hervé; Charmantier, Guy; Spivak, Eduardo; Lorin-Nebel, Catherine

    2016-06-01

    The euryhaline shrimp Palaemonetes argentinus exemplifies an evolutionary transition from brackish to freshwater habitats that requires adequate osmoregulatory capacities. Hyperosmoregulation is functional at hatching and it likely begins during the embryonic phase allowing this species to develop entirely in fresh water. Here, we investigated the Na(+)/K(+)-ATPase α-subunit gene (nka-α) expression using quantitative real-time PCR and localized Na(+)/K(+)-ATPase (NKA) in ion-transporting epithelia through immunofluorescence microscopy. We reared shrimps from spawning to juvenile stages at two salinities (1, 15 ‰) and maintained adults for 3 weeks at three salinity treatments (1, 15, 25 ‰). nka-α gene expression was measured in: (1) embryos at an early (SI), intermediate (SII) and late (SIII) stage of embryonic development; (2) newly hatched larvae (Zoea I, ZI); and (3) isolated gill tissue of adults. The nka-α expression was low in SI and SII embryos and reached maximum levels prior to hatching (SIII), which were similar to expression levels detected in the ZI. The nka-α expression in SIII and ZI was highest at 15 ‰, whereas salinity did not affect expression in earlier embryos. In SIII, in ZI and in a later zoeal stage ZIV, NKA was localized in epithelial cells of pleurae, in the inner-side epithelium of branchiostegite and in the antennal glands. Gills appeared in the ZIV but NKA immunolabeling of the cells of the gill shaft occurred in a subsequent developmental larval stage, the decapodid. Extrabranchial organs constitute the main site of osmoregulation in early ontogenetic stages of this freshwater shrimp.

  17. Nodal Cilia Dynamics and the Specification of the Left/Right Axis in Early Vertebrate Embryo Development

    PubMed Central

    Buceta, Javier; Ibañes, Marta; Rasskin-Gutman, Diego; Okada, Yasushi; Hirokawa, Nobutaka; Izpisúa-Belmonte, Juan Carlos

    2005-01-01

    Nodal cilia dynamics is a key factor for left/right axis determination in mouse embryos through the induction of a leftward fluid flow. So far it has not been clearly established how such dynamics is able to induce the asymmetric leftward flow within the node. Herein we propose that an asymmetric two-phase nonplanar beating cilia dynamics that involves the bending of the ciliar axoneme is responsible for the leftward fluid flow. We support our proposal with a host of hydrodynamic arguments, in silico experiments and in vivo video microscopy data in wild-type embryos and inv mutants. Our phenomenological modeling approach underscores how the asymmetry and speed of the flow depends on different relevant parameters. In addition, we discuss how the combination of internal and external mechanisms might cause the two-phase beating cilia dynamics. PMID:16040754

  18. Tissue distribution of cells derived from the area opaca in heterospecific quail-chick blastodermal chimeras

    PubMed Central

    Karagenç, Levent; Sandikci, Mustafa

    2010-01-01

    The objective of the current study was to determine the tissue distribution of cells derived from the area opaca in heterospecific quail-chick blastodermal chimeras. Quail-chick chimeras were constructed by transferring dissociated cells from the area opaca of the stage X–XII (EG&K) quail embryo into the subgerminal cavity of the unincubated chick blastoderm. The distribution of quail cells in embryonic as well as extra-embryonic tissues of the recipient embryo were examined using the QCPN monoclonal antibody after 6 days of incubation in serial sections taken at 100-μm intervals. Data gathered in the present study demonstrated that, when introduced into the subgerminal cavity of a recipient embryo, cells of the area opaca are able to populate not only extra-embryonic structures such as the amnion and the yolk sac, but also various embryonic tissues derived from the ectoderm and less frequently the mesoderm. Ectodermal chimerism was confined mainly to the head region and was observed in tissues derived from the neural ectoderm and the surface ectoderm, including the optic cup, diencephalon and lens. Although the possibility of random incorporation of transplanted cells into these embryonic structures cannot be excluded, these results would suggest that area opaca, a peripheral ring of cells in the avian embryo destined to form the extra-embryonic ectoderm and endoderm of the yolk sac, might harbor cells that have the potential to give rise to various cell types in the recipient chick embryo, including those derived from the surface ectoderm and neural ectoderm. PMID:19900180

  19. Comparison of nicotinic receptor binding and biotransformation of coniine in the rat and chick.

    PubMed

    Forsyth, C S; Speth, R C; Wecker, L; Galey, F D; Frank, A A

    1996-12-31

    Coniine, an alkaloid from Conium maculatum (poison hemlock), is a known teratogen in many domestic species with maternal ingestion resulting in arthrogryposis of the offspring. We have previously shown that rats are not susceptible and rabbits only weakly susceptible to coniine-induced arthrogryposis. However, the chick embryo does provide a reproducible laboratory animal model of coniine-induced teratogenesis. The reason for this cross-species variation is unknown. The purpose of this study was to evaluate coniine binding to nicotinic receptors and to measure coniine metabolism in vitro between susceptible and non-susceptible species. Using the chick model, neither the peripheral nicotinic receptor antagonist d-tubocurarine chloride nor the central nicotinic receptor antagonist trimethaphan camsylate blocked the teratogenesis or lethality of 1.5% coniine (50 microliters/egg). Trimethaphan camsylate enhanced coniine-induced lethality in a dose-dependent manner. Neither nicotinic receptor blocker prevented nicotine sulfate-induced malformations but d-tubocurarine chloride did block lethality in a dose-dependent manner. Competition by coniine for [125I]-alpha-bungarotoxin to nicotinic receptors isolated from adult rat diaphragm and chick thigh muscle and competition by coniine for [3H]-cytisine to receptors from rat and chick brain were used to assess coniine binding to nicotinic receptors. The IC50 for coniine in rat diaphragm was 314 microM while that for chick leg muscle was 70 microM. For neuronal nicotinic receptors, the IC50s of coniine for maternal rat brain, fetal rat brain, and chick brain were 1100 microM, 820 microM, and 270 microM, respectively. There were no differences in coniine biotransformation in vitro by microsomes from rat or chick livers. Differences in apparent affinity of coniine for nicotinic receptors or differences in the quantity of the nicotinic receptor between the rat and chick may explain, in part, the differences in susceptibility of

  20. Stronger net posterior cortical forces and asymmetric microtubule arrays produce simultaneous centration and rotation of the pronuclear complex in the early Caenorhabditis elegans embryo

    PubMed Central

    Coffman, Valerie C.; McDermott, Matthew B. A.; Shtylla, Blerta; Dawes, Adriana T.

    2016-01-01

    Positioning of microtubule-organizing centers (MTOCs) incorporates biochemical and mechanical cues for proper alignment of the mitotic spindle and cell division site. Current experimental and theoretical studies in the early Caenorhabditis elegans embryo assume remarkable changes in the origin and polarity of forces acting on the MTOCs. These changes must occur over a few minutes, between initial centration and rotation of the pronuclear complex and entry into mitosis, and the models do not replicate in vivo timing of centration and rotation. Here we propose a model that incorporates asymmetry in the microtubule arrays generated by each MTOC, which we demonstrate with in vivo measurements, and a similar asymmetric force profile to that required for posterior-directed spindle displacement during mitosis. We find that these asymmetries are capable of and important for recapitulating the simultaneous centration and rotation of the pronuclear complex observed in vivo. The combination of theoretical and experimental evidence provided here offers a unified framework for the spatial organization and forces needed for pronuclear centration, rotation, and spindle displacement in the early C. elegans embryo. PMID:27733624

  1. Intracellular and Extracellular pH and Ca Are Bound to Control Mitosis in the Early Sea Urchin Embryo via ERK and MPF Activities

    PubMed Central

    Ciapa, Brigitte; Philippe, Laetitia

    2013-01-01

    Studies aiming to predict the impact on marine life of ocean acidification and of altered salinity have shown altered development in various species including sea urchins. We have analyzed how external Na, Ca, pH and bicarbonate control the first mitotic divisions of sea urchin embryos. Intracellular free Ca (Cai) and pH (pHi) and the activities of the MAP kinase ERK and of MPF regulate mitosis in various types of cells including oocytes and early embryos. We found that intracellular acidification of fertilized eggs by Na-acetate induces a huge activation of ERK at time of mitosis. This also stops the cell cycle and leads to cell death, which can be bypassed by treatment with the MEK inhibitor U0126. Similar intracellular acidification induced in external medium containing low sodium or 5-(N-Methyl-N-isobutyl) amiloride, an inhibitor of the Na+/H+ exchanger, also stops the cell cycle and leads to cell death. In that case, an increase in Cai and in the phosphorylation of tyr-cdc2 occurs during mitosis, modifications that depend on external Ca. Our results indicate that the levels of pHi and Cai determine accurate levels of Ptyr-Cdc2 and P-ERK capable of ensuring progression through the first mitotic cycles. These intracellular parameters rely on external Ca, Na and bicarbonate, alterations of which during climate changes could act synergistically to perturb the early marine life. PMID:23785474

  2. Intracellular and extracellular pH and Ca are bound to control mitosis in the early sea urchin embryo via ERK and MPF activities.

    PubMed

    Ciapa, Brigitte; Philippe, Laetitia

    2013-01-01

    Studies aiming to predict the impact on marine life of ocean acidification and of altered salinity have shown altered development in various species including sea urchins. We have analyzed how external Na, Ca, pH and bicarbonate control the first mitotic divisions of sea urchin embryos. Intracellular free Ca (Cai) and pH (pHi) and the activities of the MAP kinase ERK and of MPF regulate mitosis in various types of cells including oocytes and early embryos. We found that intracellular acidification of fertilized eggs by Na-acetate induces a huge activation of ERK at time of mitosis. This also stops the cell cycle and leads to cell death, which can be bypassed by treatment with the MEK inhibitor U0126. Similar intracellular acidification induced in external medium containing low sodium or 5-(N-Methyl-N-isobutyl) amiloride, an inhibitor of the Na(+)/H(+) exchanger, also stops the cell cycle and leads to cell death. In that case, an increase in Cai and in the phosphorylation of tyr-cdc2 occurs during mitosis, modifications that depend on external Ca. Our results indicate that the levels of pHi and Cai determine accurate levels of Ptyr-Cdc2 and P-ERK capable of ensuring progression through the first mitotic cycles. These intracellular parameters rely on external Ca, Na and bicarbonate, alterations of which during climate changes could act synergistically to perturb the early marine life.

  3. Characterizing early embryonic development of Brown Tsaiya Ducks (Anas platyrhynchos) in comparison with Taiwan Country Chicken (Gallus gallus domestics)

    PubMed Central

    Lumsangkul, Chompunut; Fan, Yang-Kwang; Chang, Shen-Chang; Ju, Jyh-Cherng

    2018-01-01

    Avian embryos are among the most convenient and the primary representatives for the study of classical embryology. It is well-known that the hatching time of duck embryos is approximately one week longer than that of chicken embryos. However, the key features associated with the slower embryonic development in ducks have not been adequately described. This study aimed to characterize the pattern and the speed of early embryogenesis in Brown Tsaiya Ducks (BTD) compared with those in Taiwan Country Chicken (TCC) by using growth parameters including embryonic crown-tail length (ECTL), primitive streak formation, somitogenesis, and other development-related parameters, during the first 72 h of incubation. Three hundred and sixty eggs from BTD and TCC, respectively, were incubated at 37.2°C, and were then dissected hourly to evaluate their developmental stages. We found that morphological changes of TCC embryos shared a major similarity with that of the Hamburger and Hamilton staging system during early chick embryogenesis. The initial primitive streak in TCC emerged between 6 and 7 h post-incubation, but its emergence was delayed until 10 to 13 h post-incubation in BTD. Similarly, the limb primordia (wing and limb buds) were observed at 51 h post-incubation in TCC embryos compared to 64 h post-incubation in BTD embryos. The allantois first appeared around 65 to 68 h in TCC embryos, but it was not observed in BTD embryos. At the 72 h post-incubation, 40 somites were clearly formed in TCC embryos while only 32 somites in BTD embryos. Overall, the BTD embryos developed approximately 16 h slower than the chicken embryo during the first 72 h of development. To our best knowledge, this is the first study to describe two distinct developmental time courses between TCC and BTD, which would facilitate future embryogenesis-related studies of the two important avian species in Taiwan. PMID:29742160

  4. Naïve Chicks Prefer Hollow Objects

    PubMed Central

    Schill, Jana; Nencini, Andrea Maria; Vallortigara, Giorgio

    2016-01-01

    Biological predispositions influence approach and avoid responses from the time of birth or hatching. Neonates of species that require parental care (e.g. human babies and chicks of the domestic fowl) are attracted by stimuli associated with animate social partners, such as face-like configurations, biological motion and self-propulsion. The property of being filled is used as a cue of animacy by 8-month-old human infants but it is not known whether this reflects the effect of previous experience. We used chicks of the domestic fowl (Gallus gallus) to investigate whether the property of being filled vs. hollow elicits spontaneous or learned preferences. To this aim we tested preferences of naïve and imprinted chicks for hollow and closed cylinders. Contrary to our expectations, we documented an unlearned attraction for hollow stimuli. The preference for hollow stimuli decreased when chicks were imprinted on filled stimuli but did not increase when chicks were imprinted on hollow stimuli, suggesting that hollowness is not crucial to determine affiliative responses for imprinting objects. When chicks were imprinted on occluded stimuli that could be either filled or hollow, the preference for hollow stimuli emerged again, showing that imprinting does not disrupt the spontaneous preference for hollow objects. Further experiments revealed that hollow objects were mainly attractive by means of depth cues such as darker innards, more than as places to hide or as objects with high contrast. Our findings point to predisposed preferences for hollow objects, and suggest that early predispositions might be driven by factors different from animacy cues. PMID:27851773

  5. Prospective study of automated versus manual annotation of early time-lapse markers in the human preimplantation embryo.

    PubMed

    Kaser, Daniel J; Farland, Leslie V; Missmer, Stacey A; Racowsky, Catherine

    2017-08-01

    How does automated time-lapse annotation (Eeva™) compare to manual annotation of the same video images performed by embryologists certified in measuring durations of the 2-cell (P2; time to the 3-cell minus time to the 2-cell, or t3-t2) and 3-cell (P3; time to 4-cell minus time to the 3-cell, or t4-t3) stages? Manual annotation was superior to the automated annotation provided by Eeva™ version 2.2, because manual annotation assigned a rating to a higher proportion of embryos and yielded a greater sensitivity for blastocyst prediction than automated annotation. While use of the Eeva™ test has been shown to improve an embryologist's ability to predict blastocyst formation compared to Day 3 morphology alone, the accuracy of the automated image analysis employed by the Eeva™ system has never been compared to manual annotation of the same time-lapse markers by a trained embryologist. We conducted a prospective cohort study of embryos (n = 1477) cultured in the Eeva™ system (n = 8 microscopes) at our institution from August 2014 to February 2016. Embryos were assigned a blastocyst prediction rating of High (H), Medium (M), Low (L), or Not Rated (NR) by Eeva™ version 2.2 according to P2 and P3. An embryologist from a team of 10, then manually annotated each embryo and if the automated and manual ratings differed, a second embryologist independently annotated the embryo. If both embryologists disagreed with the automated Eeva™ rating, then the rating was classified as discordant. If the second embryologist agreed with the automated Eeva™ score, the rating was not considered discordant. Spearman's correlation (ρ), weighted kappa statistics and the intra-class correlation (ICC) coefficients with 95% confidence intervals (CI) between Eeva™ and manual annotation were calculated, as were the proportions of discordant embryos, and the sensitivity, specificity, positive predictive value (PPV) and NPV of each method for blastocyst prediction. The distribution of

  6. Early morphological nuclear events and developmental capacity of embryos reconstructed with fetal fibroblasts at the M or G1 phase after intracytoplasmic nuclear injection in cattle.

    PubMed

    Ideta, Atsushi; Urakawa, Manami; Aoyagi, Yoshito; Saeki, Kazuhiro

    2005-04-01

    We examined morphological nuclear events during the first cell cycle of bovine embryos reconstructed with somatic cells at the M and G1 phases (M-embryos and G1-embryos, respectively) by intracytoplasmic nuclear injection, and the subsequent development of these embryos in vitro and in vivo. Bovine fetal fibroblasts (BFFs) at the M or G1 phase were directly injected into enucleated oocytes, and activated immediately. Only half (48%) of the M-embryos extruded polar body-like cells (PBCs) at 6 h post injection (hpi). At 15 to 19 hpi, 54% of the M-embryos formed a single pronucleus-like nucleus. Nuclear envelope-breakdown, premature chromosome condensation and single nuclear clusters were observed in most of the G1-embryos (88%) within 30 min following the nuclear injection. At 15 to 19 hpi, single pronucleus-like nuclei were formed in most G1-embryos (83%). The potential of G1-embryos to develop to blastocysts was significantly higher than that of M-embryos (31% vs 16%). Three of five recipients following transfer of blastocysts derived from the G1-embryos became pregnant on Day 30, and one recipient delivered a calf. Our results indicate that almost a half of the M-embryos failed to extrude PBCs and that the G1-embryos developed to blastocysts at a higher rate than the M-embryos.

  7. The avian embryo responding to microgravity of space flight

    NASA Technical Reports Server (NTRS)

    Hullinger, Ronald L.

    1993-01-01

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

  8. Toxicity of glyphosate as Glypro and LI700 to red-eared slider (trachemys scripta elegans) embryos and early hatchlings.

    PubMed

    Sparling, Donald W; Matson, Cole; Bickham, John; Doelling-Brown, Paige

    2006-10-01

    More than 8.2 billion ha of cropland, gardens, and forests are treated with the herbicide glyphosate each year. Whereas the toxicity of glyphosate and associated adjuvants has been measured in other vertebrates, few, if any, studies have looked at their effects in reptiles. In some instances, management of turtle habitat requires control of successional stages through application of herbicides. Adults and juvenile turtles may be exposed directly, whereas embryos may contact the chemicals through the soil. In the present study, we exposed eggs of red-eared sliders (Trachemys scripta elegans) to single applications of herbicide ranging from 0 to 11,206 ppm wet weight of glyphosate in Glypro and 0 to 678 ppm of the surfactant, LI700. Hatching success at the highest concentration was significantly lower (73%) than in other treatments (80-100%). At hatch, turtles at the highest concentration weighed less than those at other concentrations. During a 14-d holding period, we observed dose-response relationships in the ability of hatchlings to right themselves when turned on their backs. At the end of the holding period, hatchlings at the highest dose level were still lighter, and somatic indices were lower, than those in other treatments. Genetic damage, as measured by flow cytometry, increased with treatment concentration except for the highest dose. We conclude that because of the high concentrations needed to produce effects and the protection offered by several centimeters of soil or sediment, glyphosate with LI700 poses low levels of risk to red-eared slider embryos under normal field operations with regards to the endpoints measured in the present study. Carelessness in handling glyphosate or failure to follow label directions may produce adverse effects. There also is a risk that the health of turtle embryos may be affected in ways not measured in the present study.

  9. Subtractive and differential hybridization molecular analyses of Ceratitis capitata XX/XY versus XX embryos to search for male-specific early transcribed genes.

    PubMed

    Salvemini, Marco; D'Amato, Rocco; Petrella, Valeria; Ippolito, Domenica; Ventre, Giuseppe; Zhang, Ying; Saccone, Giuseppe

    2014-01-01

    The agricultural pest Ceratitis capitata, also known as the Mediterranean fruit fly or Medfly, is a fruit crop pest of very high economic relevance in different continents. The strategy to separate Ceratitis males from females (sexing) in mass rearing facilities is a useful step before the sterilization and release of male-only flies in Sterile Insect Technique control programs (SIT). The identification of genes having early embryonic male-specific expression, including Y-linked genes, such as the Maleness factor, could help to design novel and improved methods of sexing in combination with transgenesis, aiming to confer conditional female-specific lethality or female-to-male sexual reversal. We used a combination of Suppression Subtractive Hybrydization (SSH), Mirror Orientation Selection (MOS) anddifferential screening hybridization (DSH) techniques to approach the problem of isolating corresponding mRNAs expressed in XX/XY embryos versus XX-only embryos during a narrow developmental window (8-10 hours after egg laying, AEL ). Here we describe a novel strategy we have conceived to obtain relatively large amounts of XX-only embryos staged at 8-10 h AEL and so to extract few micrograms of polyA+ required to apply the complex technical procedure. The combination of these 3 techniques led to the identification of a Y-linked putative gene, CcGm2, sharing high sequence identity to a paralogous gene, CcGm1, localized either on an autosome or on the X chromosome. We propose that CcGm2 is a first interesting putative Y-linked gene which could play a role in sex determination. The function exterted by this gene should be investigated by novel genetic tools, such as CRISPR-CAS9, which will permit to target only the Y-linked paralogue, avoiding to interfere with the autosomal or X-linked paralogue function.

  10. Subtractive and differential hybridization molecular analyses of Ceratitis capitata XX/XY versus XX embryos to search for male-specific early transcribed genes

    PubMed Central

    2014-01-01

    The agricultural pest Ceratitis capitata, also known as the Mediterranean fruit fly or Medfly, is a fruit crop pest of very high economic relevance in different continents. The strategy to separate Ceratitis males from females (sexing) in mass rearing facilities is a useful step before the sterilization and release of male-only flies in Sterile Insect Technique control programs (SIT). The identification of genes having early embryonic male-specific expression, including Y-linked genes, such as the Maleness factor, could help to design novel and improved methods of sexing in combination with transgenesis, aiming to confer conditional female-specific lethality or female-to-male sexual reversal. We used a combination of Suppression Subtractive Hybrydization (SSH), Mirror Orientation Selection (MOS) and differential screening hybridization (DSH) techniques to approach the problem of isolating corresponding mRNAs expressed in XX/XY embryos versus XX-only embryos during a narrow developmental window (8-10 hours after egg laying, AEL ). Here we describe a novel strategy we have conceived to obtain relatively large amounts of XX-only embryos staged at 8-10 h AEL and so to extract few micrograms of polyA+ required to apply the complex technical procedure. The combination of these 3 techniques led to the identification of a Y-linked putative gene, CcGm2, sharing high sequence identity to a paralogous gene, CcGm1, localized either on an autosome or on the X chromosome. We propose that CcGm2 is a first interesting putative Y-linked gene which could play a role in sex determination. The function exterted by this gene should be investigated by novel genetic tools, such as CRISPR-CAS9, which will permit to target only the Y-linked paralogue, avoiding to interfere with the autosomal or X-linked paralogue function. PMID:25472628

  11. Mouse Mix gene is activated early during differentiation of ES and F9 stem cells and induces endoderm in frog embryos.

    PubMed

    Mohn, Deanna; Chen, Siming W; Dias, Dora Campos; Weinstein, Daniel C; Dyer, Michael A; Sahr, Kenneth; Ducker, Charles E; Zahradka, Elizabeth; Keller, Gordon; Zaret, Kenneth S; Gudas, Lorraine J; Baron, Margaret H

    2003-03-01

    In frog and zebrafish, the Mix/Bix family of paired type homeodomain proteins play key roles in specification and differentiation of mesendoderm. However, in mouse, only a single Mix gene (mMix) has been identified to date and its function is unknown. We have analyzed the expression of mouse Mix RNA and protein in embryos, embryoid bodies formed from embryonic stem cells and F9 teratocarcinoma cells, as well as several differentiated cell types. Expression in embryoid bodies in culture mirrors that in embryos, where Mix is transcribed transiently in primitive (visceral) endoderm (VE) and in nascent mesoderm. In F9 cells induced by retinoic acid to differentiate to VE, mMix is coordinately expressed with three other endodermal transcription factors, well before AFP, and its protein product is localized to the nucleus. In a subpopulation of nascent mesodermal cells from embryonic stem cell embryoid bodies, mMix is coexpressed with Brachyury. Intriguingly, mMix mRNA is detected in a population (T+Flk1+) of cells which may contain hemangioblasts, before the onset of hematopoiesis and activation of hematopoietic markers. In vitro and in vivo, mMix expression in nascent mesoderm is rapidly down-regulated and becomes undetectable in differentiated cell types. In the region of the developing gut, mMix expression is confined to the mesoderm of mid- and hindgut but is absent from definitive endoderm. Injection of mouse mMix RNA into early frog embryos results in axial truncation of developing tadpoles and, in animal cap assays, mMix alone is sufficient to activate expression of several endodermal (but not mesodermal) markers. Although these observations do not exclude a possible cell-autonomous function for mMix in mesendodermal progenitor cells, they do suggest an additional, non-cell autonomous role in nascent mesoderm in the formation and/or patterning of adjacent definitive endoderm. Copyright 2003 Wiley-Liss, Inc.

  12. Polarity of the ascidian egg cortex and relocalization of cER and mRNAs in the early embryo.

    PubMed

    Prodon, François; Dru, Philippe; Roegiers, Fabrice; Sardet, Christian

    2005-06-01

    The mature ascidian oocyte is a large cell containing cytoplasmic and cortical domains polarized along a primary animal-vegetal (a-v) axis. The oocyte cortex is characterized by a gradient distribution of a submembrane monolayer of cortical rough endoplasmic reticulum (cER) and associated maternal postplasmic/PEM mRNAs (cER-mRNA domain). Between fertilization and first cleavage, this cER-mRNA domain is first concentrated vegetally and then relocated towards the posterior pole via microfilament-driven cortical contractions and spermaster-microtubule-driven translocations. The cER-mRNA domain further concentrates in a macroscopic cortical structure called the centrosome attracting body (CAB), which mediates a series of asymmetric divisions starting at the eight-cell stage. This results in the segregation of determinant mRNAs and their products in posterior cells of the embryo precursors of the muscle and germ line. Using two species of ascidians (Ciona intestinalis and Phallusia mammillata), we have pursued and amplified the work initiated in Halocynthia roretzi. We have analysed the cortical reorganizations in whole cells and in cortical fragments isolated from oocytes and from synchronously developing zygotes and embryos. After fertilization, we observe that a cortical patch rich in microfilaments encircles the cER-mRNA domain, concentrated into a cortical cap at the vegetal/contraction pole (indicating the future dorsal pole). Isolated cortices also retain microtubule asters rich in cER (indicating the future posterior pole). Before mitosis, parts of the cER-mRNA domain are detected, together with short microtubules, in isolated posterior (but not anterior) cortices. At the eight-cell stage, the posteriorly located cER-mRNA domain undergoes a cell-cycle-dependant compaction into the CAB. The CAB with embedded centrosomal microtubules can be isolated with cortical fragments from eight-cell-stage embryos. These and previous observations indicate that cytoskeleton

  13. Ovarian adipocytokines are associated with early in vitro human embryo development independent of the action of ovarian insulin.

    PubMed

    Li, Liyun; Ferin, Michel; Sauer, Mark V; Lobo, Roger A

    2012-12-01

    We aimed to characterize the association between levels of serum and follicular fluid (FF) adipocytokines, reflected by the leptin to adiponectin ratio (L:A ratio), and oocyte quality and in vitro embryo development in women undergoing assisted reproduction. We also aimed to assess whether follicular hormonal pathways mediate this interaction. We prospectively collected FF from up to four individual preovulatory follicles (n = 76) and fasting sera from women (n = 31) without endocrinopathies undergoing in vitro fertilization (IVF) at a university-based center for assisted reproduction. Leptin, total adiponectin, insulin, insulin-like growth factor 1 (IGF-1), and ovarian steriods were measured using enzyme immunoassay. Oocyte maturity, fertilization, and embryo development were assessed. FF leptin was similar to serum levels while FF adiponectin was lower. FF leptin (27.10 ± 4.05 ng/mL) and the L:A ratio (11.48E-3 ± 2.57E-3) were related to FF insulin (R (2) = 0.370 and 0.419, p < 0.001) but not to ovarian steroids or IGF-1, whereas FF adiponectin ( 4.22 ± 0.52 ug/mL) correlated only with leptin (R (2) = -0.138, p = 0.001). Oocytes from a high FF L:A ratio environment were 81 % (RR 1.81 [95%CI 0.97-3.37]) more likely to undergo successful cleavage and 117 % (RR 2.17 [95 % CI 1.06-4.44]) more likely to obtain viable cleavage morphology compared to a low FF L:A ratio environment, even when adjusted for FF insulin, an independent predictor of cleavage. Certain adipocytokines, particularly the L:A ratio in the FF of the preovulatory follicle, are related to successful in vitro embryo development. This action may be independent of FF insulin.

  14. Convergence of topological domain boundaries, insulators, and polytene interbands revealed by high-resolution mapping of chromatin contacts in the early Drosophila melanogaster embryo

    PubMed Central

    Stadler, Michael R; Haines, Jenna E

    2017-01-01

    High-throughput assays of three-dimensional interactions of chromosomes have shed considerable light on the structure of animal chromatin. Despite this progress, the precise physical nature of observed structures and the forces that govern their establishment remain poorly understood. Here we present high resolution Hi-C data from early Drosophila embryos. We demonstrate that boundaries between topological domains of various sizes map to DNA elements that resemble classical insulator elements: short genomic regions sensitive to DNase digestion that are strongly bound by known insulator proteins and are frequently located between divergent promoters. Further, we show a striking correspondence between these elements and the locations of mapped polytene interband regions. We believe it is likely this relationship between insulators, topological boundaries, and polytene interbands extends across the genome, and we therefore propose a model in which decompaction of boundary-insulator-interband regions drives the organization of interphase chromosomes by creating stable physical separation between adjacent domains. PMID:29148971

  15. Normal development of spinal axons in early embryo stages and posterior locomotor function is independent of GAL-1.

    PubMed

    Pasquini, Juana M; Barrantes, Francisco J; Quintá, Héctor R

    2017-09-01

    It was recently described that Galectin-1 (Gal-1) promotes axonal growth after spinal cord injury. This effect depends on protein dimerization, since monomeric Gal-1 fails to stimulate axonal re-growth. Gal-1 is expressed in vivo at concentrations that favor the monomeric species. The aim of the present study is to investigate whether endogenous Gal-1 is required for spinal axon development and normal locomotor behavior in mice. In order to characterize axonal development, we used a novel combination of 3-DISCO technique with 1-photon microscopy and epifluorescence microscopy under high power LED illumination, followed by serial image section deconvolution and 3-D reconstruction. Cleared whole lgals-1 -/- embryos were used to analyze the 3-D cytoarchitecture of motor, commissural, and sensory axons. This approach allowed us to evaluate axonal development, including the number of fibers, fluorescence density of the fiber tracts, fiber length as well as the morphology of axonal sprouting, deep within the tissue. Gal-1 deficient embryos did not show morphological/anatomical alterations in any of the axonal populations and parameters analyzed. In addition, specific guidance receptor PlexinA4 did not change its axonal localization in the absence of Gal-1. Finally, Gal-1 deficiency did not change normal locomotor activity in post-natal animals. Taken together, our results show that development of spinal axons as well as the locomotor abilities observed in adult mice are independent of Gal-1. Supporting our previous observations, the present study further validates the use of lgals-1 -/- mice to develop spinal cord- or traumatic brain injury models for the evaluation of the regenerative action of Gal-1. © 2017 Wiley Periodicals, Inc.

  16. Early embryonic expression of FGF4/6/9 gene and its role in the induction of mesenchyme and notochord in Ciona savignyi embryos.

    PubMed

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

    2002-04-01

    In early Ciona savignyi embryos, nuclear localization of beta-catenin is the first step of endodermal cell specification, and triggers the activation of various target genes. A cDNA for Cs-FGF4/6/9, a gene activated downstream of beta-catenin signaling, was isolated and shown to encode an FGF protein with features of both FGF4/6 and FGF9/20. The early embryonic expression of Cs-FGF4/6/9 was transient and the transcript was seen in endodermal cells at the 16- and 32-cell stages, in notochord and muscle cells at the 64-cell stage, and in nerve cord and muscle cells at the 110-cell stage; the gene was then expressed again in cells of the nervous system after neurulation. When the gene function was suppressed with a specific antisense morpholino oligo, the differentiation of mesenchyme cells was completely blocked, and the fate of presumptive mesenchyme cells appeared to change into that of muscle cells. The inhibition of mesenchyme differentiation was abrogated by coinjection of the morpholino oligo and synthetic Cs-FGF4/6/9 mRNA. Downregulation of beta-catenin nuclear localization resulted in the absence of mesenchyme cell differentiation due to failure of the formation of signal-producing endodermal cells. Injection of synthetic Cs-FGF4/6/9 mRNA in beta-catenin-downregulated embryos evoked mesenchyme cell differentiation. These results strongly suggest that Cs-FGF4/6/9 produced by endodermal cells acts an inductive signal for the differentiation of mesenchyme cells. On the other hand, the role of Cs-FGF4/6/9 in the induction of notochord cells is partial; the initial process of the induction was inhibited by Cs-FGF4/6/9 morpholino oligo, but notochord-specific genes were expressed later to form a partial notochord.

  17. Coumafuryl (Fumarin) toxicity in chicks.

    PubMed

    Munger, L L; Su, J J; Barnes, H J

    1993-01-01

    Coumafuryl (Fumarin) toxicity was diagnosed in chickens less than 1 week of age. Mortality rate was 100%. Necropsy showed crops and gizzards to be full of feed. There was diffuse hemorrhage and unclotted blood in the abdomen and thorax. Histological examination showed congestion and hemorrhage. Chemical analysis of crop and gizzard contents contained approximately 340 ppm coumafuryl. The source of the coumafuryl was found to be the wood-straw mats in the chick boxes.

  18. Penguin chicks benefit from elevated yolk androgen levels under sibling competition.

    PubMed

    Poisbleau, Maud; Müller, Wendt; Carslake, David; Demongin, Laurent; Groothuis, Ton G G; Van Camp, Jeff; Eens, Marcel

    2012-01-01

    Crested penguins (genus Eudyptes) have a peculiar hatching pattern, with the first-laid egg (A-egg) hatching after the second-laid egg (B-egg) and chicks from A-eggs typically having a much lower survival probability. Maternal yolk androgens have been suggested to contribute to the competitive superiority of the B-chick in southern rockhopper penguins Eudyptes chrysocome, given their important role in mediating sibling competition in other species. We therefore increased the yolk androgen levels in freshly-laid eggs and examined the consequences for sibling competition--via effects on embryonic developmental times, chick growth and early survival. We placed one androgen-treated egg and one control egg into each foster nest, matching them for mass, laying date and laying order. The androgen treatment did not significantly affect embryonic developmental times or chick measurements at hatching. However, elevated yolk androgen levels benefitted chick growth in interaction with the number of siblings in a brood. Chicks from androgen-treated eggs had faster growth in the presence of a sibling than chicks from control eggs. Under these circumstances they also had a higher survival probability. Thus maternal androgens appear to reinforce the observed hatching pattern, facilitating brood reduction. This contrasts to most previous studies in other species where yolk androgens have been shown to compensate for the negative consequences of delayed hatching within the brood hierarchy.

  19. [Breeding biology of the whiskered auklet (Aethia pygmaea): incubation, chick growth, and feather ontogenesis].

    PubMed

    Koniukhov, N B; Zubakin, V A; Williams, J; Fischer, J

    2000-01-01

    This is the second publication on the results of studies of the whiskered auklet breeding biology on Buldir Island. In 1993, according to the time of chick hatching the egg-laying period began in late April-early May. The single egg was incubated by both parents, which relieved each other once a day, usually at early night. Among the chicks under observation, the first hatched on June 11; the median of hatching was on June 17. After hatching, the egg shell usually remained in the nest. The chick was continuously brooded by one of parents for the first several days but since the age of 7-9 days, it stayed in the nest by itself during the daytime. Each parent fed the chick once a day, at early night, although occasional feedings were recorded also during daytime and at dawn. A newly hatched chick was covered by typical embryonal (natal) down. The remiges, rectrices, their coverts, and large feathers on pteryla humeralis had a mesoptile stage during their growth (the second down generation). The juvenile plumage consisted of semiplumes of types I and II and contour feathers of type I and, possibly, type II. In fledglings, the ornamental feathers were underdeveloped: the crest was absent and facial plumes only began growing. By reaching 100 g, the chicks left their nests at the age of about 37 days. A longer nesting period than in other Aethia auklets appears to be related to a low feeding rate due to the nocturnal activity in the colony.

  20. Identification, emergence and mobilization of circulating endothelial cells or progenitors in the embryo.

    PubMed

    Pardanaud, Luc; Eichmann, Anne

    2006-07-01

    Using quail-chick parabiosis and QH1 monoclonal antibody analysis, we have identified circulating endothelial cells and/or progenitors in the embryo. These cells were already present early in ontogeny, before the third embryonic day. Under normal conditions, they integrated into most tissues but remained scarce. When experimental angiogenic responses were induced by wounding or grafts onto the chorioallantoic membrane, circulating endothelial cells were rapidly mobilized and selectively integrated sites of neoangiogenesis. Their mobilization was not dependent on the presence of the bone marrow as it was effective before its differentiation. Surprisingly, mobilization was not effective during sprouting angiogenesis following VEGF treatment of chorioallantoic membrane. Thus, embryonic circulating endothelial cells were efficiently mobilized during the establishment of an initial vascular supply to ischemic tissues following wounding or grafting, but were not involved during classical sprouting angiogenesis.

  1. Egg laying characteristics, egg weight, embryo development, hatching weight and post-hatch growth in relation to oviposition time of broiler breeders.

    PubMed

    Akil, R; Zakaria, A H

    2015-05-01

    Two experiments were conducted to determine egg laying characteristics and the effects of oviposition time on egg weight, embryo development and post-hatch growth in broiler breeders. In experiment 1, eggs collected for 3 consecutive days on hourly basis between 06:30 and 17:30h were categorized to early, middle and late oviposition times in the clutch. In experiment 2, eggs were incubated to study embryo development, remaining albumen, liver weight, heart weight and the tibia length of embryos at 12, 14, 16 and 18 days of incubation as well as the body weight of hatchlings and chickens at 7, 21 and 42 days of age in relation to oviposition time. About 76% of nest eggs were laid from 06:30 to 11:30h. A similar pattern was observed in floor eggs. Egg weight decreased (P<0.01) with advanced position in the clutch. Generally, oviposition time had no effect on embryo growth parameters. At hatch, body weight of chicks derived from eggs of late oviposition times was less (P<0.01) than that of chicks from eggs produced earlier in the clutch. From 3-week-old onwards, chickens of early oviposition time sustained heavier (P<0.05) weight than chickens of middle oviposition time whereas chickens of late oviposition time obtained a middle weight. Differences in egg weights, body weight at hatch and post-hatch growth due to time of oviposition suggest that oviposition time together with incubation conditions should be considered for obtaining greater uniformity and growth of chickens. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. SMK-1/PPH-4.1–mediated silencing of the CHK-1 response to DNA damage in early C. elegans embryos

    PubMed Central

    Kim, Seung-Hwan; Holway, Antonia H.; Wolff, Suzanne; Dillin, Andrew; Michael, W. Matthew

    2007-01-01

    During early embryogenesis in Caenorhabditis elegans, the ATL-1–CHK-1 (ataxia telangiectasia mutated and Rad3 related–Chk1) checkpoint controls the timing of cell division in the future germ line, or P lineage, of the animal. Activation of the CHK-1 pathway by its canonical stimulus DNA damage is actively suppressed in early embryos so that P lineage cell divisions may occur on schedule. We recently found that the rad-2 mutation alleviates this checkpoint silent DNA damage response and, by doing so, causes damage-dependent delays in early embryonic cell cycle progression and subsequent lethality. In this study, we report that mutations in the smk-1 gene cause the rad-2 phenotype. SMK-1 is a regulatory subunit of the PPH-4.1 (protein phosphatase 4) protein phosphatase, and we show that SMK-1 recruits PPH-4.1 to replicating chromatin, where it silences the CHK-1 response to DNA damage. These results identify the SMK-1–PPH-4.1 complex as a critical regulator of the CHK-1 pathway in a developmentally relevant context. PMID:17908915

  3. From the Cover: Exposing Imidacloprid Interferes With Neurogenesis Through Impacting on Chick Neural Tube Cell Survival.

    PubMed

    Liu, Meng; Wang, Guang; Zhang, Shi-Yao; Zhong, Shan; Qi, Guo-Long; Wang, Chao-Jie; Chuai, Manli; Lee, Kenneth Ka Ho; Lu, Da-Xiang; Yang, Xuesong

    2016-09-01

    As a neonicotinoid pesticide, imidacloprid is widely used to control insects in agriculture and fleas on domestic animals. However, it is not known whether imidacloprid exposure negatively affects neurogenesis during embryonic development. In this study, using a chick embryo model, we investigated the effects of imidacloprid exposure on neurogenesis at the earliest stage and during late-stage embryo development. Exposing HH0 chick embryos to imidacloprid in EC culture caused neural tube defects (NTDs) and neuronal differentiation dysplasia as determined by NF/Tuj1 labeling. Furthermore, we found that F-actin accumulation on the apical side of the neural tube was suppressed by exposure to imidacloprid, and the expression of BMP4 and Shh on the dorsal and ventral sides of the neural tubes, respectively, were also reduced, which in turn affects the dorsolateral hinge points during bending of the neural plate. In addition, exposure to imidacloprid reduced cell proliferation and increased cell apoptosis, as determined by pHIS3 labeling and TUNEL staining, respectively, also contributing to the malformation. We obtained similar results in late-stage embryos exposed to imidacloprid. Finally, a bioinformatics analysis was employed to determine which genes identified in this study were involved in NTDs. The experimental evidence and bioinformatics analysis suggested that imidacloprid exposure during chick embryo development could increase the risk of NTDs and neural dysplasia. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  4. Microcystin-LR affects the hypothalamic-pituitary-inter-renal (HPI) axis in early life stages (embryos and larvae) of zebrafish.

    PubMed

    Chen, Liang; Wang, Yeke; Giesy, John P; Chen, Feng; Shi, Ting; Chen, Jun; Xie, Ping

    2018-05-22

    Frequencies and durations of blooms of cyanobacteria are increasing. Some cyanobacteria can produce cyanotoxins including microcystins (MCs). MCs are the most common toxic products of hazardous algal blooms (HABs), with the greatest potential for exposure and to cause toxicity. Recently, MCs have been shown to disrupt endocrine functions. In this study, for the first time, effects of MC-LR on the hypothalamic-pituitary-inter-renal (HPI) axis during early embryonic development (embryos/larvae) of zebrafish (Danio rerio), were investigated. Embryos/larvae of zebrafish were exposed to 1, 10, 100, or 300 μg MC-LR/L during the period of 4-168 h post-fertilization (hpf). Exposure to 300 μg MC-LR/L resulted in significantly greater concentrations of whole-body cortisol than those in controls. Expressions of genes along the HPI axis and mineralocorticoid receptor (MR-) and glucocorticoid receptor (GR-) centered gene networks were evaluated by use of quantitative real-time PCR. Expression of mRNA for crh was significantly down-regulated by exposure to 300 μg MC-LR/L, while expressions of crhbp, crhr1, and crhr2 were significantly up-regulated, relative to controls. MC-LR caused significantly lesser levels of mRNA for steroidogenic genes including hmgra, star, and cyp17, but expression of mRNA for hsd20b was significantly greater than that of controls. Treatment with MC-LR also altered profiles of transcription of MR- and GR-centered gene networks, which might result in multiple responses. Taken together, these results demonstrated that MC-LR affected the corticosteroid-endocrine system of larvae of zebrafish. This study provided valuable insights into molecular mechanisms behind potential toxicity and endocrine disruption of MCs. Copyright © 2018 Elsevier Ltd. All rights reserved.

  5. Critical role of mTOR, PPARγ and PPARδ signaling in regulating early pregnancy decidual function, embryo viability and feto-placental growth.

    PubMed

    Roberti, Sabrina L; Higa, Romina; White, Verónica; Powell, Theresa L; Jansson, Thomas; Jawerbaum, Alicia

    2018-06-01

    What are the consequences of inhibiting mTOR, the mechanistic target of rapamycin (mTOR), and the peroxisome proliferator activated receptor gamma (PPARγ) and PPARδ pathways in the early post-implantation period on decidual function, embryo viability and feto-placental growth in the rat? mTOR inhibition from Days 7 to 9 of pregnancy in rats caused decidual PPARγ and PPARδ upregulation on Day 9 of pregnancy and resulted in embryo resorption by Day 14 of pregnancy. PPARγ and PPARδ inhibition differentially affected decidual mTOR signaling and levels of target proteins relevant to lipid histotrophic nutrition and led to reduced feto-placental weights on Day 14 of pregnancy. Although mTOR, PPARγ and PPARδ are nutrient sensors important during implantation, the role of these signaling pathways in decidual function and how they interact in the early post-implantation period are unknown. Perilipin 2 (PLIN2) and fatty acid binding protein 4 (FABP4), two adipogenic proteins involved in lipid histotrophic nutrition, are targets of mTOR and PPAR signaling pathways in a variety of tissues. Rapamycin (mTOR inhibitor, 0.75 mg/kg, sc), T0070907 (PPARγ inhibitor, 0.001 mg/kg, sc), GSK0660 (PPARδ inhibitor, 0.1 mg/kg, sc) or vehicle was injected daily to pregnant rats from Days 7 to 9 of pregnancy and the studies were performed on Day 9 of pregnancy (n = 7 per group) or Day 14 of pregnancy (n = 7 per group). On Day 9 of pregnancy, rat decidua were collected and prepared for western blot and immunohistochemical studies. On Day 14 of pregnancy, the resorption rate, number of viable fetuses, crown-rump length and placental and decidual weights were determined. Inhibition of mTOR in the early post-implantation period led to a reduction in FABP4 protein levels, an increase in PLIN2 levels and an upregulation of PPARγ and PPARδ in 9-day-pregnant rat decidua. Most embryos were viable on Day 9 of pregnancy but had resorbed by Day 14 of pregnancy. This denotes a key function of

  6. Gene Expression in Pre-MBT Embryos and Activation of Maternally-Inherited Program of Apoptosis to be Executed at around MBT as a Fail-Safe Mechanism in Xenopus Early Embryogenesis

    PubMed Central

    Shiokawa, Koichiro; Aso, Mai; Kondo, Takeshi; Uchiyama, Hiroaki; Kuroyanagi, Shinsaku; Takai, Jun-Ichi; Takahashi, Senji; Kajitani, Masayuki; Kaito, Chikara; Sekimizu, Kazuhisa; Takayama, Eiji; Igarashi, Kazuei; Hara, Hiroshi

    2008-01-01

    S-adenosylmethionine decarboxylase (SAMDC) is an enzyme which converts S-adenosylmethione (SAM), a methyl donor, to decarboxylated SAM (dcSAM), an aminopropyl donor for polyamine biosynthesis. In our studies on gene expression control in Xenopus early embryogenesis, we cloned the mRNA for Xenopus SAMDC, and overexpressed the enzyme by microinjecting its mRNA into Xenopus fertilized eggs. In the mRNA-injected embryos, the level of SAMDC was enormously increased, the SAM was exhausted, and protein synthesis was greatly inhibited, but cellular polyamine content did not change appreciably. SAMDC-overexpressed embryos cleaved and developed normally up to the early blastula stage, but at the midblastula stage, or the stage of midblastula transition (MBT), all the embryos were dissociated into cells, and destroyed due to execution of apoptosis. During cleavage SAMDC-overexpressed embryos transcribed caspase-8 gene, and this was followed by activation of caspase-9. When we overexpressed p53 mRNA in fertilized eggs, similar apoptosis took place at MBT, but in this case, transcription of caspase-8 did not occur, however activation of caspase-9 took place. Apoptosis induced by SAMDC-overexpression was completely suppressed by Bcl-2, whereas apoptosis induced by p53 overexpression or treatments with other toxic agents was only partially rescued. When we injected SAMDC mRNA into only one blastomere of 8- to 32-celled embryos, descendant cells of the mRNA-injected blastomere were segregated into the blastocoel and underwent apoptosis within the blastocoel, although such embryos continued to develop and became tadpoles with various extents of anomaly, reflecting the developmental fate of the eliminated cells. Thus, embryonic cells appear to check themselves at MBT and if physiologically severely-damaged cells occur, they are eliminated from the embryo by activation and execution of the maternally-inherited program of apoptosis. We assume that the apoptosis executed at MBT is a

  7. Monitoring the hatch time of individual chicken embryos.

    PubMed

    Romanini, C E B; Exadaktylos, V; Tong, Q; McGonnel, I; Demmers, T G M; Bergoug, H; Eterradossi, N; Roulston, N; Garain, P; Bahr, C; Berckmans, D

    2013-02-01

    This study investigated variations in eggshell temperature (T(egg)) during the hatching process of broiler eggs. Temperature sensors monitored embryo temperature by registering T(egg) every minute. Measurements carried out on a sample of 40 focal eggs revealed temperature drops between 2 to 6°C during the last 3 d of incubation. Video cameras recorded the hatching process and served as the gold standard reference for manually labeling the hatch times of chicks. Comparison between T(egg) drops and the hatch time of individuals revealed a time synchronization with 99% correlation coefficient and an absolute average time difference up to 25 min. Our findings suggest that attaching temperature sensors to eggshells is a precise tool for monitoring the hatch time of individual chicks. Individual hatch monitoring registers the biological age of chicks and facilitates an accurate and reliable means to count hatching results and manage the hatch window.

  8. Effects of age, vitamin D3, and fructooligosaccharides on bone growth and skeletal integrity of broiler chicks.

    PubMed

    Kim, W K; Bloomfield, S A; Ricke, S C

    2011-11-01

    A study was conducted to evaluate the effects of age, vitamin D(3), and fructooligosaccharides (FOS) on bone mineral density (BMD), bone mineral content (BMC), cortical thickness, cortical and trabecular area, and mechanical properties in broiler chicks using peripheral quantitative computed tomography and mechanical testing. A total of 54 male broiler chicks (1 d old) were placed in battery brooders and fed a corn-soybean starter diet for 7 d. After 7 d, the chicks were randomly assigned to pens of 3 birds each. Each treatment was replicated 3 times. There were 6 treatments: 1) early age control (control 1); 2) control 2; 3) 125 µg/kg of vitamin D(3); 4) 250 µg/kg of vitamin D(3); 5) 2% FOS); and 6) 4% FOS. The control 1 chicks were fed a control broiler diet and killed on d 14 to collect femurs for bone analyses. The remaining groups were killed on d 21. Femurs from 3-wk-old chicks showed greater midshaft cortical BMD, BMC, bone area, thickness, and marrow area than those from 2-wk-old chicks (P = 0.016, 0.0003, 0.0002, 0.01, and 0.0001, respectively). Total, cortical, and trabecular BMD of chick proximal femurs were not influenced by age. However, BMC and bone area were significantly affected by age. The femurs of 2-wk-old chicks exhibited significantly lower stiffness and ultimate load than those of 3-wk-old chicks (P = 0.0001), whereas ultimate stress and elastic modulus of the femurs of 2-wk-old chicks were significantly higher than that of femurs of 3-wk-old chicks (P = 0.0001). Chicks fed 250 µg/kg of vitamin D(3) exhibited significantly greater midshaft cortical BMC (P = 0.04), bone area (P = 0.04), and thickness (P = 0.03) than control 2, 2% FOS, or 4% FOS chicks. In summary, our study suggests that high levels of vitamin D(3) can increase bone growth and mineral deposition in broiler chicks. However, FOS did not have any beneficial effects on bone growth and skeletal integrity. Age is an important factor influencing skeletal integrity and mechanical

  9. Effect of two different commercially available white light LED fixtures on broiler hatchability and chick quality.

    PubMed

    Archer, G S

    2018-06-01

    1. The objective of this study was to evaluate the effect of two different spectra of white light on hatchability and chick quality of broiler chickens. 2. A total of 8424 Ross 708 broiler eggs were divided over three light exposure treatments during incubation: No light (DARK), a light emitting diode (LED) light with high levels of blue light (BLUE), or a LED light with high levels of red light (RED). Hatchability, embryo mortality and chick quality were assessed at hatch. 3. DARK had a higher percentage of chicks with unhealed navels (21.1 ± 1.3%) and fewer overall chicks with no defects (77.0 ± 1.2%) compared to both the BLUE (9.9 ± 0.9% and 88.4 ± 1.1%) and RED treatments (7.7 ± 0.8% and 90.2 ± 1.0%). 4. Both the BLUE (80.9 ± 1.0%) and RED treatments (82.9 ± 1.4%) had higher hatchability of fertile eggs than the DARK treatment (76.0 ± 2.2%). 5. These results indicate that either of these LED fixtures could be used to improve hatchability and chick quality in broiler chickens. Utilising these types of lightings in commercial hatcheries will improve the efficiency via increased hatchability and quality of the chicks hatched.

  10. Cell cycle accumulation of the proliferating cell nuclear antigen PCN-1 transitions from continuous in the adult germline to intermittent in the early embryo of C. elegans.

    PubMed

    Kocsisova, Zuzana; Kornfeld, Kerry; Schedl, Tim

    2018-05-30

    The proliferating cell nuclear antigen (PCNA or PCN-1 in C. elegans), an essential processivity factor for DNA polymerase δ, has been widely used as a marker of S-phase. In C. elegans early embryos, PCN-1 accumulation is cyclic, localizing to the nucleus during S-phase and the cytoplasm during the rest of the cell cycle. The C. elegans larval and adult germline is an important model systems for studying cell cycle regulation, and it was observed that the cell cycle regulator cyclin E (CYE-1 in C. elegans) displays a non-cyclic, continuous accumulation pattern in this tissue. The accumulation pattern of PCN-1 has not been well defined in the larval and adult germline, and the objective of this study was to determine if the accumulation pattern is cyclic, as in other cells and organisms, or continuous, similar to cyclin E. To study the larval and adult germline accumulation of PCN-1 expressed from its native locus, we used CRISPR/Cas9 technology to engineer a novel allele of pcn-1 that encodes an epitope-tagged protein. S-phase nuclei were labeled using EdU nucleotide incorporation, and FLAG::PCN-1 was detected by antibody staining. All progenitor zone nuclei, including those that were not in S-phase (as they were negative for EdU staining) showed PCN-1 accumulation, indicating that PCN-1 accumulated during all cell cycle phases in the germline progenitor zone. The same result was observed with a GFP::PCN-1 fusion protein expressed from a transgene. pcn-1 loss-of-function mutations were analyzed, and pcn-1 was necessary for robust fertility and embryonic development. In the C. elegans early embryo as well as other organisms, PCN-1 accumulates in nuclei only during S-phase. By contrast, in the progenitor zone of the germline of C. elegans, PCN-1 accumulated in nuclei during all cell cycle stages. This pattern is similar to accumulation pattern of cyclin E. These observations support the model that mitotic cell cycle regulation in the germline stem and progenitor

  11. Rapid Induction of Apoptosis in Gastrulating Mouse Embryos by Ethanol and Its Prevention by HB-EGF

    PubMed Central

    Kilburn, Brian A.; Chiang, Po Jen; Wang, Jun; Flentke, George R.; Smith, Susan M.; Armant, D. Randall

    2006-01-01

    Background Ethanol exposure during gastrulation and early neurulation induces apoptosis within certain embryonic cell populations, leading to craniofacial and neurological defects. There is currently little information about the initial kinetics of ethanol-induced apoptosis, and interest in the ability of endogenous survival factors to moderate apoptosis is growing. Ethanol alters intracellular signaling, leading to cell death in chick embryos, suggesting that apoptosis could occur rapidly and that signaling pathways activated by survival factors might reduce apoptosis. Methods Pregnant mice were intubated with 1, 2, or 4 g/kg ethanol on day 7.5 of embryogenesis (E7.5) 1, 3, or 6, hours before harvesting gastrulation-stage embryos. Control animals received maltose/dextran. Blood alcohol concentrations (BAC) were determined by gas chromatography. E7.5 embryos isolated from untreated dams were cultured in vitro for 1 or 3 hr with 0 or 400 mg% ethanol and 0 or 5 nM heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF). Apoptosis was quantified using fluorescence microscopy to detect annexin V binding and DNA fragmentation [terminal deoxynucleotidyl transferase-mediated dUTP-X nick end labeling (TUNEL)] in whole-mount or sectioned embryos. Results Both annexin V binding and TUNEL were elevated (p<0.05) in embryos exposed in utero to 1 g/kg ethanol for 3 hours, increasing linearly with time and ethanol concentration. Apoptosis increased (p<0.05) in all germ cell layers. Mice treated with 4 g/kg sustained BAC of 400 mg% for nearly 3 hours, significantly increasing apoptosis within the first hour. Cultured embryos exposed to 400 mg% ethanol displayed 2- to 3-fold more TUNEL than vehicle-treated embryos (p<0.05); however, exogenous HB-EGF prevented apoptosis. Conclusions Ethanol rapidly produced apoptosis in gastrulation-stage embryos, consistent with induction by intracellular signaling. The ethanol-induced apoptotic pathway was blocked by the

  12. 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).

  13. Distinct spatiotemporal expression of ISM1 during mouse and chick development.

    PubMed

    Osório, Liliana; Wu, Xuewei; Zhou, Zhongjun

    2014-01-01

    Isthmin 1 (ISM1) constitutes the founder of a new family of secreted proteins characterized by the presence of 2 functional domains: thrombospondin type 1 repeat (TSR1) and adhesion-associated domain in MUC4 and other proteins (AMOP). ISM1 was identified in the frog embryo as a member of the FGF8 synexpression group due to its expression in the brain midbrain-hindbrain boundary (MHB) or isthmus. In zebrafish, ISM1 was described as a WNT- and NODAL-regulated gene. The function of ISM1 remains largely elusive. So far, ISM1 has been described as an angiogenesis inhibitor that has a dual function in endothelial cell survival and cell death. For a better understanding of ISM1 function, we examined its spatiotemporal distribution in mouse and chick using RT-PCR, ISH, and IHC analyses. In the mouse, ISM1 transcripts are found in tissues such as the anterior mesendoderm, paraxial and lateral plate mesoderm, MHB and trunk neural tube, as well as in the somites and dermomyotome. In the newborn and adult, ISM1 is prominently expressed in the lung and brain. In addition to its putative role during embryonic and postnatal development, ISM1 may also be important for organ homeostasis in the adult. In the chick embryo, ISM1 transcripts are strongly detected in the ear, eye, and spinal cord primordia. Remarkable differences in ISM1 spatiotemporal expression were found during mouse and chick development, despite the high homology of ISM1 orthologs in these species.

  14. Evolution of antero‐posterior patterning of the limb: Insights from the chick

    PubMed Central

    2017-01-01

    Summary The developing limbs of chicken embryos have served as pioneering models for understanding pattern formation for over a century. The ease with which chick wing and leg buds can be experimentally manipulated, while the embryo is still in the egg, has resulted in the discovery of important developmental organisers, and subsequently, the signals that they produce. Sonic hedgehog (Shh) is produced by mesenchyme cells of the polarizing region at the posterior margin of the limb bud and specifies positional values across the antero‐posterior axis (the axis running from the thumb to the little finger). Detailed experimental embryology has revealed the fundamental parameters required to specify antero‐posterior positional values in response to Shh signaling in chick wing and leg buds. In this review, the evolution of the avian wing and leg will be discussed in the broad context of tetrapod paleontology, and more specifically, ancestral theropod dinosaur paleontology. How the parameters that dictate antero‐posterior patterning could have been modulated to produce the avian wing and leg digit patterns will be considered. Finally, broader speculations will be made regarding what the antero‐posterior patterning of chick limbs can tell us about the evolution of other digit patterns, including those that were found in the limbs of the earliest tetrapods. PMID:28734068

  15. MAP kinase dependent cyclinE/cdk2 activity promotes DNA replication in early sea urchin embryos

    PubMed Central

    Kisielewska, J.; Philipova, R.; Huang, J.-Y.; Whitaker, M.

    2009-01-01

    Sea urchins provide an excellent model for studying cell cycle control mechanisms governing DNA replication in vivo. Fertilization and cell cycle progression are tightly coordinated by Ca2+ signals, but the mechanisms underlying the onset of DNA replication after fertilization remain less clear. In this study we demonstrate that calcium-dependent activation of ERK1 promotes accumulation of cyclinE/cdk2 into the male and female pronucleus and entry into first S-phase. We show that cdk2 activity rises quickly after fertilization to a maximum at 4 min, corresponding in timing to the early ERK1 activity peak. Abolishing MAP kinase activity after fertilization with MEK inhibitor, U0126, substantially reduces the early peak of cdk2 activity and prevents cyclinE and cdk2 accumulation in both sperm pronucleus and zygote nucleus in vivo. Both p27kip1 and roscovitine, cdk2 inhibitors, prevented DNA replication suggesting cdk2 involvement in this process in sea urchin. Inhibition of cdk2 activity using p27kip1 had no effect on the phosphorylation of MBP by ERK, but completely abolished phosphorylation of retinoblastoma protein, a cdk2 substrate, indicating that cdk2 activity is downstream of ERK1 activation. This pattern of regulation of DNA synthesis conforms to the pattern observed in mammalian somatic cells. PMID:19665013

  16. Early development of circadian rhythmicity in the suprachiamatic nuclei and pineal gland of teleost, flounder (Paralichthys olivaeus), embryos.

    PubMed

    Mogi, Makoto; Uji, Susumu; Yokoi, Hayato; Suzuki, Tohru

    2015-08-01

    Circadian rhythms enable organisms to coordinate multiple physiological processes and behaviors with the earth's rotation. In mammals, the suprachiasmatic nuclei (SCN), the sole master circadian pacemaker, has entrainment mechanisms that set the circadian rhythm to a 24-h cycle with photic signals from retina. In contrast, the zebrafish SCN is not a circadian pacemaker, instead the pineal gland (PG) houses the major circadian oscillator. The SCN of flounder larvae, unlike that of zebrafish, however, expresses per2 with a rhythmicity of daytime/ON and nighttime/OFF. Here, we examined whether the rhythm of per2 expression in the flounder SCN represents the molecular clock. We also examined early development of the circadian rhythmicity in the SCN and PG. Our three major findings were as follows. First, rhythmic per2 expression in the SCN was maintained under 24 h dark (DD) conditions, indicating that a molecular clock exists in the flounder SCN. Second, onset of circadian rhythmicity in the SCN preceded that in the PG. Third, both 24 h light (LL) and DD conditions deeply affected the development of circadian rhythmicity in the SCN and PG. This is the first report dealing with the early development of circadian rhythmicity in the SCN in fish. © 2015 Japanese Society of Developmental Biologists.

  17. IN VITRO CULTURE OF POSTIMPLANTATION HAMSTER EMBRYOS

    EPA Science Inventory

    In vitro culture of intact rat and mouse embryos has been described extensively, but information on the culture of other species is sparse. The present study examined some culture requirements of early somite stage hamster embryos and assessed the embryotoxic effects of sodium sa...

  18. Single cell transcriptome profiling of developing chick retinal cells.

    PubMed

    Laboissonniere, Lauren A; Martin, Gregory M; Goetz, Jillian J; Bi, Ran; Pope, Brock; Weinand, Kallie; Ellson, Laura; Fru, Diane; Lee, Miranda; Wester, Andrea K; Liu, Peng; Trimarchi, Jeffrey M

    2017-08-15

    The vertebrate retina is a specialized photosensitive tissue comprised of six neuronal and one glial cell types, each of which develops in prescribed proportions at overlapping timepoints from a common progenitor pool. While each of these cells has a specific function contributing to proper vision in the mature animal, their differential representation in the retina as well as the presence of distinctive cellular subtypes makes identifying the transcriptomic signatures that lead to each retinal cell's fate determination and development challenging. We have analyzed transcriptomes from individual cells isolated from the chick retina throughout retinogenesis. While we focused our efforts on the retinal ganglion cells, our transcriptomes of developing chick cells also contained representation from multiple retinal cell types, including photoreceptors and interneurons at different stages of development. Most interesting was the identification of transcriptomes from individual mixed lineage progenitor cells in the chick as these cells offer a window into the cell fate decision-making process. Taken together, these data sets will enable us to uncover the most critical genes acting in the steps of cell fate determination and early differentiation of various retinal cell types. © 2017 Wiley Periodicals, Inc.

  19. Confocal imaging of whole vertebrate embryos reveals novel insights into molecular and cellular mechanisms of organ development

    NASA Astrophysics Data System (ADS)

    Hadel, Diana M.; Keller, Bradley B.; Sandell, Lisa L.

    2014-03-01

    Confocal microscopy has been an invaluable tool for studying cellular or sub-cellular biological processes. The study of vertebrate embryology is based largely on examination of whole embryos and organs. The application of confocal microscopy to immunostained whole mount embryos, combined with three dimensional (3D) image reconstruction technologies, opens new avenues for synthesizing molecular, cellular and anatomical analysis of vertebrate development. Optical cropping of the region of interest enables visualization of structures that are morphologically complex or obscured, and solid surface rendering of fluorescent signal facilitates understanding of 3D structures. We have applied these technologies to whole mount immunostained mouse embryos to visualize developmental morphogenesis of the mammalian inner ear and heart. Using molecular markers of neuron development and transgenic reporters of neural crest cell lineage we have examined development of inner ear neurons that originate from the otic vesicle, along with the supporting glial cells that derive from the neural crest. The image analysis reveals a previously unrecognized coordinated spatial organization between migratory neural crest cells and neurons of the cochleovestibular nerve. The images also enable visualization of early cochlear spiral nerve morphogenesis relative to the developing cochlea, demonstrating a heretofore unknown association of neural crest cells with extending peripheral neurite projections. We performed similar analysis of emb