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[Embryonic development of 2 Phyllobothriidae (Cestoda: Tetraphyllidea)].

Caulobothrium longicolle (Linton, 1890) and Phyllobothrium gracile (Weld, 1855) (Cestoda: Tetraphyllidea, Phyllobothriidae) have the same embryonic development with the following characteristic data: --a small number of vitelline cells (2 or 3) pass with the zygote in the ootype;--a non operculate thin egg-shell;--the entire and equal zyhote cleavage following by unequal divisions leading to the formation of four blastomere types (Macromeres, secondary Macromere, Mesomeres and Micromeres);--the differentiation of two syncytial embryonic envelopes during the preoncospheral phase. The outer envelope encloses the vitelline material remnant and three Macromeres among which the secondary Macromere. The inner envelope or embryophore, originates from five or six Mesomeres;--the presence of the oncospheral membrane;--the Phyllobothriidae ontogenesis is similar to that of the Onchobothriidae. By their embryonic features, the Tetraphyllidea are close to the Cyclophyllidea. This similarity suggests a phylogenic relationship between those two Cestoda orders.

Animals↗

The lethal (2) giant larva (l(2)gl), a recessive oncogene, is required during embryonic and post-embryonic development in Drosophila.

Recessive oncogenes have genetic functions important for the regulation of tissue growth and differentiation. Defining the role of these genes in normal developmental and physiological processes is important to the development of the accurate models of the normal regulation of growth and differentiation. We report here a genetic analysis of the requirement for the lethal (2) giant larva function during development. The results demonstrate that the lethal (2) giant larva function is required during embryonic and post-embryonic development to maintain the normal developmental capacity.

Animals↗

Estradiol reverses the limiting effects of clomiphene citrate on early embryonic development in the in vitro perfused rabbit ovary.

This study examined whether the addition of estradiol (E2) to the perfused rabbit ovary would reverse the deleterious effects of clomiphene citrate (CC) on early embryonic development. Ovaries were perfused with CC (10(-5) M) or CC + E2 (1 to 1000 ng/ml). Human chorionic gonadotropin (hCG, 50 IU) was added to the perfusate of each ovary. In vitro ovulated ova in cumulus were retrieved and inseminated in vitro. E2 significantly increased the percentage of ovulated ova achieving (1) the 2-cell stage at 36 hours, (2) the morula stage by 84 hours, and (3) the blastocyst stage at 132 hours. The percentage of inseminated ova showing evidence of degeneration was reduced in ovaries treated with E2. These data suggest that CC may exert an antiestrogenic effect on the intrafollicular oocyte, which interferes with postfertilization development.

Animals↗

The LDL receptor gene family, apolipoprotein B and cholesterol in embryonic development.

In recent years, a number of genes that are involved in cholesterol synthesis, its systemic or intercellular transport or lipid metabolism in general have been found to play important roles during embryonic development. In this article, we present a brief overview of these genes, their molecular functions as we understand them to date and our current interpretation of possible mechanisms by which genetic deficiency states might affect the development of the embryo, in particular the formation of the central nervous system.

Animals↗

Clathrin adaptor AP-2 is essential for early embryonal development.

The heterotetrameric adaptor protein (AP) complexes AP-1, AP-2, AP-3, and AP-4 play key roles in transport vesicle formation and cargo sorting in post-Golgi trafficking pathways. Studies on cultured mammalian cells have shown that AP-2 mediates rapid endocytosis of a subset of plasma membrane receptors. To determine whether this function is essential in the context of a whole mammalian organism, we carried out targeted disruption of the gene encoding the mu2 subunit of AP-2 in the mouse. We found that mu2 heterozygous mutant mice were viable and had an apparently normal phenotype. In contrast, no mu2 homozygous mutant embryos were identified among blastocysts from intercrossed heterozygotes, indicating that mu2-deficient embryos die before day 3.5 postcoitus (E3.5). These results indicate that AP-2 is indispensable for early embryonic development, which might be due to its requirement for cell viability.

Adaptor Protein Complex 2↗

Fertilization rates and embryonic development in CD-1 mice fed fungal endophyte-infected tall fescue seed.

The objective of this study was to assess the effects of endophyte-infected tall fescue (KY-31) seed (80% infected, containing Acremonium coenophialum ) on the fertilization rates and embryonic development of CD-1 mice. Twenty-four pairs of mature CD-1 mice were randomly allocated to two dietary treatments containing 40% mouse chow and 1) 60% noninfected tall fescue seed or 2) 60% infected tall fescue seed (w/w). The mice were preconditioned on their respective diets for 60 d prior to 96 h of cohabitation between pairs of males and females. Following the removal of the males, equal numbers of females (n = 4) from each dietary treatment were sacrificed at 80 h, 6 and 10 d after vaginal plug establishment. The female reproductive tracts were excised and flushed to recover eggs and embryos or dissected to identify the number and status of the implanted fetuses. The number of eggs-embryos and/or fetuses per mouse present at the time of sacrifice were 9.8, 8.8 and 11.5 vs 8.2, 5.3 and 4.8 for the mice fed the noninfected and infected fescue seeds, respectively. Similarly, the conception rates (%) were 100, 100 and 100 vs 73, 50 and 50 for the two treatments. The mice sacrificed at 80 h and fed the noninfected fescue had 9.8 expanded blastocysts per animal, whereas those fed the infected fescue had only 4.0 expanded blastocysts. The results suggest that the infected fescue seed consumed by these mice had a significant effect on fertilization rates. More importantly, the infected fescue seed influenced negatively the development of these embryos during the embryonic-preimplantation period.

Journal Article↗

[Effect of bushen huoxue decoction on in vitro fertilization and early embryonic development in mice].

OBJECTIVE: To explore the effect of Bushen Huoxue decoction (BSHX) on female reproduction and elucidate its therapeutic mechanism to infertility. METHODS: The BSHX medicated serum of rabbit, as a supplement, was co-cultured with the sperm and ovum of non-copulated mice, and the 2-cell embryos of copulated female mice separately, to observe the changes of in vitro fertilization rate (IVF) and early embryogenesis rate. RESULTS: By co-cultured with BSHX medicated serum, IVF rate was increased obviously (P < 0.01), and the follow-up early embryogenesis rate at various period was promoted, particularly that of the 4-cell and 8-cell embryos (P < 0.05). No influence on developmental rate of in vitro 4-cell embryos obtained from the in vivo 2-cell embryos. But the development of the 8-cell embryos, morula, blastula and hatching were promoted significantly. CONCLUSION: BSHX could raise the fertilization rate and promote the early embryonic development.

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Expression of carbonic anhydrases IX and XII during mouse embryonic development.

BACKGROUND: Of the thirteen active carbonic anhydrase (CA) isozymes, CA IX and XII have been linked to carcinogenesis. It has been suggested that these membrane-bound CAs participate in cancer cell invasion, which is facilitated by an acidic tumor cell environment. Since active cell migration is a characteristic feature of embryonic development, we set out to explore whether these isozymes are expressed in mouse embryos of different ages. The studies were focused on organogenesis stage. RESULTS: Immunohistochemistry demonstrated that both CA IX and XII are present in several tissues of the developing mouse embryo during organogenesis. Staining for CA IX revealed a relatively wide distribution pattern with moderate signals in the brain, lung, pancreas and liver and weak signals in the kidney and stomach. The expression pattern of CA XII in the embryonic tissues was also relatively broad, although the intensity of immunostaining was weak in most tissues. The CA XII-positive tissues included the brain, where the most prominent staining was seen in the choroid plexus, and the stomach, pancreas, liver and kidney. CONCLUSION: Membrane-bound CA isozymes IX and XII are expressed in various tissues during mouse organogenesis. These enzymes may regulate ion and pH homeostasis within the developing embryo.

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Constitutive genomic methylation during embryonic development of Xenopus.

Methylation of CpG dinucleotides is a predominant modification of genomic DNA in many species, especially in vertebrates. This modification, generally associated with transcriptional repression, is rapidly and globally lost during mammalian pre-implantation development. This loss of methylation is gradually reversed during subsequent stages of development. Here we show that the amphibian Xenopus laevis maintains high levels of DNA methylation during early embryonic development. The methylation status of specific loci is independent of the temporal expression profile. The observations have profound implications for the regulation of early embryonic gene regulation and genome function.

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Replacement of K-Ras with H-Ras supports normal embryonic development despite inducing cardiovascular pathology in adult mice.

Ras proteins are highly related GTPases that have key roles in regulating growth, differentiation and tumorigenesis. Gene-targeting experiments have shown that, out of the three mammalian ras genes, only K-ras is essential for normal mouse embryogenesis, and that mice deprived of H-ras and/or N-ras show no major phenotype. We generated mice (HrasKI) in which the K-ras gene had been modified to encode H-Ras protein. HrasKI mice produce undetectable amounts of K-Ras but-in contrast to mice homozygous for a null K-ras allele-they are born at the expected mendelian frequency, indicating that H-Ras can be substituted for K-Ras in embryonic development. However, adult HrasKI mice show dilated cardiomyopathy associated with arterial hypertension. Our results show that K-Ras can be replaced by H-Ras in its essential function in embryogenesis, and indicate that K-Ras has a unique role in cardiovascular homeostasis.

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Embryonic development of the house shrew (Suncus murinus). II. Embryos at stages 11 and 12 with 13 to 29 pairs of somites, showing limb bud formation and closed cephalic neural tube.

The embryonic development of the laboratory house shrew (Suncus murinus) was observed during the period from 13 to 29 pairs of somites and was compared with that of other mammalian species. This period ranged from 10 to 12 days after fertilization. Embryos were allocated to stages 11A-C and 12A-C according to somitic count. These embryos were characterized by the following features. 1. The curvature of the body became gradually enhanced, changing from a parenthesis-like to a bracket-like shape. 2. The swelling of the future hindlimb bud appeared at stage 11A, while the forelimb bud became apparent at stage 12A. The latter developed rapidly and had exceeded the former in size by the end of stage 12C. 3. The timing of closure of the cephalic neuropores varied, but the order of closure is constant. The prosencephalic neuropore closed later. The caudal neuropore still existed even at the end of stage 12C. 4. In all of the cranial nerve ganglia except for the auditory one, crest-derived cells seemed to contribute to the initial formation, while placodal cells seemed to be involved later. A different pattern of primordial formation was observed between trigeminal/facial and glossopharyngeal/vagal ganglia. 5. By the end of stage 12C, the eye cup and nasal pit began to form, and the endolymphatic appendage also appeared. 6. In the heart, the sinus valves were well developed, and the atrium and ventricle were being divided by the developing interatrial septum I and interventricular septum, respectively. 7. The thyroid primordium was temporarily everted and projected into the pharynx. The internal structure of this protruding thyroid resembled "tuberculum thyroideum" observed in the human embryo.

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Effect of titanium exposure on embryonic development during pre-implantation period in rats.

Doses of titanium trichloride (1/10th and 1/5th of LD50) were administered once and daily to pregnant rats to assess their effect on embryonic development. 1/5th dose of TiCl3 administered once orally on 1st, 2nd and 3rd day post-coitum. Similarly 1/10th of LD50 was administered daily. Results revealed that 1/10th LD50 dose of TiCl3 was more effective during pre-implantation period as number of 4 and 8-celled embryos decreased as compared to 1/5th. Delayed hatching of the blastocysts on day 5 was registered in TiCl3 treated dam.

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Lucifer yellow stains displaced amacrine cells of the chicken retina during embryonic development.

We have used Lucifer Yellow for histological tracing of displaced amacrine cells within the ganglion cell layer (GCL) during the embryonic development of the chicken retina. Incubating whole eyes in the dye leads to bright staining of all displaced amacrine cells, whereas ganglion cells and glial cells are not stained. A subpopulation of cells of the inner part of the inner nuclear layer (INL) are also stained (for further details see ref. 13). Kainic acid, which is known to interfere with and kill amacrine cell systems, blocks the staining of these cells fully. This in addition to histological evidence confirms that the LY-stained cells in the GCL are displaced amacrine cells. Of the cells in the GCL, 23% (+/- 3%) are of the displaced amacrine type. Further, we find that the cytoarchitectural arrangement of these cells changes significantly during development.

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Isolation and characterization of a Drosophila gene essential for early embryonic development and formation of cortical cleavage furrows.

We have isolated a new female sterile mutant from Drosophila melanogaster, which arrests the embryonic development during the transition from syncytial to cellular blastoderm. Cytological analysis of the mutant embryos indicates that pseudocleavage furrows in the syncytial blastoderm are abnormal but not completely disrupted. However, cleavage furrows during cellularization are totally disorganized, and no embryos can develop beyond this stage. Consistent with this observation, the expression of this gene peaks around the cellular blastoderm and not in any later developmental stages. Based on immunofluorescence experiments, the protein product of this gene is localized in both pseudocleavage furrows at the syncytial blastoderm and in the cleavage furrows during the cellularization stage. Sequence homology analysis demonstrates a modest, but statistically significant, similarity of this protein with the carboxyl-terminal domains of dystrophin and a family of proteins collectively known as apodystrophins. It is possible that this protein may play an essential role in organizing and maintaining a specialized cytoskeletal structure, a function also suggested for dystrophin and apodystrophins.

Amino Acid Sequence↗

Abnormal embryonic development induced by antibodies to rat visceral yolk-sac endoderm: isolation of the antigen and localization to microvillar membrane.

An antigenic substance was isolated from rat visceral yolk-sac endoderm of the 18th-20th days of gestation by extraction with the nonionic detergent Nonidet P-40, Sephacryl S-300 gel filtration, and Ricinus communis agglutinin affinity chromatography. The rabbit antiserum directed against this antigenic substance when injected into pregnant rats during the period of organogenesis caused abnormal embryonic development, fetal growth retardation, and embryonic death. Ouchterlony gel diffusion analysis demonstrated that the antiserum formed one immunoprecipitin band against the crude detergent extract and a complete identity between the present visceral yolk-sac antigen and the renal glycoprotein antigen previously isolated (C. C. K. Leung, (1982) J. Exp. Med. 156, 372-384). The antigen eluted from the antibody affinity column appeared to consist of two major peptides of 60 and 30 kDa when analyzed by SDS-polyacrylamide gel electrophoresis. Indirect immunofluorescent and immunoperoxidase localization studies at the light microscopic level demonstrated that both rat renal proximal tubule and embryonic visceral yolk-sac endoderm at various gestational stages (including the organogenetic period) shared the same antigen. Indirect immunoperoxidase localization studies at the electron microscopic level demonstrated that the antigen was a part of (or associated with) the microvillar membrane and membrane invaginations at the base of the microvilli of the renal proximal tubule and visceral yolk-sac endoderm. In vivo immunoperoxidase localization studies demonstrated that the teratogenic antibodies localized within the large phagolysosomes and the apical vesicles of the visceral yolk-sac endoderm. It is postulated that visceral yolk-sac pathology was induced by the antibodies.

Animals↗

Effect of platelet activating factor on embryonic development and implantation in the mouse.

Platelet activating factor (PAF) was administered to female mice in order to investigate its effect on ovulation rate and on oocyte quality including their in-vitro embryonic development, implantation and uterine receptivity. In experiment 1, 4-week-old female mice were assigned to receive PAF or phosphate buffered saline for 4 consecutive days. On the second day of this treatment, pregnant mares' serum gonadotrophin was administered and human chorionic gonadotrophin (HCG) 48 h later, after which copulation occurred. Oocytes were collected on the following day and evaluated. The mean number of oocytes and zygotes (two pronuclear stage embryos) recovered from the PAF-treated group was not different from the control group (31 versus 27), but the proportion of zygotes was higher in PAF-treated group than in controls (83 versus 68%, P < 0.05, PAF versus controls). Although the rate of in-vitro first cleavage was not different in the two groups (82 versus 69% respectively), hatching was higher in the PAF-treated group than control mice (99 versus 83%, P < 0.01). In experiment 2, the in-vitro developed blastocysts from experiment 1 were transferred into the uterus of day 3 pseudopregnant PAF-treated or control recipients. Three different combinations of intrauterine transfer were performed; PAF embryo to control recipient (PAF-->C: n = 19), control embryo to PAF recipient (C-->PAF: n = 19), and control embryo to control recipient (C-->C: n = 22). Implantation and abortion were assessed on day 19 posttransfer. The implantation rate of C-->PAF (23.7%) was lower than C-->C (31.1%, P < 0.05), but was not different from PAF-->C (31.2%). Further, C-->PAF showed a higher abortion rate per embryo (29.6%) than PAF-->C (12.7%, P < 0.05), but was not different from C-->C (24.4%). In the present study, PAF administration enables females to produce oocytes with a higher potential for fertilization, in-vitro development and implantation, but has a detrimental effect on uterine receptivity to embryos.

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Regulation of murine TGFbeta2 by Pax3 during early embryonic development.

Previously our laboratory identified TGFbeta2 as a potential downstream target of Pax3 by utilizing microarray analysis and promoter data base mining (Mayanil, C. S. K., George, D., Freilich, L., Miljan, E. J., Mania-Farnell, B. J., McLone, D. G., and Bremer, E. G. (2001) J. Biol. Chem. 276, 49299-49309). Here we report that Pax3 directly regulates TGFbeta2 transcription by binding to cis-regulatory elements within its promoter. Chromatin immunoprecipitation revealed that Pax3 bound to the cis-regulatory elements on the TGFbeta2 promoter (GenBanktrade mark accession number AF118263). Both TGFbeta2 promoter-luciferase activity measurements in transient cotransfection experiments and electromobility shift assays supported the idea that Pax3 regulates TGFbeta2 by directly binding to its cis-regulatory regions. Additionally, by using a combination of co-immunoprecipitation and chromatin immunoprecipitation, we show that the TGFbeta2 cis-regulatory elements between bp 741-940 and bp 1012-1212 bind acetylated Pax3 and are associated with p300/CBP and histone deacetylases. The cis-regulatory elements between bp 741 and 940 in addition to associating with acetylated Pax3 and HDAC1 also associated with SIRT1. Whole mount in situ hybridization and quantitative real time reverse transcription-PCR showed diminished levels of TGFbeta2 transcripts in Pax3(-/-) mouse embryos (whose phenotype is characterized by neural tube defects) as compared with Pax3(+/+) littermates (embryonic day 10.0; 30 somite stage), suggesting that Pax3 regulation of TGFbeta2 may play a pivotal role during early embryonic development.

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Requirement for Casper (c-FLIP) in regulation of death receptor-induced apoptosis and embryonic development.

Casper (c-FLIP) associates with FADD and caspase-8 in signaling complexes downstream of death receptors like Fas. We generated Casper-deficient mice and cells and noted a duality in the physiological functions of this molecule. casper-/- embryos do not survive past day 10.5 of embryogenesis and exhibit impaired heart development. This phenotype is reminiscent of that reported for FADD-/- and caspase-8-/- embryos. However, unlike FADD-/- and caspase-8-/- cells, casper-/- embryonic fibroblasts are highly sensitive to FasL- or TNF-induced apoptosis and show rapid induction of caspase activities. NF-kappaB and JNK/SAPK activation is intact in TNF-stimulated casper-/- cells. These results suggest that Casper has two distinct roles: to cooperate with FADD and caspase-8 during embryonic development and to mediate cytoprotection against death factor-induced apoptosis.

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