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PubMed1984

Vascular abnormalities due to hyperthermia in chick embryos.

Intraembryonic vascular abnormalities were studied in chick embryos exposed to temperatures 3 degrees C and 4 degrees C above normal temperature (38 degrees C) from the beginning of incubation. The average duration of hyperthermia was 54 and 53 hours, respectively. Immediately after exposure, the embryos were examined with FITC-Dextran microangiography in vivo. Following hyperthermia various abnormalities in the heart, ventral aortae, aortic arches, omphalomesenteric arteries, and the distal dorsal aortae frequently occurred. There were also significant microvascular changes in the head, in the lateral and caudal parts of the embryos, and in the pellucid area of the yolk sac. In another series incubation at 41 degrees C, hyperthermia of 3 degrees C during the first 3 days of development produced several extraembryonic vascular abnormalities. These included duplication and abnormal branching of the cranial vitelline vein, absence or abnormal course of the omphalomesenteric vessels, aneurysmatic dilatation or abnormal course of the caudal vitelline vein, and aneurysmatic dilatation or occlusion of the abdominal venous sinus. Most frequent findings were blind, congested, and dilated microvascular segments in the pellucid area, commonly associated with an irregular microvascular pattern and perivascular swelling. The abnormalities described are assumed to be caused by the direct effects of hyperthermia upon the developing vessels resulting in microvascular insufficiencies, pathological leakage, and perivascular oedema. Such disturbances may have serious consequences for embryonic vascular development and microcirculation, which in turn may have adverse effects on further embryonic growth and development.

Angiography↗

PubMed1995

Phospholipid distribution and fatty acid composition in different brain regions during chick embryo development.

The phospholipid profile of different chick embryo brain regions was studied from 11 to 21 days of development, revealing interesting changes in content and distribution. Total phospholipid phosphorus (P), in micrograms of P per microgram of DNA, increases significantly during development of cerebral hemispheres (CHs), optic lobes (OLs), and brainstem (BS). Compared with CH and OL, the BS shows at all stages a significantly higher concentration of phospholipid P, which in contrast decreases in the cerebellum (CB) during development. Moreover, the data show interesting differences between the right and the left portion of the brain. The distribution of phospholipid P and the fatty acid composition of phospholipids were asymmetric between left and right OL and CH, as were the concentrations of DNA and cholesterol, demonstrating lateralized neurochemical development in these structures, i.e., left OL, right OL, left CH, and right CH. The data are discussed also in relation to the potential importance of neurochemical lateralization for determining lateralized embryonic and postnatal behavior of this species.

Animals↗

PubMed1992

Biochemical characterization of coumarin 7-hydroxylase activity in chick embryo liver microsomes.

Coumarin occurs naturally in the diet and can induce and inhibit cytochrome P450 enzymes. Hepatic coumarin 7-hydroxylase activity is the major pathway for coumarin metabolism in humans but not in rats, most strains of mice, or other laboratory animals. Coumarin 7-hydroxylase activity and the effects of chemical inhibitors and inducers on this activity were studied in 19-day-old chick embryo liver microsomes. Activity was between 35 and 75 nmol/mg protein/hr which is approximately 2-fold higher than reported for human liver microsomes. The pH optimum was 7.8 and the Km determined by both an ether extraction and a high performance liquid chromatography method was 7.3 +/- 0.9 (+/- SD) microM. Substrate inhibition was evident at coumarin concentrations above 250 microM (activities at 1000 and 4000 microM coumarin were 84 and 40% of Vmax, respectively). The Ki values (+/- SD) for inhibitors of microsomal coumarin 7-hydroxylase activity in vitro were: alpha-naphthoflavone, 46.9 +/- 19.8 nM; metyrapone, 0.8 +/- 0.9 microM; aniline, 12.3 +/- 8.2 microM; cimetidine, 70.9 +/- 27.9 microM; N-nitrosodimethylamine, 0.7 +/- 0.9 mM; and dimethyl sulfoxide, 7.9 +/- 1.9 mM. Treatment of chick embryos with pyrazole (40 mumol) increased coumarin 7-hydroxylase by 50% at 24 hr, but this activity was unaffected by treatment of embryos with 3-methylcholanthrene (2 mumol) or glutethimide (8 mumol). Thus, hepatic coumarin 7-hydroxylase activity in 19-day-old chick embryos is higher than in most laboratory animals and has similar biochemical properties as the enzyme in humans and mice. The chick embryo liver may be a useful system for studies on the biochemical effects of coumarin and the regulation of cytochrome P450-dependent coumarin 7-hydroxylase.

Animals↗

PubMed1988

Fluorescence and electron microscopic studies of the cytoskeletal organization of normal, established and transformed chick embryo cells.

The cytoskeletons of two established chick embryo cell (CEC) lines were examined by fluorescence and electron microscopy and compared with those of control cells and cells transformed by Rous sarcoma virus (RSV). In normal CEC, many stress fibers were observed. On the other hand, stress fibers were disorganized in nontransformed spontaneously established CEC, non-tumorigenic CEC partially transformed with a chemical carcinogen, and tumorigenic RSV-transformed CEC. In the normal CEC, actin filaments formed several bundles along the processes of the cell. Stereo-images of the peripheral region revealed bundles of filaments which were located along the attached side to the substrate. A fine well preserved network of filaments was also observed. On the other hand, in spontaneously established, partially transformed and RSV-transformed CEC, a fine network of filaments, but no actin cables, was found. These results support previous evidence that the cytoskeletal changes themselves are not directly related to the transformation or tumorigenicity of cells.

Actin Cytoskeleton↗

PubMed1979

Chondroitin SO4 catabolism in chick embryo chondrocytes.

An enzyme preparation from cultured chick embryo vertebral chondrocytes attacks chondroitin SO4 oligosaccharides from the nonreducing terminal in a recycling pathway involving the sequential action of a beta-glucuronidase, a 4- or a 6-sulfatase, and a beta-N-acetylgalactosaminidase. The sequence is blocked by saccharo-1,4-lactone, an inhibitor of the beta-glucuronidase, or by 2-acetamido-2-deoxy-D-galactonolactone, an inhibitor of the beta-N-acetylgalactosaminidase. The level of 4-sulfatase activity is low relative to the other activities and limits the rate of catabolism of hybrid oligosaccharide structures containing both 6-sulfated galactosamine residues and 4-sulfated galactosamine residues. This results in the accumulation of shortened oligosaccharides, most of which have galactosamine-4-SO4 residues at their nonreducing terminals. In the presence of the lactone inhibitors, polymeric chondroitin SO4 is broken down by the enzyme preparation to oligosaccharides which are 10 to 15 monosaccharides long, indicating that degradation of chondroitin SO4 chains is initiated by an endoglycosidase which generates oligosaccharide substrates for the recycling exoglycosidase system.

Acetylgalactosamine↗

PubMed1990

Glycoprotein synthesis in the subcommissural organ of the chick embryo. II. An immunochemical study.

In the chick embryo, A74 immunoaffinity chromatography allowed to purify specific glycoproteins relevant to the SCO ventricular secretory process. The eluted fractions of the subcommissural organ (SCO), the cerebral hemispheres (CH) and the medulla oblongata (MO) were compared using the Concanavalin A (Con A) and wheat germ agglutinin (WGA) staining procedures after western-blotting. Analysis of the optical density of the reactive bands allowed to estimate the relative concentration of the various glycopeptides in the eluted fractions. In the SCO-eluted fractions at least ten Con A-positive glycopeptides were identified, their apparent molecular weight ranging from 240 to 42 kD. Only three of these appeared to be WGA-positive (98, 88, and 52 kD). In the CH-eluted fractions only a 52 kD Con A- and WGA-positive glycopeptide was revealed, while in the MO-eluted fractions a 32 kD glycopeptide was also Con A- and WGA-positive. These results are discussed in regard to the known biosynthesis pathway of complex type glycoproteins.

Animals↗

PubMed1991

Patterns of cell division and interkinetic nuclear migration in the chick embryo hindbrain.

Early in its development, the chick embryo hindbrain manifests an axial series of bulges, termed rhombomeres. Rhombomeres are units of cell lineage restriction, and both they and their intervening boundaries form a series that reiterates various features of neuronal differentiation, cytoarchitecture, and molecular character. The segmented nature of hindbrain morphology and cellular development may be related to early patterns of cell division. These were explored by labeling with BrdU to reveal S-phase nuclei, and staining with basic fuchsin to visualise mitotic cells. Whereas within rhombomeres, S-phase nuclei were located predominantly toward the pial surface of the neuroepithelium, at rhombomere boundaries S-phase nuclei were significantly closer to the ventricular surface. The density of mitotic figures was greater toward the centres of rhombomeres than in boundary regions. Mitotic cells did not show any consistent bias in the orientation of division, either in the centres of rhombomeres, or near boundaries. Our results are consistent with the idea that rhombomeres are centres of cell proliferation, while boundaries contain populations of relatively static cells with reduced rates of cell division.

Actins↗

PubMed1987

In vitro methods for the analysis of motor function in the developing spinal cord of the chick embryo.

The isolated spinal cord of the chick embryo spontaneously generates episodes of motor activity in vitro that can be recorded from muscle nerves and ventral roots. In vitro systems provide stable conditions for intra- and extra-cellular recordings and enable pharmacological and ionic manipulations of the neuronal environment. Studies of motor activity generated by isolated spinal cord have revealed the existence of co-ordinated motor output from early in development, in which antagonist motoneurons alternate in their activity and synergists are co-active. Intra-cellular recordings from single neurons and electronic recordings from muscle nerves have provided insight into the mechanism of flexor and extensor alternation. These studies have revealed that flexor and extensor motoneurons receive a similar de-polarization during each cycle of motor activity, but that the two classes of motoneuron process the de-polarization differently. Flexors fire late in each cycle whereas extensors fire early, which leads to a pattern of alternation. The cellular mechanisms responsible for the differences in the firing behavior of flexor and extensor motoneurons are currently being investigated using techniques that are only possible using the in vitro preparation.

Animals↗

PubMed1994

Regulation of SC1/DM-GRASP during the migration of motor neurons in the chick embryo brain stem.

The hindbrain of the chick embryo contains three classes of motor neurons: somatic, visceral, and branchial motor. During development, somata of neurons in the last two classes undergo a laterally directed migration within the neuroepithelium; somata translocate towards the nerve exit points, through which motor axons are beginning to extend into the periphery. All classes of motor neuron are immunopositive for the SC1/DM-GRASP cell surface glycoprotein. We have examined the relationship between patterns of motor neuron migration, axon outgrowth, and expression of the SC1/DM-GRASP mRNA and protein, using anterograde or retrograde axonal tracing, immunohistochemistry, and in situ hybridization. We find that as motor neurons migrate laterally, SC1/DM-GRASP is down-regulated, both on neuronal somata and axonal surfaces. Within individual motor nuclei, these lateral, more mature neurons are found to possess longer axons than the young, medial cells of the population. Labelling of sensory or motor axons growing into the second branchial arch also shows that motor axons reach the muscle plate first, and that SC1/DM-GRASP is expressed on the muscle at the time growth cones arrive.

Activated-Leukocyte Cell Adhesion Molecule↗

PubMed1982

In vitro survival of chick embryo spinal neurons capable of forming synapses on skeletal muscle cells.

Survival of chick embryo spinal neurons capable of forming synapses on skeletal muscle cells (junction-forming neurons) was examined electrophysiologically in culture. Tissue extracts (skeletal muscle, heart, liver, brain), serum-free muscle-conditioned medium, horse serum (HS), chick embryo extract (CEE) showed survival-promoting activity. The highest activity was observed in a mixture of HS and CEE. This mixture did not promote the survival of all of the spinal cord cells, but selectively supported the survival of junction-forming neurons.

Animals↗

PubMed1979

Gel electrophoretic analysis of histones in lens epithelium, lens fiber, liver, brain, and erythrocytes of late chick embryos.

Histones from 19-day-old chick embryo lens epithelium, lens fibers, liver, brain, and erythrocytes were electrophoresed in polyacrylamide gels using buffers containing sodium dodecylsulfate, acetic acid urea, or mixtures of Triton X-100 acetic acid urea. In the last two buffer systems, histone bands were characterized by their apparent molecular weights determined by electrophoresis in the second dimension in sodium dodecylsulfate containing polyacrylamide gels. From the densitograms of the stained gels, the relative proportion of protein in different histone bands was estimated. With the exception of the erythrocyte-specific histone H5, all histones from different tissues examined at any of the gel systems migrated with the same mobilities. In lens epithelium and lens fibers, all histones were present in identical proportions. As compared to liver and brain, the total amount of histone Hl was significantly lower in lens cells and erythrocytes, possibly reflecting differences between the differentiated states. However, no tissue-specific differences were found in the relative distribution of histone Hl I and Hl II among lens epithelium, lens fiber, liver and, brain, but a threefold higher Hl I : Hl II ratio (0.5--0.7) was found in erythrocytes.

Animals↗

PubMed1985

[Effect of N-nitrosodiethylamine on the liver cells of chick embryos in primary culture].

The morphology and proliferative activity of liver cells from chick embryos were studied. In vitro, the cells were treated with N-nitrosodiethylamine (NDEA) in doses of 100 and 75 mg/ml of culture medium for a 10-day period. The primary culture from liver cells of 9-day chick embryos was prepared on the highly permeable celloidine membrane. In early periods of cultivation there were degenerative signs, mainly in the parenchymal cells, which shows a cytotoxic effect of NDEA. Proliferation of a small population of light epithelioid cells was observed towards the end of the experiment. There were also some atypical and pathological mitoses in the fibroblasts. The obtained results show that a neoplastic transformation of the chick embryonal liver cells occurs under in vitro conditions.

Animals↗

PubMedDate not supplied

Analysis of the cardiotoxic effect of isoproterenol in chick embryo.

Intraamnial administration of isoproterenol (IPRO) to chick embryo induces different types of pathological changes of the myocardial and liver tissue: their character depends upon the developmental period during which the drug was administered. Analysis of the embryotoxic effect of IPRO has revealed that the time-course of in vivo uptake of [3H]IPRO by the developing heart appears to present the same pattern as the embryonic liver tissue; both peak values (10 min and 6 h after injection) were, however, significantly higher in the liver as compared with the myocardium. IPRO is rapidly metabolized to 3-o-methyl-IPRO; the proportion of this fraction in both organs represents approximately 40% of total radioactivity as early as 5 min after administration. Intraamnial administration of IPRO significantly increases the cAMP content in the myocardium; dibutyryl cAMP-induced myocardial lesions occurred, however, only occasionally and in comparison with IPRO they were significantly less pronounced. The cardiotoxic effect of IPRO can be significantly reduced by beta-blockade; calcium antagonist, verapamil, was found to have no protective effect. Our results suggest that the following factors may participate in the IPRO-induced embryotoxicity: (1) IPRO, (2) its toxic metabolites, (3) cAMP.

Absorption↗

PubMed1981

Insulin and thyroid hormone stimulation of histotypic organogenesis and proteoglycan synthesis in seven-day chick embryo liver.

Livers from seven-day chick embryos were dissociated into single cells and the cells were allowed to aggregate in Dulbecco's Modified Eagle's Medium-12% foetal calf serum, 1-3H-glycosamine, 2-14C-glycine and hormones. Insulin, 10(-6)M, accelerated the rate of aggregation and resulted in larger aggregates than the controls, without added hormone . 3, 3', 5 Tri-iodo-thyronine, 10(-8)M, gave rise to slightly smaller aggregates than control aliquots. Incorporation of 2-14C-glycine and 1-3H-glucosamine was almost linear the first 36 hours whereupon incorporation reached a plateau at 48 hours. Insulin, 10(-6)M, enhanced 1-3H-glucosamine incorporation 54% and glycine incorporation to 42%. Tri-iodo-thyronine stimulated 1-3H-glucosamine 110% and glycine incorporation by 68%. When both insulin and tri-iodo-thyronine were added to the culture medium, glucosamine incorporation was augmented by 156% and glycine incorporation was stimulated by 77%. These findings are interpreted to indicate that synthesis of glycoproteins and proteo-glycosaminylglycans favours histogenesis or specific cell orientation and cell-cell interactions. Hormones had only a small effect on the incorporation of glycine and glycosamine into the soluble media components.

Animals↗

PubMed1988

Experimental induction of microphthalmia in the chick embryo with a single dose of cisplatin.

Chick embryos were injected on the fifth day of incubation with 75 ng cis-diamminedichloroplatinum II (cisplatin) and killed at daily intervals. Bilateral microphthalmia appeared in 88% of the surviving embryos; the decrease in eye size was noticeable 2 or 3 days after injection. Coinciding with this, macroscopic, histological, and ultrastructural changes started to appear in the ciliary body: ciliary processes failed to form and the cells in the inner layer of the ciliary epithelium underwent degenerative changes. Changes in the retina appeared somewhat later. Despite the decreased growth rate of the whole eye the neural layer of the retina continued to grow rapidly; as a result, it formed numerous folds and acquired a glandular appearance. In the most severe cases the rapidly growing retina would invade the ciliary region and replace completely the degenerated inner layer of the ciliary epithelium. It has been shown by previous authors that intraocular pressure is a determinant of eye expansion and also that the secretion of water and ions by the ciliary epithelium is important for the maintenance of that intraocular pressure. On this basis, our results are interpreted as indicating that the primary lesion induced by cisplatin was in the ciliary epithelium and that microphthalmia was the consequence of decreased pressure. It is also concluded that the retinal changes were due to the fact that the retina continued to grow despite the lack of expansion of the eye as a whole.

Age Factors↗

PubMed1998

Neural fold fusion in the cranial region of the chick embryo.

Cranial neural fold fusion in the chick embryo is known to commence in the midbrain region before progressing cranially and caudally to involve the fore- and hindbrain regions, respectively. The two epithelial layers at the tips of the neural folds that participate in fusion are the surface ectoderm and the neuroepithelium. We have examined and compared cranial neural fold fusion in both layers, and our results show that fusion of the neuroepithelial component of the neural folds, unlike that of the surface ectoderm, starts in the caudal portion of the forebrain. Second, contrary to the widely accepted opinion, we have demonstrated that in the hindbrain region, fusion of the neuroepithelial component of the neural folds does not occur. Soon after neural fold apposition, a neuroepithelial eminence appears in rhombomeres 1 and 2, and this, together with other neuroepithelial cells in the dorsal midline of the hindbrain, undergoes massive apoptosis. The absence of neuroepithelial fusion in the hindbrain may be due to the presence of massive apoptosis among neuroepithelial cells that should have participated in the fusion process. The events described above may predispose the hindbrain to the development of neural tube defects. The appearance of cranial neural crest cells in the midline during their migration may enhance the fusion of the surface ectodermal portion of the neural folds.

Animals↗

PubMedDate not supplied

[Immunohistochemical study of fibronectin localization during the formation of the vascular coat under the influence of the pigment epithelium in chick embryos].

The role of fibronectin (FN) in cell interactions of retinal pigment epithelium (RPE) and mesenchyme surrounding the optic cup during choroid formation in chick embryos was studied by indirect immunofluorescence using antibodies against FN. Experimental coloboma of retina and choroid was used as a model. During the initial stages of coloboma the regions structured like retina rudiment appear in the outer layer of the optic cup. Such regions were formed in microphthalmic eyes obtained by excision of lens from the eyes of 3.5 day old chick embryos (stage 21). At stage 21 bright FN-specific immunofluorescence was observed in basal membrane located along the external surface of the normally differentiated RPE. Later on, FN-specific immunofluorescence appeared in mesenchyme condensing along the RPE. The most intensive FN-specific immunofluorescence was observed in chorio-capillary layer of choroid after 5-7 days of incubation. In microphthalmic eyes retina-like regions of RPE and adjacent mesenchyme showed negative reaction, and the choroid was not formed from the adjacent mesenchyme in such zones. The data obtained suggest that the presence of normally differentiated RPE producing FN-containing basal membrane is necessary for the formation of chorio-capillary layer of the choroid in chick embryos.

Animals↗

PubMed2005

Human and rat glioma growth, invasion, and vascularization in a novel chick embryo brain tumor model.

The mechanisms that control the insidiously invasive nature of malignant gliomas are poorly understood, and their study would be facilitated by an in vivo model that is easy to manipulate and inexpensive. The developing chick embryo brain was assessed as a new xenograft model for the production, growth, and study of human and rat glioma cell lines. Three established glioma lines (U-87 MG, C6, and 9L) were injected into chick embryo brain ventricles on embryonic day (E) 5 and brains were examined after several days to two weeks after injection. All glioma lines survived, produced vascularized intraventricular tumors, and invaded the brain in a manner similar to that in rodents. Rat C6 glioma cells spread along vasculature and also invaded the neural tissue. Human U-87 glioma cells migrated along vasculature and exhibited slight invasion of neural tissue. Rat 9L gliosarcoma cells were highly motile, but migrated only along the vasculature. A derivative of 9L cells that stably expressed the cell surface adhesion molecule NgCAM/L1 was produced and also injected into chick embryo brain ventricles to see if this protein could facilitate tumor cell migration away from the vasculature into areas such as axonal tracts. 9L/NgCAM cells, however, did not migrate away from the vasculature and, thus, this protein alone cannot be responsible for diffuse invasiveness of some gliomas. 9L/NgCAM cell motility was assessed in vitro using sophisticated time-lapse microscopy and quantitative analysis, and was significantly altered compared to parental 9L cells. These studies demonstrate that the chick embryo brain is a successful and novel xenograft model for mammalian gliomas and demonstrate the potential usefulness of this new model for studying glioma tumor cell growth, vascularization, and invasiveness.

Animals↗