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Biomedical subjects

L Vakaet

Publications and source records attributed to L Vakaet.

At least 37 records · Page 2Linked to original sources
PubMed1991

Features of polyingression and primitive streak ingression through the basal lamina in the chicken blastoderm.

The de-epithelialization of cells of the upper layer during the phenomena of polyingression and primitive streak ingression was studied by analyzing, from the time of laying to the end of gastrulation, the ultrastructure of the basal lamina underlying the upper layer. The electron density of the basal lamina and associated extracellular materials was enhanced by addition of tannic acid to the fixative. Special attention was also paid to the spatial and temporal distribution of blebs at the basal surface of the upper layer, and to the contribution of the de-epithelialized cells to the formation of the deep layer. The results indicate that a nascent basal lamina is already present at the time of laying, especially beneath regions of the area pellucida where polyingression is not apparent. From the onset of incubation, the basal lamina rapidly develops, and it is interrupted by a large number of blebs. However, during the first 6-8 h of incubation, i.e., stages 1-2 of Vakaet (Arch. Biol. (Liège) 81:387-426, 1970), a downward movement of de-epithelialized cells that insert into the deep layer and form the endophyll persists cranially. This phenomenon of polyingression, which starts during the intrauterine period, probably extends from caudal to cranial and comes to an end by stage 3. During these first three stages, the number of blebs progressively decreases, especially in the cranial part of the area pellucida, and a thicker, continuous basal lamina associated with numerous interstitial bodies is laid down. The caudal part of the upper layer is still actively blebbing at that time. Due to the convergence of this area toward the axis of the blastoderm, which leads to ingression at and elongation of the primitive streak up to and including stage 6, the number of blebs at the basal surface of the upper layer progressively decreases. From stage 7 on, blebs are virtually absent; shortening of the primitive streak and formation of the head process begin. At the level of the head process, primitive streak ingression has ceased and a novel basal lamina is progressively deposited beneath the upper layer. By stage 9, a thick, smooth basal lamina physically separates the upper layer from the head mesenchyme. Summarizing, at the time of gastrulation, the presence of blebs that perforate the basal lamina is correlated with the de-epithelialization of cells. Before incubation, however, de-epithelialization of upper-layer cells occurs before the assembly of the basal lamina.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Esophageal cancer.

The authors invited eminent Belgian specialists to report on their experience in esophageal cancer. A summary is given of the most striking features of their reports.

Combined Modality Therapy↗
PubMed1991

Numerical evaluation of the invasion of closely related cell lines into collagen type I gels.

Cells were seeded on top of a reconstituted collagen gel layer, and their migration into the gel was evaluated as an assay for invasive behavior. The method was standardized by measuring the depth of migration of each cell in a defined volume of the gel. We developed a microscope stage, controlled by a computer program. This semiautomatic counting method allowed precise vertical localization of each cell in a collagen gel with an error of less than 0.1 micron. To test the discriminative power of the assay, we used cell lines which were known to be invasive or noninvasive in other assays. Closely related variants of 2 cell families were chosen: (1) one family derived from a mouse mammary gland (NMuMG), and (2) one derived from a mouse T cell lymphoma (BW5147). The assay could discriminate between invasive and noninvasive variants of related cell lines within the same family. The profile of the number of cells in each layer of the gel provided additional discrimination between the different cell lines. Furthermore, the assay allowed direct microscopic observation of cells migrating in the collagen gel. The present standardization makes the collagen assay suitable for semiautomatic testing of the invasive phenotypes in cell populations from the same as well as from different cell families.

Animals↗
PubMed1989

Fate mapping the avian neural plate with quail/chick chimeras: origin of prospective median wedge cells.

The origin of prospective M cells, which are median neuroepithelial cells that become wedge-shaped during bending of the neural plate and eventually form the midline floor of the neural tube, was determined by constructing quail/chick chimeras and using the quail nucleolar marker to identify quail donor cells in chick host blastoderms. Two possible sites of prospective M-cell origin in the epiblast were examined: a single, midline rudiment located just rostral to Hensen's node and paired rudiments flanking the cranial part of the primitive streak. Our results suggest that M cells arise exclusively from the midline, prenodal rudiment. From this rudiment, M cells extend caudally throughout the entire length of the neuroepithelium. This new information on the origin of prospective M cells will aid in the analysis of their role in neurulation.

Animals↗
PubMed1989

Neural plate- and neural tube-forming potential of isolated epiblast areas in avian embryos.

Formation, shaping, and bending of the neural plate and closure of the neural groove are complex processes resulting in formation of the neural tube. Two experiments were performed using avian embryos as model systems to examine these events. First, we transected blastoderms near the level of Hensen's node to determine the potential of prenodal neural plate to form neural tube in isolation from primitive streak regression. Our results demonstrate that shaping and bending of the prenodal neural plate occur under these conditions, but neural groove closure is inhibited. Second, we isolated various areas of postnodal epiblasts to determine their potential to form neural plate. Our results suggest that the area of the postnodal epiblast that can form neural plate consists of paired tracts lying adjacent to the definitive primitive streak and extending caudally at least 1 mm from its cranial end.

Animals↗
PubMed1989

Immunohistochemistry of laminin in early chicken and quail blastoderms.

We have used immunohistochemical techniques to study laminin in quail blastoderms milked from the oviduct and the distribution of laminin in laid chicken and quail blastoderms. Laminin is a constituent of the basement membrane in both chicken and quail blastoderms. It is found at the ventral side of the upper layer cells. Laminin is first observed under individual upper layer cells in prelaid quail blastoderms 15 h post-ovulation, but is absent at the ingression site of endophyll cells. The presence of a continuous laminin layer coincides with the epithelialization of the epiblast after 5-10 h incubation. The laminin layer is discontinuous at the primitive streak and at Hensen's node. It is thinner and partly discontinuous at the median part of the neural plate. By induction, either of an ectopic primitive streak or a neural plate, we have demonstrated, using the chicken-quail nucleolar marker technique, that at these sites the laminin layer is interrupted. A laminin layer might confer rigidity onto the epiblast, whereas disruption of a laminin layer seems to be correlated with ingression of cells or bending of the neural plate.

Animals↗
PubMed1989

Morphology of a columellaform stapes.

A columellaform stapes found in male patient 18 years old with a conductive hearing loss, has been examined by SEM and light microscopy. The ossicle was smaller than a normal stapes. The head was linked to the base by a crural plate, probably formed by the fusion of the material of the two crura. Microscopy showed a slightly less interwoven structure of the fibrillar bone present mainly at the cranial aspect of the crural plate. Honeycomb-like bone was confined to the caudal part of the crural plate. Microscopy revealed a cavity in the centre of the stapes at the transition between the base and the plate. During surgery, a bony structure was found linking the posterior side of the head of the stapes to the pyramidal eminence. Post-operative audiograms revealed improved hearing with some residual conductive loss of about 10 dB. This case illustrates that a columellaform stapes may occur without forming part of a syndrome.

Adolescent↗
PubMed1989

The chicken blastoderm: in vitro methods of investigation of developmental processes.

A brief description of the structure and a critical survey of the methods of explantation, cultivation, and experimental manipulation of chicken blastoderms are presented. Special attention has been paid to those methods that have enabled major developments in cell and developmental biology of gastrulation. The data suggest that the chicken blastoderm is an appropriate system for the study of developmental processes that are often analogous to medical problems.

Animals↗
PubMed1988

Morphological and immunocytochemical studies of fibronectin-coated, plasma membrane-limited vesicles in the early chicken embryo.

Vesticles with a mean outside diameter of 32.8 nm have been observed in the early chicken embryo after fixation with a mixture of glutaraldehyde and tannic acid. Densitometric tracing has revealed that the vesicles are limited by a unit membrane. The presence of complex carbohydrates is suggested by the increased electron density of the vesticles after addition of tannic acid to the fixative. Immunocytochemical staining with a monoclonal antibody directed against chicken cellular fibronectin demonstrated the presence of this glycoprotein along the surface of the vesicles. These results suggest a cellular origin of the vesicles, since their surface shares morphological and biochemical similarities with the cell surfaces of the embryonic tissue layers. Recycling of plasma-membrane vesicles may occur, as vesicles were found in the vicinity of coated vesicles. We postulate that extracellular materials of the cell surface, which may affect cell and tissue interactions, are shed in the environment together with plasma-membrane vesicles. The difficulties encountered in observing the vesicles stems from the facts that an adequate visualization method is necessary and that they are few in number. The latter reason suggests their transient nature. The vesicles probably rapidly disintegrate in the extracellular milieu or are recycled by the cell surface.

Animals↗
PubMed1988

Connecting cords and morphogenetic movements in the quail blastoderm.

Connecting cords are elongated telophase bridges persisting between separating daughter cells. We have studied them with Scanning Electron Microscopy in the upper cell layer of the quail blastoderm where a high mitotic activity accompanied by interkinetic nuclear migration coincides with morphogenetic movements. The predominant orientation of the connecting cords is parallel to the direction of the morphogenetic movements.

Animals↗
PubMed1987

The anti-invasive flavonoid (+)-catechin binds to laminin and abrogates the effect of laminin on cell morphology and adhesion.

To study the effect of the flavonoid (+)-catechin on cell-matrix interactions two cell types with a different morphology on and adhesion to laminin were used. MO4 virally transformed fetal mouse cells adhere and spread when cultured on top of laminin-coated coverslips or on human amnion basement membrane. M5076 mouse reticulum cell sarcoma cells poorly adhere to these substrates and remain round. Both cell types are invasive in confronting cultures with embryonic chick heart fragments. (+)-Catechin binds to laminin in a pH-dependent way. Pretreatment of laminin-coated coverslips or amnion basement membrane with 0.5 mM (+)-catechin abrogates the effect of laminin on cell morphology and adhesion. MO4 cells do not adhere to the pretreated substrates and remain round, while M5076 cells now adhere and spread. (+)-Catechin inhibits the invasion of MO4 cells but not of M5076 cells into embryonic chick heart in vitro. We speculate that the anti-invasive activity of the flavonoid to MO4 cells is the result of its interference with MO4 cell adhesion to laminin. Invasion of M5076 cells does not imply adhesion to and spreading on laminin.

Animals↗
PubMed1986

Confrontation of an invasive (MO4) and a noninvasive (MDCK) cell line with embryonic chick heart fragments in serum-free culture media.

Confronting cultures of precultured embryonic chick heart fragments (PHF) with aggregates of malignant cells in vitro have been shown to be relevant for a number of aspects of tumor invasion in vivo. Preculture of the heart fragments, formation of cell aggregates, and subsequent culture of confronting pairs have so far been done only in serum-containing culture media. We describe here confronting cultures of PHF with invasive MO4 mouse cell aggregates or noninvasive MDCK dog kidney cell aggregates in serum-free media. Heart fragments precultured in the absence of serum seemed to be necrotic after confronting culture in serum-free media. However, preculturing in media supplemented with 10% fetal bovine serum allowed us to do subsequent confronting cultures in absence of serum. Cell aggregates were also prepared in serum-containing medium. MO4 cells occupied and replaced the heart tissue within 4 d, whereas MDCK cells remained at the periphery of the PHF. This indicates that serum-free confronting cultures can discriminate between invasive and noninvasive cells. The viability of individual PHF and cell aggregates cultured in the same way as in confrontations was ascertained by histology and by explantation and postculturing on a solid tissue culture substrate. Growth of the cultures was smaller in serum-free media than in media supplemented with 10% fetal bovine serum. The main advantage of serum-free culture conditions in vitro is the elimination of the influence of serum components on invasion, and the ability to examine the effect on invasion of drugs that are susceptible to inactivation by serum.

Animals↗
PubMed1986

Microinjection of glycosaminoglycan-degrading enzymes in the chicken blastoderm. An ultrastructural study.

The relationship between the presence of glycosaminoglycans (GAGs) and the morphology of the middle layer or mesoblast was examined by performing transmission electron microscopy of chicken blastoderms microinjected with GAG-degrading enzymes. The controls included microinjections with saline or trypsin, as well as solid-phase assays for proteolytic activity in commercially available GAG-degrading preparations. The results indicate that, in normal as well as in saline-injected blastoderms, middle-layer cells are rounded or cuboidal in shape, and are linked to each other by small intercellular junctions in the primitive-streak region. As they migrate laterally along the basal lamina, they appear as typical mesenchymal cells, being separated by large intercellular spaces and covered by cell processes. The removal of hyaluronate (by the microinjection of hyaluronidases) led to compaction of the middle-layer cells in the area lateral to the primitive streak. These cells lost their mesenchymal aspect and retracted their processes, and intercellular junctions were observed. The presence of proteolytic activity in the enzyme preparations did not interfere with the results. On the basis of the results obtained using this microinjection technique, we were able to confirm at the ultrastructural level that hyaluronate, due to its space-creating properties, promotes the detachment of ingressed primitive-streak cells and preserves the mesenchymal aspect of the middle layer during the lateral migration of single cells along the basal lamina. Whether the presence of hyaluronate is necessary to allow positioning of the mesoblast could not be inferred using our experimental procedure. We present evidence that this molecule, as well as having physicochemical properties, is also involved in the modulation of tissue interactions during gastrulation.

Animals↗
PubMed1985

Fibronectin and its relation to the basal lamina and to the cell surface in the chicken blastoderm.

The ultrastructural distribution of fibronectin immunoreactivity was investigated in the chicken embryo during late gastrulation. Sites of binding of anti-fibronectin antibodies were ascribed to the basal lamina and associated structures, and to the cell surface. The fibronectin-rich basal lamina was resolved into a lamina densa, which appears as a continuous, dense sheet, a lamina lucida, consisting of anchoring cords between lamina densa and epithelial cells, and a lamina intima, closely juxtaposed to the cell surface. Cell-surface labelling was also observed in mesoblast cells, and along the dorsal side of the deep-layer cells. The ventral side of the latter cells was poorly stained in the endophyllic crescent, except in coated pits, and more regularly stained at the level of definitive endoblast. Some structures associated with the basal lamina reacted intensely with anti-fibronectin antibodies. These are the interstitial bodies, which are aggregates of extracellular material, and a kind of fibril or tubule, embedded in a fibronectin matrix and mainly found in the endophyllic crescent. Some intracellular labelling was found in most deep-layer cells, in few epiblast cells, never in mesoblast cells. These results extend previous studies on the localization of fibronectin, and correlate its presence and surface topology with its postulated role in migration of mesoblast cells on the basal lamina which, chemically, constitutes an appropriate substrate.

Animals↗
PubMed1985

Transfer of extracellular matrix components between germ layers in chimaeric chicken-quail blastoderms.

A chemical basis for the transmission of signals during gastrulation has been investigated by using chimaeric embryos resulting from the combination of 3H-glucosamine-labelled and unlabelled hypoblast with epiblast taken from chicken and quail embryos at stage 3 of Vakaet (1970). The ability to distinguish chicken from quail cells on the basis of their different nuclear distribution of heterochromatin after Feulgen staining made it possible to determine the origin of the cells in the chimaerae. Tritiated quail hypoblast (after incubation of the embryo in the presence of 3H-glucosamine) was transplanted onto unlabelled chicken blastoderm deprived of its hypoblast. After culture of the chimaera for 5 h, the autoradiographic pattern shows silver grains not only over the graft, but also at the ventral surface of the epiblast of the host. Transfer of label may occur to mesoblast cells, but not between chicken and quail hypoblast cells. Chase experiments exclude the possibility that unprocessed, tritiated glucosamine is transferred. Chemical fixation of the host before transplantation of a labelled quail hypoblast also allows visualization of a transfer of macromolecules from hypoblast to the basement membrane of the epiblast, suggesting that an intervention of the epiblast cells in this process is not necessary. The morphology of the chimaeric embryos, as studied by scanning electron microscopy, suggests a direct deposition of these macromolecules by filopodia of the dorsal surface of the hypoblast. The possibility of diffusion of free macromolecules has been considered and can reasonably be discarded on the basis of several observations.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Inhibition of cell spreading on the band of extracellular fibres in early chick and quail embryos.

The ventral surface of the upper layer shows a band of extracellular fibrils around the anterior and lateral border of the area pellucida during gastrulation of the chick embryo. Using scanning electron microscopy, we found that this disposition is correlated with the motility of the middle-layer cells of gastrulating chick and quail embryos. Outside the fibrous band, single middle-layer cells and a sheet of mesoblast cells were spread out and possessed lamellae. Single cells on the fibrous band did not form lamellae. The same cell behaviour was obtained with the explants of deep layer on the fibrous band. The fibrous band is assumed to operate as a barrier that inhibits cell motility during gastrulation.

Animals↗
PubMed1985

Cell junctions and locomotion of the blastoderm edge in gastrulating chick and quail embryos.

The blastoderm edge migrates by the active locomotion of a multilayer of epithelial cells, the so-called margin of overgrowth (MO), that uses the vitelline membrane as its substratum. The structural unity formed by the margin of overgrowth cells and their rapid migration suggest coordination of locomotion between individual cells. Using transmission electron microscopy of thin sections and freeze-fracture, we attempted to determine if the pattern of junctions of the migrating margin of overgrowth is related to the suggested cell--cell cooperation between individual cells in this region. In the leading edge there are large areas of closely apposed cell membranes. Incipient desmosomes and small gap junctions were observed. Tight junctions consisted of isolated strands or isolated networks of tight-junctional strands. In the proximal part of the margin of overgrowth the size of the gap junctions increased and the desmosomes were fully developed. Tight-junctional strands were either isolated or arranged into an isolated network. A broad belt of tight junctions was observed at the transition between margin of overgrowth and non-marginal cells. The distribution of the junctional elements in the MO suggests that junctions contribute to the maintenance of the structural and functional organization of the margin of overgrowth. Furthermore, the spatial distribution of the junctions might give information about the mechanism of locomotion of the margin of overgrowth.

Animals↗