Embryo Development (embryo + development)

Distribution by Scientific Domains
Distribution within Life Sciences

Kinds of Embryo Development

  • early embryo development
  • mouse embryo development

  • Selected Abstracts

    Consequences of Nitric Oxide Synthase Inhibition During Bovine Oocyte Maturation on Meiosis and Embryo Development

    KRL Schwarz
    Contents The importance of nitric oxide synthase (NOS) in bovine oocyte maturation was investigated. Oocytes were in vitro matured with the NOS inhibitor Nw - l -nitro-arginine methyl-ester (10,7, 10,5 and 10,3 m l -NAME) and metaphase II (MII) rates and embryo development and quality were assessed. The effect of l -NAME (10,7 m) during pre-maturation and/or maturation on embryo development and quality was also assessed. l -NAME decreased MII rates (78,82%, p < 0.05) when compared with controls without l -NAME (96%). Cleavage (77,88%, p > 0.05), Day 7 blastocyst rates (34,42%, p > 0.05) and total cell numbers in blastocysts were similar for all groups (146,171 cells, p > 0.05). Day 8 blastocyst TUNEL positive cells (3,4 cells) increased with l -NAME treatment (p < 0.05). For oocytes cultured with l -NAME during pre-maturation and/or maturation, Day 8 blastocyst development (26,34%) and Day 9 hatching rates (15,22%) were similar (p > 0.05) to controls pre-matured and matured without NOS inhibition (33 and 18%, respectively), while total cell numbers (Day 9 hatched blastocysts) increased (264,324 cells, p < 0.05) when compared with the controls (191 cells). TUNEL positive cells increased when NOS was inhibited only during the maturation period (8 cells, p < 0.05) when compared with the other groups (3,4 cells). NO may be involved in meiosis progression to MII and its deficiency during maturation increases apoptosis in embryos produced in vitro. Nitric oxide synthase inhibition during pre-maturation and/or maturation affects embryo quality. [source]

    Effect of Antioxidants During Bovine In Vitro Fertilization Procedures on Spermatozoa and Embryo Development

    FS Gonçalves
    Contents Increased amounts of reactive oxygen species (ROS) during in vitro fertilization (IVF) may cause cytotoxic damage to gametes, whereas small amounts of ROS favour sperm capacitation. The aim of this study was to investigate the effect of antioxidants [50 ,m,-mercaptoethanol (,-ME) and 50 ,m cysteamine (Cyst)] or a pro-oxidant (5 mm buthionine sulfoximine) on the quality and penetrability of spermatozoa into bovine oocytes and on the subsequent embryo development and quality when added during IVF. Sperm quality, evaluated by the integrity of plasma and acrosomal membranes, and mitochondrial function, was diminished (p < 0.05) after 4-h culture in the presence of antioxidants. Oocyte penetration rates were similar between treatments (p > 0.05), but antioxidants adversely affected the normal pronuclear formation rates (p < 0.05). The incidence of polyspermy was high for ,-ME (p < 0.05). No differences were observed in cleavage rates between treatments (p > 0.05). However, the developmental rate to the blastocyst stage was adversely affected by Cyst treatment (p < 0.05). The quality of embryos that reached the blastocyst stage, evaluated by total, inner cell mass (ICM) and trophectoderm cell numbers and ICM/total cell ratio was unaffected (p > 0.05) by treatments. The results indicate that ROS play a role in the fertilizing capacity in bovine spermatozoa, as well as in the interaction between the spermatozoa and the oocytes. It can be concluded that supplementation with antioxidants during IVF procedures impairs sperm quality, normal pronuclear formation and embryo development to the blastocyst stage. [source]

    Effect of Taurine and Melatonin in the Culture Medium on Buffalo In Vitro Embryo Development

    BM Manjunatha
    Contents This study was carried out to investigate the effect of supplementing culture medium with different concentrations of taurine and melatonin, on buffalo oocyte in vitro meiotic maturation and embryo development. In experiment 1, oocytes were matured in vitro and the cleaved embryos were cultured in the same following seven culture medium; (i) control (TCM 199 + 10% SS); (ii) control + 0.5 mm taurine; (iii) control + 1 mm taurine; (iv) control + 3 mm taurine; (v) control + 5 ,m melatonin; (vi) control + 10 ,m melatonin and (vii) control + 50 ,m melatonin. In experiment 2, based on the results of experiment 1, to examine the synergistic effect of antioxidants, the oocytes were matured in culture medium (TCM199 + 10% SS), supplemented with both taurine at 1 mm and melatonin at 10 ,m concentration and the cleaved embryos were cultured in the same medium. Supplementation of taurine at 1 mm concentration in the culture medium resulted in a higher (p < 0.05) transferable embryo (TE) yield when compared with control (20.6% vs 14.1%). Supplementation of melatonin at 10 and 50 ,m concentration in the culture medium resulted in a higher (p < 0.05) meiotic maturation rate (90.3% and 88.8% respectively) and TE yield (28.4% and 27.2% respectively), than the other treatments. In experiment 2, the TE yield did not improve by supplementing the culture medium with both taurine and melatonin, when compared with melatonin alone. In conclusion, the results of this study demonstrated that, enriching the culture medium with taurine and melatonin, improves in vitro embryo production efficiency in buffaloes. In particular, a high TE yield was obtained by enriching the culture medium with 10 ,m melatonin. [source]

    Oviductal Fluid Proteins Associated with the Bovine Zona Pellucida and the Effect on In Vitro Sperm,Egg Binding, Fertilization and Embryo Development

    RF Gonçalves
    Contents Studies have demonstrated that oviductal fluid (ODF) proteins associate with eggs of numerous species including the bovine. In this study, the association of three ODF proteins, the bovine oestrus-associated protein, osteopontin (OPN), lipocalin-type prostaglandin D synthase (L-PGDS), with the bovine zona pellucida (ZP) was demonstrated by immunohistochemistry and western blot. The biological function of ODF derived egg-associated OPN and L-PGDS in sperm binding, fertilization and embryonic development was also explored. In vitro matured bovine oocytes were pre-incubated with ODF collected by cannula from cows in oestrus, or ODF with antibodies to OPN, L-PGDS and bovine serum albumin (BSA). Following incubation, oocytes were inseminated with 1 × 105 frozen-thawed spermatozoa, and they were evaluated for sperm binding, fertilization and embryonic development in vitro. Pre-treatment of ODF with antibodies to all of proteins reduced sperm binding to the ZP and fertilization in vitro. Cleavage rates were not significantly different among incubations, but rates of embryo development were significantly decreased. We conclude that antibodies to OPN, L-PGDS and BSA react with oocytes incubated with ODF and inhibit sperm binding, fertilization and embryonic development in vitro, suggesting a potential role of these proteins in these events. [source]

    Effects of Exogenous ACTH during Oestrus on Early Embryo Development and Oviductal Transport in the Sow

    Y Brandt
    Contents This study was conducted to assess the effects of ACTH injections on the early development of embryos and their transportation to the uterus. Fifteen sows were monitored for ovulation using transrectal ultrasonography during the first two oestrous periods after weaning. The sows were randomly divided into a control group (C group, n = 8) and an ACTH-treated group (ACTH group, n = 7), and were all surgically fitted with intra-jugular catheters. From the onset of the second standing oestrus after weaning, the sows were injected (NaCl/synthetic ACTH) every 4 h. Blood samples were collected immediately before and 45 min after each injection. All sows were inseminated once 10,33 h before ovulation in their second oestrus after weaning. At 48 (n = 4) or 60 (n = 11) h after ovulation during their second oestrus, the sows were killed and the embryos retrieved from the oviduct and uterus. The embryos were counted and compared with the number of corpora lutea, cleavage rate was noted and, finally, the embryos were prepared for confocal laser scanning microscopy and transmission electron microscopy. There was no difference between the groups regarding cleavage rate, the cytoskeleton, or the number of active nucleoli. However, the ACTH group had significantly (p < 0.05) fewer ova/embryos retrieved (51%) than the C group (81%), and there was a tendency towards faster transportation to the uterus in the ACTH group, possibly because of high progesterone concentrations during treatment. To conclude, administration of ACTH every 4 h from onset of oestrus to 48 h caused significant loss of oocytes or embryos, and possibly faster transportation through the oviduct. [source]

    Effect of Days Post-Partum, Breed and Ovum Pick-Up Scheme on Bovine Oocyte Recovery and Embryo Development

    AS Lopes
    Contents The objective of this study was to investigate (i) the effect of two different ovum pick-up (OPU) schemes (once vs twice weekly aspirations) on oocyte recovery rate, quality and subsequent in vitro embryo development, (ii) the influence of days post-partum on oocyte recovery and (iii) possible differences in OPU results from two different herds. In group A, OPU was performed twice weekly in two Holstein Friesian (HF) and three Danish Red and White (DRW) cows from a private herd. In the research herd, two groups of eight HF cows were investigated: group B (OPU once weekly) and group C (OPU twice weekly). The collected oocytes were subsequently submitted to in vitro embryo production. More oocytes were recovered from the private herd when compared with the research herd. In the research herd, the twice weekly scheme aspirated more oocytes than the once weekly scheme. The quality of the retrieved oocytes was significantly different between groups B and C but not between groups A and C, and HF cows yielded higher quality oocytes than DRW cows (p = 0.029). Oocytes from group C showed higher level of embryonic development than group B oocytes. No differences in blastocyst rates were observed between groups A and C. Session affected the number of retrieved oocytes and subsequent developmental rates, with these being lower in the first compared with the last sessions. Finally, there was no significant effect of days post-partum in the number and quality of the retrieved oocytes, likely because of the small group size and high variation between sessions. [source]

    Postovulatory Effect of Intravenous Administration of Lipopolysaccharide (E. coli, O55:B5) on the Contractile Activity of the Oviduct, Ova Transport, Binding of Accessory Spermatozoa to the Zona Pellucida and Embryo Development in Sows

    AM Mwanza
    Contents The effect of lipopolysaccharide (LPS) (E. coli, O55:B5), administered 18 h after ovulation in the second oestrus after weaning, on the contractile activity of the oviduct, ova transport, sperm binding to zona pellucida (ZP) and embryo development, was studied in 14 Swedish crossbred (Landrace Yorkshire) multiparous sows. The endotoxin group (E-group) sows were administered with 300 ng/kg of LPS while the control group (C-group) sows were administered with 5 ml of saline i.v. via an indwelling jugular cannula. Immediately after evidence of standing oestrus, a Millar® pressure transducer was placed intraluminally about 3 cm into the mid-isthmus, via laparotomy. Pressure recordings of the oviduct were collected from all conscious sows until slaughter. After slaughter, the genital tract opposite to the side with the transducer was retrieved, and three equal isthmic segments and the first third of the uterine horn part adjacent to the utero-tubal-junction (UTJ) were flushed separately to recover the ova. The intervals (mean±SD) from ovulation to slaughter (OS) and insemination to ovulation (IO) were not different between the E-group (44.5±5.7 h; 13.3±6.5 h) and the C-group (42.7±5.9 h; 14.8±4.1 h), respectively. Ova recovery rate (RR) in the E-group (80.2±22.9%) did not differ from that in the C-group (85.2±4.5%). The frequency distribution of ova recovered in the different segments did not significantly (p>0.05) differ between the groups. The E-group showed higher cleavage rate than controls. A higher proportion of spermatozoa bound to the ZP was also found in the E-group compared with controls. The isthmic intraluminal pressure slightly increased (p=0.07) 18 h after ovulation and immediately following LPS in the E-group, compared with the C-group. The frequencies of phasic pressure fluctuations were significantly (p<0.05) lower at 30 and 38 h after ovulation in the E- than in the C-group. It can be concluded from the present study that a single i.v. administration of LPS (300 ng/kg body weight) to sows, 18 h after ovulation might be associated with changes in isthmic pressure and the frequency of phasic pressure fluctuations, increased numbers of spermatozoa attached to the ZP and an enhanced embryo development but not with ova transport rates. [source]

    Embryo development of Corticium candelabrum (Demospongiae: Homosclerophorida)

    Sonia De Caralt
    Abstract. Corticium candelabrum is a homosclerophorid sponge widespread along the rocky Mediterranean sublittoral. Scanning and transmission electron microscopy were used to describe the gametes and larval development. The species is hermaphroditic. Oocytes and spermatocytes are clearly differentiated in April. Embryos develop from June to July when the larvae are released spontaneously. Spermatic cysts originate from choanocyte chambers and spermatogonia from choanocytes by choanocyte mitosis. Oocytes have a nucleolate nucleus and a cytoplasm filled with yolk granules and some lipids. Embryos are surrounded by firmly interlaced follicular cells from the parental tissue. A thin collagen layer lies below the follicular cells. The blastocoel is formed by migration of blastomeres to the morula periphery. Collagen is spread through the whole blastocoel in the embryo, but is organized in a dense layer (basal lamina) separating cells from the blastocoel in the larva. The larva is a typical cinctoblastula. The pseudostratified larval epithelium is formed by ciliated cells. The basal zone of the ciliated cells contains lipid inclusions and some yolk granules; the intermediate zone is occupied by the nucleus; and the apical zone contains abundant electron-lucent vesicles and gives rise to cilia with a single cross-striated rootlet. Numerous paracrystalline structures are contained in vacuoles within both apical and basal zones of the ciliated cells. Several slightly differentiated cell types are present in different parts of the larva. Most cells are ciliated, and show ultrastructural particularities depending on their location in the larvae (antero-lateral, intermediate, and posterior regions). A few smaller cells are non-ciliated. Several features of the C. candelabrum larva seem to support the previously proposed paraphyletic position of homoscleromorphs with respect to the other demosponges. [source]

    A note on the development of the embryos of anglerfish Lophius piscatorius

    C. H. Laurenson
    Embryo development over a 5 day period are shown and described from a rare find of anglerfish Lophius piscatorius spawn that was obtained at the Shetland Islands, U.K. The stage of development shown is earlier than that previously available for the species. [source]

    Embryo development of porcine oocytes after injection with miniature pig sperm and their extracts

    Daizou MATSUURA
    ABSTRACT This study examined embryo development of porcine oocytes after microinjection of sperm extracts (SE) in porcine intracytoplasmic sperm injection (ICSI). SE was prepared from miniature pig sperm by a nonionic surfactant, and various concentrations (0.02, 0.04 and 0.08 mg/mL) of SE were injected into the matured oocytes with a first polar body. In the pronuclear stage, the rate of oocytes with two pronuclei and a second polar body (21.4%) in the sperm and SE (0.04 mg/mL) injection group was significantly higher (P < 0.05) compared to other groups. The rate of 2,4-cell stage in sperm and SE (0.04 mg/mL) injection group was 38.1%, and it was significantly higher than that in the sperm injection group (22.9%). The rate of blastocyst stage in sperm and SE (0.04 mg/mL) injection group was 21.4%, the value was significantly higher than those in SE (0.08 mg/mL) injection group (0%), sperm injection group (5.7%), and sperm and SE (0.08 mg/mL) injection group (2.6%). These results suggest that SE induces activation of porcine oocytes and their further embryonic development, and that SE is effective for porcine ICSI. [source]

    Characterization and expression of AmphiBMP3,/3b gene in amphioxus Branchiostoma japonicum

    Yi Sun
    Bone morphogenetic proteins (BMPs) are responsible for regulating embryo development and tissue homeostasis beyond osteogenesis. However, the precise biological roles of BMP3 and BMP3b remain obscure to a certain extent. In the present study, we cloned an orthologous gene (AmphiBMP3/3b) from amphioxus (Branchiostoma japonicum) and found its exon/intron organization is highly conserved. Further, in situ hybridization revealed that the gene was strongly expressed in the dorsal neural plate of the embryos. The gene also appeared in Hatschek's left diverticulum, neural tube, preoral ciliated pit and gill slit of larvae, and adult tissues including ovary, neural tube and notochordal sheath. Additionally, real-time quantitative polymerase chain reaction (RTqPCR) analysis revealed that the expression displayed two peaks at gastrula and juvenile stages. These results indicated that AmphiBMP3/3b, a sole orthologue of vertebrate BMP3 and BMP3b, might antagonize ventralizing BMP2 orthologous signaling in embryonic development, play a role in the evolutionary precursors of adenohypophysis, as well as act in female ovary physiology in adult. [source]

    Identification of asymmetrically localized transcripts along the animal,vegetal axis of the Xenopus egg

    Kensuke Kataoka
    In many organisms, proper embryo development depends on the asymmetrical distribution of mRNA in the cytoplasm of the egg. Here we report comprehensive screening of RNA localized in the animal or vegetal hemisphere of the Xenopus egg. Macroarrays including over 40 000 independent embryonic cDNA clones, representing at least 17 000 unigenes, were differentially hybridized with labeled probes synthesized from the mRNA of animal or vegetal blastomeres. After two rounds of screening, we identified 33 clones of transcripts that may be preferentially distributed in the vegetal region of the early stage embryo, but transcripts localized in the animal region were not found. To assess the array results, we performed northern blot and quantitative real-time reverse transcription,polymerase chain reaction analysis. As a result, 21 transcripts of the 33 were confirmed to be localized in the vegetal region of the early stage embryo. Whole-mount in situ hybridization analysis revealed that 11 transcripts, including 7 previously reported genes, were localized in the vegetal hemisphere of the egg. These 11 transcripts were categorized into three groups according to their expression patterns in the egg. The first group, which contained four transcripts, showed uniform expression in the vegetal hemisphere, similar to VegT. The second group, which contained three transcripts, showed gradual expression from the vegetal pole to the equator, similar to Vg1. The last group, which contained three transcripts, was expressed at the germ plasm, similar to Xdazl. One transcript, Xwnt11, showed both the second and the third expression patterns. [source]

    IFN-, induces apoptosis in mouse embryonic stem cells, a putative mechanism of its embryotoxicity

    Gang-Ming Zou
    It has been reported that interferon (IFN)-, should inhibit in vitro mouse embryo growth by direct cell toxicity. However, the mechanism involved has not been clearly established. In the present study, this question was addressed using the embryonic stem (ES) cell model. It was found that IFN-, induces a dose-dependent apoptosis in ES cells, as assessed by trypan-blue staining, by Annexin-V labeling and DNA analysis. Moreover, IFN-, treatment cooperates with Fas-mediated apoptosis, a phenomenon that has been recently reported. As Bcl-2 oncoprotein functions as a death repressor molecule in an evolutionarily conserved cell death pathway, its expression was analyzed by flow cytometry. It was demonstrated that Bcl-2 is expressed in ES cells. When compared to untreated ES cells, IFN-,-treated, apoptotic cells expressed a lower Bcl-2 level and a normal level of Fas, whereas surviving cells expressed a normal level of Bcl-2 but a lower Fas expression. Altogether, these data suggest that IFN-, may influence early mouse embryo development by promoting apoptosis, which may constitute a novel mechanism of IFN-, embryotoxicity. [source]

    Medaka Oct4 is expressed during early embryo development, and in primordial germ cells and adult gonads

    Ana V. Sánchez-Sánchez
    Abstract Oct4 is a crucial transcription factor for controlling pluripotency in embryonic stem cells and the epiblast of mouse embryos. We have characterized the expression pattern of medaka (Oryzias latipes) Ol-Oct4 during embryonic development and in the adult gonads. Genomic analysis showed that Ol-Oct4 is the ortholog of zebrafish spg/pou2. However, their expression patterns are not the same, suggesting that Oct4 may play different roles in zebrafish and medaka. Using specific antibodies for the Ol-Oct4 protein, we showed that Ol-Oct4 is also expressed in primordial germ cells, in the spermatogonia (male germ stem cells), and during different stages of oocyte development. These results suggest that Ol-Oct4 plays a post-embryonic role in the maturing gonads and gametes. The Ol-Oct4 mRNA and protein expression patterns are similar to those of mammalian Oct4 and introduce medaka fish as a valid model for the functional and evolutionary study of pluripotency genes in vivo. Developmental Dynamics 239:672,679, 2010. © 2009 Wiley-Liss, Inc. [source]

    MicroRNA expression during chick embryo development

    Diana K. Darnell
    Abstract MicroRNAs (miRNAs) are small, abundant, noncoding RNAs that modulate protein abundance by interfering with target mRNA translation or stability. miRNAs are detected in organisms from all domains and may regulate 30% of transcripts in vertebrates. Understanding miRNA function requires a detailed determination of expression, yet this has not been reported in an amniote species. High-throughput whole mount in situ hybridization was performed on chicken embryos to map expression of 135 miRNA genes including five miRNAs that had not been previously reported in chicken. Eighty-four miRNAs were detected before day 5 of embryogenesis, and 75 miRNAs showed differential expression. Whereas few miRNAs were expressed during formation of the primary germ layers, the number of miRNAs detected increased rapidly during organogenesis. Patterns highlighted cell-type, organ or structure-specific expression, localization within germ layers and their derivatives, and expression in multiple cell and tissue types and within sub-regions of structures and tissues. A novel group of miRNAs was highly expressed in most tissues but much reduced in one or a few organs, including the heart. This study presents the first comprehensive overview of miRNA expression in an amniote organism and provides an important foundation for investigations of miRNA gene regulation and function. Developmental Dynamics 235:3156,3165, 2006. © 2006 Wiley-Liss, Inc. [source]

    Platypus Pou5f1 reveals the first steps in the evolution of trophectoderm differentiation and pluripotency in mammals

    Hitoshi Niwa
    SUMMARY Uterine nourishment of embryos by the placenta is a key feature of mammals. Although a variety of placenta types exist, they are all derived from the trophectoderm (TE) cell layer of the developing embryo. Egg-laying mammals (platypus and echidnas) are distinguished by a very short intrauterine embryo development, in which a simple placenta forms from TE-like cells. The Pou5f1 gene encodes a class V POU family transcription factor Oct3/4. In mice, Oct3/4 together with the highly conserved caudal -related homeobox transcription factor Cdx2, determines TE fate in pre-implantation development. In contrast to Cdx2, Pou5f1 has only been identified in eutherian mammals and marsupials, whereas, in other vertebrates, pou2 is considered to be the Pou5f1 ortholog. Here, we show that platypus and opossum genomes contain a Pou5f1 and pou2 homolog, pou2-related, indicating that these two genes are paralogues and arose by gene duplication in early mammalian evolution. In a complementation assay, we found that platypus or human Pou5f1, but not opossum or zebrafish pou2, restores self-renewal in Pou5f1 -null mouse ES cells, showing that platypus possess a fully functional Pou5f1 gene. Interestingly, we discovered that parts of one of the conserved regions (CR4) is missing from the platypus Pou5f1 promoter, suggesting that the autoregulation and reciprocal inhibition between Pou5f1 and Cdx2 evolved after the divergence of monotremes and may be linked to the development of more elaborate placental types in marsupial and eutherian mammals. [source]

    Microevolutionary support for a developmental hourglass: gene expression patterns shape sequence variation and divergence in Drosophila

    Tami Cruickshank
    SUMMARY A central goal of evolutionary developmental biology (Evo-Devo) is to synthesize comparative molecular developmental genetics and its description of the dynamic relationship between genotype and phenotype with the microevolutionary processes (mutation, random drift, and selection) of population genetics. To this end, we analyzed sequence variation of five gene classes that act sequentially to shape early embryo development in Drosophila: maternal, gap, pair-rule, segment polarity, and segment identity genes. We found two related patterns: (1) a microevolutionary pattern, wherein relative sequence variation within species is 2- to 3-fold higher for maternal-effect genes than for any other gene class; and, (2) a macroevolutionary pattern, wherein the relative sequence divergence among species for maternal-effect genes is 2- to 4-fold greater than for any other gene class. Both patterns are qualitatively and quantitatively consistent with the predictions of microevolutionary theory. Our findings connect within-species genetic variation to between-species divergence and shed light on the controversy over the existence of a "developmental hourglass," where mid-embryonic stages are more evolutionarily constrained than either earlier or later stages. Because maternal-effect genes experience relaxed selective constraint relative to zygotic-effect genes, they explore a wider mutational and phenotypic space. As a result, early acting maternal-effect genes diverge more widely across taxa and thereby broaden the base of the developmental hourglass. In contrast, later acting zygotic genes are relatively more constrained and limited in their diversification across taxa, narrowing the waist of the developmental hourglass. This pattern is obscured by genes with both maternal and zygotic expression, which experience the strongest evolutionary constraint. [source]

    Molecular characterization of artemin and ferritin from Artemia franciscana

    FEBS JOURNAL, Issue 1 2003
    Tao Chen
    Embryos of the brine shrimp, Artemia franciscana, exhibit remarkable resistance to physiological stress, which is temporally correlated with the presence of two proteins, one a small heat shock/,-crystallin protein termed p26 and the other called artemin, of unknown function. Artemin was sequenced previously by Edman degradation, and its relationship to ferritin, an iron storage protein, established. The isolation from an Artemia expressed sequence tag library of artemin and ferritin cDNAs extends this work. Artemin cDNA was found to contain an ORF of 693 nucleotides, and its deduced amino-acid sequence, except for the initiator methionine, was identical with that determined previously. Ferritin cDNA is 725 bp in length with an ORF of 516 nucleotides. Artemin amino-acid residues 32,185 are most similar to ferritin, but artemin is enriched in cysteines. The abundance of cysteines and their intramolecular spatial distribution suggest that artemin protects embryos against oxidative damage and/or that its function is redox regulated. The conserved regions in artemin and ferritin monomers are structurally similar to one another and both proteins assemble into oligomers. However, modeling of the quaternary structure indicated that artemin multimers lack the central space used for metal storage that characterizes ferritin oligomers, implying different roles for this protein. Probing of Northern blots revealed two artemin transcripts, one of 3.5 kb and another of 2.2 kb. These transcripts decreased in parallel and had almost disappeared by 16 h of development. The ferritin transcript of 0.8 kb increased slightly during reinitiation of development, then declined, and was almost completely gone by 16 h. Clearly, the loss of artemin and ferritin during embryo development is due to transcriptional regulation and proteolytic degradation of the proteins. [source]

    Lymphatics at the crossroads of angiogenesis and lymphangiogenesis

    JOURNAL OF ANATOMY, Issue 6 2004
    Claudio Scavelli
    Abstract The lymphatic system is implicated in interstitial fluid balance regulation, immune cell trafficking, oedema and cancer metastasis. However, the sequence of events that initiate and coordinate lymphatic vessel development (lymphangiogenesis) remains obscure. In effect, the understanding of physiological regulation of lymphatic vasculature has been overshadowed by the greater emphasis focused on angiogenesis, and delayed by a lack of specific markers, thereby limiting this field to no more than a descriptive characterization. Recently, new insights into lymphangiogenesis research have been due to the discovery of lymphatic-specific markers and growth factors of vascular endothelial growth factor (VEGF) family, such as VEGF-C and VEGF-D. Studies using transgenic mice overexpressing VEGF-C and VEGF-D have demonstrated a crucial role for these factors in tumour lymphangiogenesis. Knowledge of lymphatic development has now been redefined at the molecular level, providing an interesting target for innovative therapies. This review highlights the recent insights and advances into the field of lymphatic vascular research, outlining the most important aspects of the embryo development, structure, specific markers and methods applied for studying lymphangiogenesis. Finally, molecular mechanisms involved in the regulation of lymphangiogenesis are described. [source]

    Cryopreservation of fish sperm: applications and perspectives

    E. Cabrita
    Summary Cryopreservation is of interest not only for fish farming but also for the conservation and genetic improvement of resources. This technique has been well established in some freshwater fish species mainly, salmonid, sturgeons and carps, however, only in the last decade research was focused in marine fish species. The benefits of sperm cryopreservation include: (i) synchronization of gamete availability of both sexes, (ii) sperm economy; (iii) simplification of broodstock management, (iv) transport of gametes from different fish farms, and (v) germplasm storage for genetic selection programs or conservation of species. These issues would certainly benefit the aquaculture industry. The tremendous impact that biotechnology is having in aquaculture has been particularly obvious in recent years. Several species are being used as research models not only for aquaculture development applications but also for medical research. Sperm cryopreservation can give an important contribution in the germ storage of all transgenic lines. However, in all applications in fish sperm, cryopreservation needs to overcome a lack in standardization of methodologies and procedures, a correct assay of seminal quality and the development of tools to characterize cryoinjury. Many efforts have recently been made in the study of DNA using different approaches such as the comet assay (single cell gel electrophoresis), TUNEL (terminal deoxynucleotidyl transferase-nick-end-labelling), SCSA (sperm chromatin structure assay) and the analysis of specific DNA sequences using RT-PCR, since DNA damage may impair fertility or embryo development. Cryopreservation of gametes would certainly benefit from a higher concern on male improvement, basically through nutrition or selection of resistant stocks (e.g. stress resistant individuals or highly adapted to captivity) producing gametes of higher quality. There is a huge window of opportunities for improve the resistance of cells to cryopreservation through diet supplementation of certain compounds such as amino acids (taurine and hypotaurine), vitamins (Vit. E and C) and lipids or through a direct supplementation of the extender media. An equilibrium of those compounds will improve spermatozoa and seminal plasma composition protecting cells against oxidative stress (lipid peroxidation, protein oxidation, DNA fragmentation, enzyme protection) that is gaining each day more importance in cryodamage research. [source]

    Developmental biology of medaka fish (Oryzias latipes) exposed to alkalinity stress

    Z. L. Yao
    Summary Alkalinity stress is common in cultured aquatic animals and considered to be one of the major stress factors for fishes when they are transferred to saline-alkali waters. To evaluate potential effects of alkalinity on the developmental biology of Oryzias latipes, fertilized eggs, larvae and breeding fish were exposed to different carbonate alkalinity concentrations of 1.5,64.5 meq l,1, for 9, 120, and 60 days, respectively. The mortality of embryos significantly increased when exposed to the high concentrations (16.5,64.5 meq l,1). Although more than 50% of survived embryos hatched in 16.5 and 31.4 meq l,1 concentrations of carbonate alkalinity, most were not able to swim up after hatching. Morphological abnormalities such as coagulated embryos, halted embryo development, and hatching failure were observed at stages 15, 29,33 and 38 in high concentrations (31.4, 64.5 meq l,1). Almost all larvae in 16.5 and 31.4 meq l,1 treatments died 70 d post-hatch. Growth of juveniles exposed to carbonate alkalinity of 5.3 and 8.8 meq l,1 was not significantly different at 70 d and 120 d post-hatch. The number of eggs released by breeders, the fertilization rate and the hatching rate of eggs were significantly lower in the 31.4 meq l,1 treatment than in other treatments. Although medaka are capable of surviving in high alkalinities (31.4, 64.5 meq l,1) for an extended period of time, these conditions are stressful to the fish, especially at the embryonic and reproductive stages. [source]

    The effect of different kinds of electrolyte and non-electrolyte solutions on the survival rate and morphology of zebrafish Danio rerio embryos

    F. Lahnsteiner
    The effect of electrolyte and non-electrolyte solutions on the survival and on the morphology of zebrafish Danio rerio embryos was investigated. Embryos in different ontogenetic stages were incubated in electrolyte (NaCl, KCl, MgCl2 and CaCl2) and non-electrolyte solutions [sucrose and polyvinylalcohol (PVA)] of different concentrations for 5 , 15 min. The embryos were hatched to the long-pec stage and the effective concentrations which caused a 50% decrease in embryo development (EC50) were determined. The morphometric changes, which were caused by the test solutions, were measured. Ion channel blockers were used to see if active ion transport played a role for embryo survival. Finally, dechorionated embryos were exposed to the test solutions to get indications about the importance of chorion and perivitelline space. For 12 hours post fertilization (hpf) embryos and a 15 min exposure period, EC50 was highest for MgCl2 (1·60 mol l,1), followed by sucrose (0·73 mol l,1), NaCl (0·49 mol l,1), KCl (0·44 mol l,1), CaCl2 (0·43 mol l,1) and PVA [0·0005 mol l,1 (2·2%)]. EC50 were lower for early embryonic stages than for advanced stages for all solutions with exception of MgCl2 and sucrose. At the EC50, MgCl2 and CaCl2 solutions did not induce morphometric changes. NaCl and sucrose solutions induced reversible morphometric changes, which were compensated within 10 min. Only the EC50 of KCl and PVA solutions induced permanent morphometric changes, which could not be compensated. Incubation of embryos in electrolyte and non-electrolyte solutions together with ouabain (blocker of Na+, K+ ATPase), HgCl3 (dose-dependent inhibition of aquaporine channels), verapamil (inhibition of calcium and magnesium uptake) and amiloride (inhibition of sodium uptake) significantly decreased the per cent of embryos developing to the long-pec stage in comparison to the same solutions without blockers. Ouabain and HgCl3 also induced morphometric changes. For dechorionated embryos the survival rates in water and in the different test solutions were similar to untreated embryos. [source]

    Embryo developmental events and the egg case of the Aleutian skate Bathyraja aleutica (Gilbert) and the Alaska skate Bathyraja parmifera (Bean)

    G. R. Hoff
    Embryo development events were correlated with egg-case changes for the Aleutian skate Bathyraja aleutica and the Alaska skate Bathyraja parmifera. Yolk absorption underwent two phases: that of steady absorption during early development and that of rapid yolk absorption during the final development stages. Total length (LT) for 50% of the pre-hatching embryos egg-case jelly disappearance was 92·04 mm (range 81,102 mm) and 99·36 mm (range 81,100 mm) for B. aleutica and B. parmifera, respectively, allowing the inner chamber to open to seawater flow. The tail filament underwent three phases of growth: rapid elongation during early development (<100 mm embryo LT), stasis of tail filament length during the remainder of embryo development and rapid absorption soon after hatching. Complete tail filament development coincided with the disappearance of egg-case jelly. Clasper buds first developed at embryos >70 mm LT for both species and the sex ratio was 1:1 well before hatching. Egg cases that were devoid of an ova or developing embryo were c. 5·0 and 6·5% of the egg cases examined for B. aleutica and B. parmifera, respectively. Measurements showed that egg cases containing only egg jelly were smaller in both width and length than those possessing an ova. Embryo stages were punctuated with distinct events that correlated with egg case changes controlling the internal environment of the developing embryo. [source]

    The effect of elevated oocyte triiodothyronine content on development of rainbow trout embryos and expression of mRNA encoding for thyroid hormone receptors

    J. C. Raine
    The ability of developing rainbow trout Oncorhynchus mykiss embryos to compensate for elevated oocyte triiodothyronine (T3) content and whether elevation of oocyte T3 content within a physiologically meaningful range affects growth rates of the embryo or the expression of genes encoding for thyroid hormone receptors ,(TR,) and ,(TR,) were examined. Oocytes were immersed in ovarian fluid alone (control) or T3 -enriched ovarian fluid prior to fertilization and water hardening, to induce a dose-dependant increase in oocyte T3 content of c. 3 (control), c. 30 (LT3) or c. 110 ng egg,1(HT3). To examine the interaction of embryo somatic growth with altered thyroid state more effectively, the embryos were reared at two ambient temperatures (8·5 and 5·5°C ) to induce different growth rates. A significant decline in whole embryo T3 content was measured in the T3 -treatment groups reared at both water temperatures by 3 weeks post-fertilization (dpf), and may have reflected the action of outer ring monodeiodinase, which was present in microsomes prepared from embryos 23 dpf. Whole embryo T3 levels in the HT3 group, however, remained higher than controls until phase 2 of development [the onset of endogenous thyroid hormone (TH) release]. This suggested that the embryos exerted some control over their response to exogenous TH, but that there was a limit to the level of control exerted by the embryonic tissues. Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed the presence of mRNA encoding for the two TR isoforms as early as 26 dpf, and quantitative real-time RT-PCR (qPCR) was used to examine the effect of elevated oocyte T3 content on the expression of these TR genes in embryos raised at 8·5 and 5·5° C, and sampled at similar developmental stages prior to the onset of embryonic TH synthesis, to ensure that the oocyte T3 was the only source of TH exposure to the embryo. There was a suppression of the TR, gene expression in the control 5·5° C group relative to the control 8·5° C group. In addition, both TR, and TR, mRNA accumulation was lower, relative to the controls, in the LT3 treatment group reared at 8·5° C suggesting a suppressive effect of the lower level of T3 treatment on the TR gene expression. Conversely, there were no differences from controls in the HT3 treatment group, possibly indicating that this level of exposure overrides the down-regulating capacity of the embryo. Similar patterns were seen for TR, and TR, mRNA accumulation in embryos reared at 5·5° C, but because of the temperature suppressed level of TR, mRNA in the controls, significant affects of the LT3 treatment were only found for TR,. There were no measurable effects of T3 treatment on oocyte fertility or embryo somatic growth for either temperature treatment group, nor was somatic growth hormone content (measured only in the 8·5° C treatment group) apparently related to in ovo T3 levels. The results suggest that altered in ovo T3 levels, within the ranges used here, do not induce marked affects on embryo development, probably because of the ability of the embryo to maintain the integrity of its TH milieu. [source]

    The effects of 1.5T magnetic resonance imaging on early murine in-vitro embryo development

    MMed (O&G), MRACOG, Stephen Chew MBBS
    Abstract Although no ionizing radiation is involved, patients undergoing magnetic resonance imaging (MRI) are exposed to powerful static magnetic fields, magnetic field gradients, and radio-frequency fields that may be potentially damaging. Our study aims to document the effect of MRI imaging sequences on early murine embryo development (two-cell to blastocyst stage) in vitro. Two-cell murine embryos were exposed to various lengths of MRI using pulse sequences employed in present day clinical imaging. Early murine embryo development was documented in vitro, and blastocyst development rates were computed for both the control and exposed groups. There were no significant differences detected in the rate of blastocyst formation between the control groups and the embryos exposed to MRI. J. Magn. Reson. Imaging 2001;13:417,420. © 2001 Wiley-Liss, Inc. [source]

    Characterization of an immortalized oviduct cell line from the cynomolgus monkey (Macaca fascicularis)

    H. Okada
    Abstract:, To establish reproductive biological techniques in mammals, it is important to understand the growth environment of the embryo. Oviduct epithelial cells are in close proximity to the embryo during pre-implantation development. We, therefore, established an immortalized oviduct epithelial cell line from the cynomolgus monkey, evaluated the usefulness of these cells as feeder cells for embryo culture, and investigated the gene expression of several growth factors and cytokines in the cells. The immortalized cells were positive for the anti-cytokeratin antibody, as determined by immunocytochemistry, indicating that they are epithelial. They also expressed oviductin, which is specific to oviduct epithelial cells, glyceraldehyde-3-phosphate dehydrogenase (control), leukemia inhibitory factor, vascular endothelial growth factor, epidermal growth factor, insulin-like growth factor 1, transforming growth factor beta-2, and interleukin 4. Mouse embryo development was improved when the immortalized cells were used as feeder cells. This cell line is also useful for studying the factors secreted by oviduct epithelial cells. [source]

    The significance of platelet-activating factor and fertility in the male primate: a review

    William E. Roudebush
    Abstract:, Since its discovery nearly 30 years ago platelet-activating factor (PAF) has emerged as one of the more important lipid mediators known. PAF (1- O -alkyl-2- O -acetyl- sn -glycero-3-phosphorylcholine) exists endogenously as a mixture of molecular species with structural variants of the alkyl moiety. PAF is a novel potent signaling phospholipid that has unique pleiotropic biological properties in addition to platelet activation. PAF also plays a significant role in reproduction and is present in the sperm of a number of primate species. PAF content in squirrel monkey sperm is significantly higher during the breeding season than the non-breeding season. PAF content in rhesus sperm has a significant relationship with sperm motility. PAF content in human sperm has a positive correlation with seminal parameters and pregnancy outcomes. The enzymes (lyso-PAF-acetyltransferase and PAF-acetylhydrolase) necessary for PAF activation and deactivation are present in primate sperm. PAF-acetylhydrolase may act as a ,decapacitation factor'. Removal of this enzyme during capacitation promotes PAF synthesis increasing primate motility and fertilization. PAF also plays a significant role in the fertilization process, enhancing the fertilization rates of oocytes. Enhanced embryo development has also been reported in oocytes fertilized with PAF-treated sperm. Exogenous PAF will also significantly improve primate artificial insemination pregnancy outcomes. PAF antagonists inhibit sperm motility, acrosome reaction, and fertilization thus suggesting the presence of receptors for PAF. The PAF-receptor is present on primate sperm, with altered transcript levels and distribution patterns on abnormal cells. Whereas, the exact mechanism of PAF in sperm function and reproduction is uncertain, its importance in normal primate fertility is substantial. [source]

    Yin yang 1 directly regulates the transcription of RE-1 silencing transcription factor

    Lichun Jiang
    Abstract The RE-1 silencing transcription factor (REST) is a master transcription factor that plays a critical role in embryo development, especially during the process of neurogenesis and neural plasticity. However, the mechanism of REST gene transcription regulation is still an open question. Here, by combining bioinformatics analysis and experimental studies, we report that the transcription factor Yin Yang 1 (YY1) bound to a conserved YY1 binding site in the promoter of the mouse REST gene and positively regulated activity of this promoter in SH-SY5Y cells. Furthermore, analysis of microarray data revealed a significant correlation between the expression of YY1 and REST genes. Overall, this study suggests that YY1 directly regulates expression of the REST gene. © 2007 Wiley-Liss, Inc. [source]

    Estrogen administration during superovulation increases oocyte quality and expressions of vascular endothelial growth factor and nitric oxide synthase in the ovary

    Choong-Sik Ha
    Abstract Aims:, This study investigated whether estrogen administration during superovulation enhances oocyte quality using a mice model. We also investigated whether this estrogen treatment regulates the expressions of angiogenic factors, such as vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS), in the ovary. Method:, Female mice were co-injected with various doses of estrogen (1 µM, 10 µM and 100 µM) and pregnant mare serum gonadotrophin during superovulation, followed by human chorionic gonadotrophin injection 48 hours later. Then they were mated with individual males. After 18 hours, zygotes were flushed and cultured to blastocyst. The expression of VEGF and eNOS in the ovary was examined using Western blot and immunohistochemistry. The control group was superovulated without estrogen. Results:, Both numbers of ovulated zygotes and the rate of embryo development to blastocyst were significantly increased in the 1-µM estrogen dose compared to the control group. VEGF and eNOS expressions were stimulated by estrogen treatment. In particular, VEGF expression was significantly increased at 1-µM estrogen concentration, whereas, eNOS expression was significantly increased in all estrogen concentrations compared to controls. Conclusions:, The study showed that estrogen co-injection during superovulation increased the ovarian response, embryo developmental competence and expressions of VEGF and eNOS in the ovary. [source]

    Alcohol-Induced Lipid and Morphological Changes in Chick Retinal Development

    ALCOHOLISM, Issue 5 2004
    Yolanda Aguilera
    Abstract: Background: Alcohol exposure causes alterations in the lipid content of different organs and a reduction of long-chain fatty acids. During embryo development, the central nervous system is extremely vulnerable to the teratogenic effects of alcohol, and the visual system is particularly sensitive. Methods: White Leghorn chick embryos were injected with 10- and 20-,l alcohol doses into the yolk sac at day 6 of incubation. The lipid composition of the retina was analyzed in embryos at day 7 of incubation (E7), E11, E15, and E18. The percentages of phospholipids, free cholesterol, esterified cholesterol, diacylglycerides, and free fatty acids were estimated by using an Iatroscan thin layer chromatography flame ionization detector. Gas chromatography and mass spectrometry were used to determine fatty acid composition. The morphological study was performed at E7, E11, and E19 by means of semithin and immunohistochemical techniques. Results: In the retina, alcohol causes the total lipid content to change, with a remarkable increase in free cholesterol and a dramatic decrease in esterified cholesterol. Diacylglycerides and free fatty acids tend to increase. Phosphatidylcholine and phosphatidylethanolamine decrease, whereas phosphatidylserine, sphingomyelin, and phosphatidylinositol increase. The main fatty acids of the retina also undergo changes. At E7, myriotic acid increases, and oleic acid and polyunsaturated fatty acids such as arachidonic acid and docosahexaenoic acid decrease. From E18 onward, there is some recovery, except for fatty acids, which recover earlier. From a morphological point of view, alcohol effects on retinal development are various: increase of intercellular spaces in all cell layers, pyknosis with loss of cellularity in the inner nuclear cell layer and ganglion cell layer, retarded or disorderly cell migration, early cell differentiation, and loss of immunoreactivity for myelin oligodendrocyte,specific protein. Conclusions: Acute alcohol exposure during embryo development causes the lipid composition of the retina to change, with a trend to recovery in the last stages. These alterations are in line with the changes observed at a morphological level. [source]