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PubMed1981

p-Chloro-N-methylaniline demethylase activity in liver and extrahepatic tissues of the human fetus and in chick embryo liver.

We achieved a 50-fold increase in sensitivity of a fluorescence assay for p-chloro-N-methylaniline (p-CNMA) demethylase enabling us to examine the kinetics of the reaction in chick embryo liver and human fetal tissues and its inducibility in chick embryo liver. In chick embryo liver, p-CNMA demethylase activity was maximally induced (to about 200% of control levels) by both phenobarbital and beta-naphthoflavone. p-CNMA demethylase is therefore nonselective in its response to inducers of the P-450 and P-448 classes, respectively. Line-weaver-Burk plots of the data suggest that single kinetic components predominated in the reaction for both control and induced livers (mean apparent KmS: control, 3.8 X 10(-5) M; phenobarbital treated, 3.8 X 10(-5) M). Differences in the KmS and the pH-activity curves for the phenobarbital- and beta-naphthoflavone-induced enzymes suggested, however, that the induced enzymes were not identical. p-CNMA demethylase activity was measurable in liver and extrahepatic organs of the human fetus and was 1.5 times higher in human fetal liver than in chick embryo liver. At least two kinetic components participated in the reaction in the human liver. Reaction rates in human fetal adrenal, lung and brain were about 60, 2 and 1% of the liver rates, respectively. Detection of mixed function oxidase activity in human fetal brain, not usually considered a site of biotransformation, suggests that the fetal brain could be a site of cytotoxic activation of chemicals during human fetal development.

Animals↗

PubMed1999

SDS/PAGE characteristics of protein kinases tightly associated with chick embryo brain ribosomes.

Protein kinases tightly associated with chick embryo brain ribosomes washed with Triton X-100 and KCl were characterized by their ability to phosphorylate ribosomes and two exogenous substrates, histone IIA and casein. c-AMP-dependent kinase (PKA) and casein kinases (CK1, CK2) were examined in the presence of specific modulators by SDS/PAGE followed by renaturation in gel assay according to Kameshita & Fujisawa (Anal. Biochem. 1989, 183, 139-143). Basing on these data it can be presumed that PKA activity increases, but the levels of CK2 and CK1 decrease during chick embryo development.

Animals↗

PubMed1983

Drug withdrawal prior to hatch in the morphine tolerant chick embryo.

Morphine tolerance and dependence were produced in chick embryos by injecting 20 mg/kg into the air space of the egg daily from incubation day 12. Starting on day 16, two groups of eggs were withdrawn from morphine by either substituting water or by treating with naloxone. Chick embryo activities were monitored on incubation day 18. Activities of the embryos withdrawn from morphine did not differ from control, but naloxone injection produced a substantial increase in activity. Neither regimen of morphine withdrawal affected the hatchability of the eggs. When these chicks were 1 day old, the effect of 1 mg morphine/kg on the rate of distress vocalization was measured. The withdrawn chicks were 1 day old, the effect of 1 mg morphine/kg on the rate of distress vocalization was measured. The withdrawn chicks vocalized at a rate that was intermediate between that of controls and that of nonwithdrawn chicks indicating that tolerance was still present one week after the last morphine injection.

Animals↗

PubMed1989

A simple method for shell-less cultivation of chick embryos.

The chorio-allantoic membrane of explanted chick embryos can be used for the study of vascular development and of the vascular response to drugs and to tissue implants or tissue extracts. Three different types of vials for shell-less cultivation of chick embryos were compared in terms of short-term survival. Plastic cups with rounded bottoms were superior to Petri dishes and polyethylene film bags, in that they combined a high survival rate of the embryos with a possibility of using a large number of embryos in each experiment. With the use of plastic cups this method is simple, reliable and inexpensive.

Allantois↗

PubMed2004

Nervous system development in normal and atresic chick embryo intestine: an immunohistochemical study.

Intestinal motility disorders are a common complication after surgery for neonatal intestinal atresia. Although intestinal atresia causes alterations in the enteric nervous system, especially in its inner structures (nervous fibers in the mucosa, submucous and deep muscular plexuses), how these alterations develop is unclear. The chick model is a useful research tool for investigating the ontogenesis of the enteric nervous system and the pathogenesis of congenital bowel diseases. More information is needed on the overlap between the developing enteric nervous system and intestinal atresia. Because vasoactive intestinal polypeptide and substance P are typical intestinal neuropeptides, and vasoactive intestinal polypeptide acts as a modulator in neurodevelopment and an inhibitor of smooth muscle cell proliferation, our aim in this study was to investigate the distribution of their immunoreactivity in the developing enteric nervous system of normal and experimental chick models. We studied gut specimens excised from normal chick embryos (aged 12-20 days) and experimental chick embryos (aged 15-20 days) that underwent surgical intervention on day 12 to induce intestinal atresia (atresic embryos) or simply to grasp the bowel loop (sham-operated embryos). In normal chick embryos we showed vasoactive intestinal polypeptide and substance P immunoreactivity from day 12 in the submucous and myenteric plexuses. The distribution of peptide immunoreactivity differed markedly in atresic and normal or sham-operated gut embryos. These differences especially affected the inner structures of the enteric nervous system of specimens proximal to atresia and were related to the severity of dilation. Because nerve structures in the gut wall mucosa and submucous and deep muscular plexuses play a role in motility control and stretch sensation in the intestinal wall, our findings in the chick embryo may help to explain how gut motility disorders develop after surgery for neonatal intestinal atresia.

Animals↗

PubMed1983

Transcriptional control of the fibronectin gene in chick embryo fibroblasts transformed by Rous sarcoma virus.

Rous sarcoma virus transformed-chick embryo fibroblasts contain decreased amounts of the adhesive protein fibronectin that results from a lowering of fibronectin mRNA levels (Fagan, J.B., Sobel, M.E., Yamada, K.M., de Crombrugghe, B., and Pastan, I. (1981) J. Biol. Chem. 256, 520-525). In the present study, nuclei isolated from chick embryo fibroblasts (CEF) and Rous sarcoma virus transformed-chick embryo fibroblasts (RSV-CEF) were used to measure relative rates of fibronectin RNA synthesis. [32P]RNA was synthesized by preinitiated endogenous RNA polymerases in nuclei, and the radioactive RNA was subsequently hybridized to DNA fragments spanning approximately 40 kilobase pairs (approximately 85%) of the fibronectin gene. Transformation of CEF by RSV resulted in a 7- to 8-fold reduction in the amount of 32P-labeled nuclear RNA hybridizing with fibronectin DNA sequences when the nuclei were incubated for either 3.5 or 45 min. These results indicate that decreased transcription and not reduced RNA stability plays a major role in lowering fibronectin mRNA levels in RSV-CEF. When nuclear RNA was transcribed in the presence of Sarkosyl, diminished RNA synthesis was still observed suggesting that reduced fibronectin transcription in RSV-CEF is not due to a block in RNA chain elongation. Our data indicate that transcriptional regulation of the fibronectin gene is an important mechanism of lowering steady state levels of fibronectin mRNA in RSV-CEF.

Animals↗

PubMed1991

Ectopic colonization of primordial germ cells in the chick embryo lacking the gonads.

Chick primordial germ cells (PGCs) first appear in the extraembryonic region in the early embryo, then temporarily circulate via the blood vascular system and finally migrate into the gonadal anlagen. In the present study, we examined the trend of ectopic distribution of PGCs in the chick embryo when its future gonadal region had been removed at an early stage. Embryos at stage 10, from which the caudal third region was excised, were incubated until they reached stages 14 to 20. In embryos at stage 14, about 80% of the total PGCs were found in the capillaries of the yolk sac, whereas others were observed in the head, mainly in the mesenchyme and small vessels close to the neural tube. From stage 18 onward, many PGCs accumulated in the embryo proper; about 90% of them colonized in the head region around the neural tube. These ectopic PGCs in the head were found in the capillaries, sometimes as thrombi or emerging from them into the adjacent mesenchyme. These results show that, when the chick embryo lacked gonads, the PGCs could be concentrated in the head region and migrated from the capillaries into the mesenchyme.

Animals↗

PubMed1975

Marek's disease virus (Kekava strain) replication in chickens, chick embryos and cell cultures.

In the course of 12 passages of Marek's disease virus (MDV) strain Kekava (MDV-Kekava) in chickens, the morbidity varied greatly (from 23 to 50 percent). MDV-Kekava produced plaques in cultures of chick embryo kidney and adult chicken kidney cells and chick embryo fibroblasts (CEF). The virus adaptation to the cultures was very slow. MDV-Kekava induced the formation of pocks on the chorioallantoic membranes (CAM) of chick embryos but the proportion of embryos with CAM lesions did not exceed 24 percent. Serial passaging of the virus in chick embryos beyond the 5th passage was unsuccessful. The results of virus isolation in chickens, cell cultures and chick embryos indicate the possibility of a long-term latent virus carrier state in chickens without development of tumours.

Animals↗

PubMed1993

Developmental regulation of insulin-like growth factor binding protein-2 in chick embryo serum and vitreous humor.

The chick embryo is a useful vertebrate model for studying developmental embryogenesis. Insulin-like growth factor I (IGF-I), a potent mitogen, is thought to contribute to the general growth of the embryo as an endocrine factor, and as a paracrine factor to the development of the early embryo and of specific organs such as the eye. Recent data suggest that a family of at least six IGF binding proteins (IGFBPs) complex IGF-I and modulate its biological actions. In the present study, we examine the expression of IGFBPs in chicken serum and vitreous humor at different stages of embryonic development, and compare it with that of IGF-I. As determined by ligand blotting, the predominant IGFBP in chick serum and vitreous humor between embryonic days 4 and 22 (E4-E22) is a 30 kDa IGFBP. This IGFBP was specifically immunoprecipitated by a polyclonal antiserum raised against rat IGFBP-2, the predominant IGFBP in fetal human and rat serum. Although IGFBP-2 is present in both chick fluids at all times examined, serum IGFBP-2 increased progressively between E10-E22, whereas vitreous IGFBP-2 was highest during eye organogenesis (E4-E8). This suggests that vitreous IGFBP-2 is synthesized locally. Like serum IGFBP-2, levels of immunoreactive IGF-I in serum are higher in the second week of embryogenesis than the first. Despite this correlation, changes in IGFBP-2 do not appear to be regulated by IGF-I: (a) serum IGF-I decreases after day 15, whereas IGFBP-2 levels remain stable until hatching; (b) vitreous IGF-I, like serum IGF-I, is higher in the second week of embryogenesis, whereas vitreous IGFBP-2 is highest in the first week; (c) embryos cultured ex ovo express IGFBP-2 at E15-E19, although they lack the normal mid-embryogenesis surge in IGF-I. We conclude that vitreous IGFBP-2 is synthesized locally in the eye, and that the expression of IGFBP-2 in chick embryos is not directly regulated by IGF-I.

Animals↗

PubMed1982

Acid and alkaline phosphatase activity in migrating primordial germ cells of the early chick embryo.

Little information is available concerning enzyme activity in primordial germ cells (PGCs) of the early chick embryo. The present study is designed to examine the disposition of alkaline and acid phosphatase activity in the PGCs during their migration into the developing gonads of the early chick embryo. White Leghorn chick embryos were sacrificed at daily intervals from 1 to 6 days of incubation. Following sacrifice the embryos were fixed, dehydrated, and embedded in glycol methacrylate (GMA). Alkaline and acid phosphatases were demonstrated by the simultaneous diazo-coupling method. The embryonic tissues at the different ages were examined for PGCs and the histochemical reactions for alkaline and acid phosphatases in these cells evaluated. Acid phosphatase activity did not appear within PGCs until 3 days of incubation, and then in only a few PGCs in the active phase of their migration in the dorsal mesentery, suggesting that there is no large wave of degeneration of these cells during migration. Alkaline phosphatase activity was observed as early as 2 days of incubation in PGCs during the passive phase of their migration in extraembryonic blood vessels. Alkaline phosphatase-positive PGCs in the active phase of migration were also found in the dorsal mesentery; however, the cellular localization of this enzyme differed from that observed in the passively migrating PCGs, indicating that there are alterations in the metabolic activities of these cells during the active and passive phases of migration.

Acid Phosphatase↗

PubMed1979

Fine structural demonstration of tyrosinase activity in the retinal pigment epithelium of normal and PTU-treated chick embryos.

The effect of phenylthiourea (PTU) on the retinal pigment epithelium of chick embryo was studied by using electron microscopy and electron-microscopic histochemistry. In the retinal pigment epithelium of PTU-treated, 10-day-old chick embryo, there is an abundance of premelanosomes and relative increases in the smooth endoplasmic reticulum (ER), and in the smooth-surfaced anastomosing tubular channels. Reaction products resulting from electron-microscopic histochemistry are first observed in some cisternae of the distal or active face of the Golgi apparatus and occasionally in small vesicles in the 5-day-old embryo, and these reactions become more intense with time. PTU treatment blocks melanin synthesis in the premelanosomes of 10-day-old chick embryo. Electron-microscopic histochemistry is completely inhibited by PTU treatment. Removal of PTU allows a weak positive reaction to histochemistry in some elements of the Golgi apparatus and in some small vesicles.

Animals↗

PubMed2001

Activin disrupts somitogenesis in cultured chick embryos.

The anatomical and cell biological aspects of somite formation in the chick embryo have been rather well studied. Molecular regulation of somitogenesis in vertebrates is just beginning to be understood. We have studied the effects of human recombinant activin on somitogenesis in gastrulating chick embryos cultured in vitro with a view to assessing the possible role of activin-related molecules in this phenomenon. Activin disrupted somitogenesis in treated embryos, resulting in the formation of abnormal, split or ectopic somites. Light microscopic examination indicated that the ability of activin to interfere with somitogenesis might be partly due to initiation of somite formation at ectopic sites. We show that these cells are indeed somitogenic by their expression of one of the earliest somite-specific marker genes, Pax3. Scanning electron microscopic examination of control and treated embryos revealed direct effects of activin on cell-cell interactions. Cells from treated embryos exhibited disrupted intercellular adhesion leading to large intercellular spaces, altered cell shapes and modification of cell surface protrusions. The effects of activin on somitogenesis appear to be specific, since the neural structures, which are generally more susceptible to chemical insults during gastrulation, were relatively less affected. The results clearly point to a role of activin-related molecules in somitogenesis in the chick embryo.

Activins↗

PubMedDate not supplied

Interaction of MF 50 Hz, 10 mT with high dose of X-rays: evaluation of embryotoxicity in chick embryos.

We studied the combined effects of exposure to magnetic field (MF) 50 Hz 10 mT and X-ray ionizing radiation on developing chick embryos. When chick embryos at early stages were repeatedly exposed to MF prior to X-ray radiation, reduction of X-ray teratogenicity was observed. When MF exposures started immediately after X-ray radiation, the adverse developmental effects of ionizing radiation were potentiated. Embryotoxicity was more pronounced in X-ray irradiated and MF exposed groups. We present a hypothesis about the mechanisms of MF effect based on enhancement of oxidative processes, followed by stimulation of antioxidant systems.

Animals↗

PubMed1980

Altered protein synthesis and creatine kinase in breast muscle cell cultures from dystrophic chick embryos.

The total protein synthesis (TPS), myosin synthesis (MS) and creatine kinase (CK) levels in muscle cell cultures obtained from 400 normal (strain 454) and 400 dystrophic chick embryos (strain 455) were investigated. The cultures were obtained from breast muscles of 12 day chick embryos by dissociation in 0.25% trypsin, preplating and plating of 5 x 10(5) floating cells on gelatin coated dishes in Minimal Essential Medium, 10% horse serum and 2% chick embryo extract. After 6 days, when electron-microscopic studies demonstrated good muscle differentiation, cell cultures were labeled with [3H]leucine. TPS and MS, respectively, showed 85% and 65% increases in breast muscle cell cultures from dystrophic chick embryos. The half-life times for total protein and myosin from dystrophics were 19 and 32 hr, respectively as compared with 36 and 48 hr from controls. Noncollagen protein content (NCP) showed 27% decrease in postfusion stage (12 days) of cell cultures from dystrophics. The CK level showed 30% lower values in the cells from dystrophics but 50% higher values in their culture medium. The addition of leupeptin plus pepstatin (50 microgram/ml) to these cultures resotred NCP content, total protein and myosin turnover to normal values and significantly increased TPS and MS. The addition of diphenylhydantoin (DPH) (20 microgram/ml) to cell cultures from dystrophics did not change the NCP content nor the turnover for total protein and myosin but significantly increased TPS, MS and CK while medium CK significantly decreased. The addition of leupeptin plus pepstatin or DPH to muscle cell cultures from normal chick embryos also significantly stimulated TPS and MS.

Animals↗

PubMed1984

Onset of the response to chorionic gonadotropin in the chick embryo testis.

The stage of development of the chick embryo testis when it begins to respond to gonadotropin stimulation was investigated. The testosterone secretion in vitro, measured by radioimmunoassay, was employed to evaluate the response to hCG in testis from 8 to 16 days of incubation. At 8 to 10 days of the chick embryo development, the testis secreted testosterone, but no increment in the steroid production has been observed after hCG treatment. On the contrary, at 12, 14, and 16 days a clear increase in testosterone secretion has been demonstrated when hCG was added to the culture medium. The absence of hCG response before 12 days of incubation agrees with the hypothesis of an early independence period between testis and adenohypophysis during embryonic development.

Animals↗

PubMed1980

The proliferative potential of chick embryo fibroblasts: population doublings vs. time in culture.

Chick embryo fibroblasts were maintained in a non-proliferating state up to 150 days in medium containing reduced concentrations of fetal bovine serum. Under these conditions the total time in culture was increased from 50 to 200 days while the total proliferative potential decreased only slightly. From these experiments we conclude that the ability of chick embryo fibroblasts to proliferate in vitro is much more dependent on the number of previous in vitro doublings than on the duration of the time in the culture.

Animals↗

PubMed2003

beta-Microseminoprotein-related molecules may participate in formation of the mesoderm in the chick embryo.

It has previously been shown that human beta-microseminoprotein enhances development of mesodermal structures in the chick embryo. The present study was carried out to elucidate the mechanism of action of human beta-microseminoprotein in the chick embryo. beta-Microseminoprotein brought about significant modulation of expression of Brachyury in gastrulating embryos. In approximately 50% of the treated embryos, Brachyury expression was enhanced around the Hensen's node. These cells not only expressed higher levels of Brachyury, but also appeared to switch off Brachyury expression prematurely, postinvagination. The spatial modulation of Brachyury is not clearly reflected in the northern blots, indicating that beta-microseminoprotein treatment results in redistribution of available transcripts or that the upregulation is compensated for by early switching off of Brachyury postinvagination. Because higher levels of Brachyury during gastrulation are believed to result in early exit of cells from the primitive streak, beta-microseminoprotein treatment appeared to have stimulated morphogenetic movements by upregulating Brachyury around the Hensen's node. This deduction was confirmed by scanning electron microscopic analysis that showed that altered morphogenetic movements accompany modulation of Brachyury. The specific responses elicited by beta-microseminoprotein indicate presence of a structurally related molecule in the chick. By western blotting, similar molecules were indeed detected in the chicken seminal plasma and in chick embryos. These data strongly suggest that beta-microseminoprotein-related molecule(s) participates in mesoderm formation in the chick embryo.

Animals↗

PubMed1975

The major cell surface glycoprotein of chick embryo fibroblasts is an agglutinin.

A major cell surface protein, CSP, of chick embryo fibroblasts has been shown to constitute up to 3% of total cell protein, and to be decreased after viral transformation. Its role in normal cell behavior is not known. We have isolated CSP from chick embryo fibroblasts by extraction with 1 M urea and find that these preparations of CSP agglutinate formalinized sheep erythrocytes at protein concentrations of under 2 mug/ml. In extracts of chick embryo cells, the quantity of such hemagglutinating activity parallels that of CSP determined by electrophoresis, and both are substantially decreased in chick cells transformed by the Bryan hightiter strain of Rous sarcoma virus. Both CSP and hemagglutinating activity are progressively adsorbed onto erythrocytes and can be released by 1 M urea. An antiserum to purified CSP specifically blocks the agglutination. The agglutinating activity is destroyed by boiling or treatment with proteases. The agglutination reaction is inhibited by the chelating agents EDTA and EGTA [ethyleneglycol-bis(beta-aminoethyl ether)N,N'-tetraacetic acid]. Agglutination is also inhibited to a lesser degres by amino sugars and other amines, increased osmolarity, and urea. Other monosaccharides, hyaluronidase, DNase, and RNase have little or not effect on the agglutination reaction. This demonstration that CSP has an agglutinating activity that is sensitive to proteases and that requires divalent cations suggests that this molecule may play a role in cell adhesion.

Agglutinins↗