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V Nurcombe

Publications and source records attributed to V Nurcombe.

51 records · Page 3Linked to original sources
PubMed1991

Basic fibroblast growth factor upregulates steady-state levels of laminin B1 and B2 chain mRNA in cultured neuroepithelial cells.

The growth of purified populations of murine neuroepithelial cells isolated from 10 day embryonic (E10) telencephalon and mesencephalon can be specifically enhanced by supplementing growth culture media with basic fibroblast growth factor (bFGF). One effect of bFGF on cultured neuroepithelial cells was to enhance the amount of laminin expressed at the protein level as detected by immunofluorescence. This was correlated with significant upregulation of steady-state levels of laminin B1 and B2 chain expression as analyzed at the mRNA level. When E12 neuroepithelial cells were split into precursor neuronal or glial subpopulations on the basis of differential expression of major histocompatibility class-1 antigens, only the glial progenitor fraction was found to be capable of detectable laminin synthesis. It is thus possible that a primary action of FGF is to increase the synthesis and release of extracellular matrix molecules from neural cells which act back in a paracrine manner to stimulate differentiation.

Animals↗
PubMed1989

The high-affinity binding of laminin to cells. Assignation of a major cell-binding site to the long arm of laminin and of a latent cell-binding site to its short arms.

The laminin proteolytic fragments 1 (derived from the intersection of the short arms of the cruciform laminin molecule) and 8 (derived from the laminin long arm) bind to distinct receptors on HT-1080 human fibrosarcoma cells; both fragments are shown here to inhibit the high-affinity binding of laminin to these cells. Inhibition of binding between fragment 8 and laminin was competitive, whereas that between fragment 1 and laminin was noncompetitive. This indicates that laminin and fragment 8 most probably share the same cellular receptors, whereas laminin and fragment 1 bind to distinct receptors, inhibition being due to steric hindrance. Surprisingly, fragment 1-4 (corresponding to the complete short arms of laminin) neither bound to HT-1080 cells nor inhibited the binding of laminin or fragment 1. After treatment of fragment 1-4 with pepsin, however, the smaller subfragment 1 was liberated, which could then bind to the cells, and so was shown to block the binding of laminin and fragment 1. We conclude that native laminin bound to HT-1080 cells via the fragment-8-binding site near the end of its long arm. Although these cells also have distinct receptors for the short arm fragment 1, this receptor-binding site was not used as it appeared to be latent within the native laminin molecule.

Binding Sites↗
PubMed1989

Locomotory competence and laminin-specific cell surface binding sites are lost during myoblast differentiation.

The specific interaction of embryonal cells with the extracellular matrix (ECM) is one of the principal forces influencing embryonal development (Hay, 1984; Trinkaus, 1984). We used a muscle satellite cell line (MM14dy) to determine the relationship between locomotory response to laminin and the expression of specific cell surface binding sites for it. Time lapse videomicroscopic analysis was used to study the locomotory response and radioligand binding assays and cell attachment assays were used to follow the expression levels of binding sites for laminin and its subfragments E8 and E1-4. We report here the novel finding that the ability of MM14dy to locomote over laminin diminishes and finally vanishes as the cells differentiate. The simultaneous drop in expression of binding sites for laminin is interpreted as being of potential significance during development and repair.

Animals↗
PubMed1989

Developmental loss of functional laminin receptors on retinal ganglion cells is regulated by their target tissue, the optic tectum.

The ability of chick retinal ganglion cells (RGCs) to extend neurites on tissue culture substrata of the extra-cellular matrix protein laminin is lost during embryonic development. In order to establish the mechanism responsible for the loss of response, the number of high affinity (KD 10(-9) M) laminin receptors on both the cell bodies and neurites of RGCs were determined throughout this period by a ligand binding assay using radio-labelled laminin. It was found that the loss of response paralleled a decrease in receptor numbers on both the cell bodies and the neurites of the RGCs. Bilateral tectal ablation at embryonic day 6 resulted in the subsequent maintenance of laminin-stimulated neurite outgrowth, together with a partial inhibition of the loss of laminin receptors. Thus, the loss of response of the RGCs to laminin reflects a decrease in the numbers of laminin receptors on these neurons, and furthermore, this down-regulation is in turn dependent on innervation of the target tissue.

Animals↗
PubMed1987

The cellular interactions of laminin fragments. Cell adhesion correlates with two fragment-specific high affinity binding sites.

The molecular interactions of laminin with several tumor cell lines and skin fibroblasts were investigated by radioligand binding studies and cell attachment assays using laminin, the laminin-nidogen complex, and laminin fragments as substrates and also domain-specific antibodies as inhibitors of cell attachment. The majority of cells showed a dual binding pattern for fragments 1 and 8 which originate from short-arm or long-arm structures of laminin, respectively. Both of these fragments in solution bind to suspended cells with high affinity (KD = 1-10 nM), with the receptor numbers for each fragment depending on the cell type. Competition studies and independent variation of receptor numbers demonstrated that the cell-binding structures on each fragment are different, implicating the existence of two distinct cellular receptors for laminin. The ability of these fragments to act as substrates for cell adhesion correlated with the presence of high affinity binding sites on the cells. However, only antibodies to fragment 8 were able to block cell adhesion to laminin, despite the presence of binding sites for fragment 1. A few cells had very low numbers of high affinity receptors for either fragment 1 or 8. The latter cell type was used to demonstrate that complex formation between laminin and nidogen, which binds to fragment 1 structures, reduces the potential of laminin for cell binding.

Animals↗
PubMed1985

Motoneurone survival and neuritic outgrowth promoted by different cell types in embryonic muscle.

Different cell types within developing chick skeletal muscle were assayed for their ability to release factors into culture media which could affect the survival and neuritic development of labelled motoneurones and lateral motor column explants. Enriched cultures of myotubes, myoblasts, fibroblasts and mesenchyme were prepared by selective preplating and trypsinisation techniques. Degrees of enrichment were assessed immunofluorescently and morphologically; fibroblasts were the main contaminating cell type. Medium conditioned over each cell type was then tested in dose-response assay against both explants and dissociated motoneurones. In both cases the myotube conditioned medium (MCM) promoted the greatest levels of both survival and neuritic outgrowth, and had the greatest relative potency of all of the cell types. When MCM was preincubated over polycationic substrata, it lost the ability to promote neuritic growth; this could be restored if fresh conditioned medium (CM) was added to the cultures. Thus it was demonstrated that within the MCM there are physically separable agents responsible for neurone survival and neurite expression. The neurite-promoting factor (NPF) within the MCM was stable to collagenase, deoxyribonuclease, neuraminidase and chondroitinase ABC, but was destroyed by trypsin and heparinase. These results imply that a heparan sulfate proteoglycan is essential for the activity of the factor.

Animals↗
PubMed1984

Motor neuron survival and neuritic extension from spinal cord explants induced by factors released from denervated muscle.

Extracts prepared from denervated adult skeletal muscle contain increased amounts of neurotrophic activity which promotes both survival of dissociated motor neurons and the outgrowth of neurites from explants of spinal cord maintained in serum-free defined media. The trophic activity is specific for motor neurons and reaches a peak within the first week post-denervation. In these most potent extracts the neurite outgrowth enhancement is a linearly increasing function of protein concentration at low concentrations; at higher concentrations the neurite activity-concentration relationship saturates and in the milligram range the relationship becomes inhibitory. When media containing active denervated muscle extract was preincubated over polycationic substrata, it lost the ability to promote neuritic growth; this could be restored if fresh extract was added to the cultures. Thus it was demonstrated that within the denervated muscle extract there are physically separable agents responsible for neuron survival and neurite expression. It is possible that the release of neurotrophic factors may be in part responsible for the in vivo phenomenon of nerve sprouting.

Animals↗
PubMed1983

The growth of neurites from explants of brachial spinal cord exposed to different components of wing bud mesenchyme.

The extension of peripheral axons from the brachial spinal cord into the embryonic chick wing bud suggests that the target premuscle cell masses may act as the source of an adhesion gradient which establishes selective nerve pathways. Wing premuscle cell masses were explanted from different stage embryos and tested against age-matched brachial spinal cord for their ability to promote directional neurite outgrowth. It was found that target premuscle which does not contain myotubes could still elicit directional outgrowth in vitro. In contrast, skin and precartilage were unable to promote neuritic outgrowth significantly. Serum-free conditioned media were prepared from stages 20-38 premuscle and tested against age-matched spinal cord explants. There was an increase in the effects of conditioned media on neuritic outgrowth up to stage 35; conditioned media from older-stage premuscles had less effect than that of stage 35 premuscles. These results were shown to be dependent on the maturation of the premuscles and not on that of the spinal cord. When the premuscle conditioned media were preincubated over polylysine substrata, the ability to induce neuritic outgrowth was abolished from media derived from premuscle at stage 27 and older. Conditioned media derived from premuscle at stage 27 or older contain a polylysine-binding neurite-promoting factor which is present in greater amounts in more differentiated muscle. The time of first detection of neurite-promoting factors in stage 27 premuscle conditioned media correlates with the in vivo stages at which muscle-specific nerves branch from the main nerve trunks.

Animals↗
PubMed1982

Evidence for neuron-survival and neurite-promoting factors from skeletal muscle: their effects on embryonic spinal cord.

Explants of 8- and 9-day embryonic chick spinal cord were cultured for 4 days in medium which had been conditioned by skeletal muscle cells. When the conditioned medium was preincubated over polylysine substrata, it lost the ability to induce extensive neuritic outgrowth from the spinal cord. This ability was restored when the spinal cord was exposed to preincubated conditioned medium on preincubated polylysine substrata. Skeletal muscle thus appears to contain two separable components; one is a soluble factor which supports neuronal survival, and the other is an adsorbable factor which binds to appropriate substrata and promotes neuritic outgrowth.

Animals↗
PubMed1981

Embryonic chick retinal ganglion cells identified "in vitro". Their survival is dependent on a factor from the optic tectum.

When HRP is injected into the optic tecta of embryonic or newly hatched chicks, the ganglion cells in the contralateral retina can be successfully dissociated into culture and identified at any time by appropriate histochemical staining. Histological examination of whole mounts of retinae both ipsilateral and contralateral to an injection site indicated that no HRP diffused out of an injected tectum, and that the only reaction product that could be visualized was restricted to the ganglion cell layer of the contralateral eye. Because retinal ganglion cells are the only retinal neurons to project to the optic tectum, the intraxonal retrograde transport of HRP to these cells allows their unequivocal identification from amongst the heterogeneous population of retinal neurons present after dispersal into single cells in monolayer culture. The presence of HRP in the cell bodies did not appear to impair their ability to survive, grow or express neurites. Counts of labeled cells from progressively aged birds confirmed that the peak number of generated ganglion cells occurs on embryonic day 10,and that is a 40% decline in the number these neurons over the following 3 days. However, when labelled ganglion cells from 10 day embryos were grown in culture with optic tectum, all the ganglion cells survived over the following 4 days, including those destined to die in vivo. This trophic effect cannot be induced by cerebellum, but is partly induced by media first conditioned over tectal cells. The trophic effect exerted by optic tectum appears therefore to be specific and chemically mediated. We suggest that the death of retinal ganglion cells in vivo may be a consequence of the inability of some cells to establish adequate supplies of a growth factor from the optic tectum.

Animals↗