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E J Sanders

Publications and source records attributed to E J Sanders.

At least 19 recordsLinked to original sources
PubMed1997

Patterns of cell death during gastrulation in chick and mouse embryos.

We have examined the distribution of cells at an early stage of the cell death process in gastrulating chick and mouse embryos, using a DNA nick end-labelling technique to label nuclei that are undergoing DNA fragmentation in situ. In the chick embryo, the incidence of nuclei showing DNA fragmentation was mapped by digitizing the occurrence of these nuclei from sections, and reconstructing the three separate layers of the entire embryo at several stages of gastrulation. In the chick, DNA fragmentation was found in nuclei throughout the embryo, in cells of all three germ layers, but most especially in the epiblast in the rostral germinal crescent and in the lateral marginal zones. This region of greatest cell death formed an arc rostrally and laterally in the epiblast, and was consistent through gastrulation and into the early neurulation stage. While the extensive cell death in the chick embryo may be due to cell redundancy, it is also possible that the pattern of death observed could be related to the compression of the embryo against the barrier of yolk at the periphery of the area pellucida during expansion. In a number of cases in the chick, local regions of elevated cell death were also observed in the primitive streak. This may be associated with the changing cell-cell and cell-matrix interactions experienced by cells traversing the primitive streak. In the gastrulating mouse embryo, by contrast, nuclei undergoing DNA fragmentation showed no consistent regions of elevated incidence, in any of the embryonic layers. DNA fragmentation in these embryos was, however, observed in nuclei of cells in the visceral endoderm and in the epiblast. The lack of any clear pattern of DNA fragmentation in the mouse embryo at this stage of development leaves the roles of the dying cells enigmatic. The death may, however, be lineage-related or be a reflection of a cellular redundancy necessary in a developing system that is undergoing extensive cell rearrangement and cellular adhesive change.

Animals↗
PubMed1997

Roles for growth and differentiation factors in avian embryonic development.

We review the evidence for a role for transforming growth factor-beta (TGF-beta) and for tumor necrosis factor-alpha (TNF-alpha) in the development of the avian embryo. Transforming growth factor-beta is expressed in a number of locations in the early embryo with a distribution consistent with a function in epithelial-mesenchymal transformation and modulation of the composition of the extracellular matrix. During gastrulation, this factor is found in the mesoderm cell layer as well as in the endoderm underlying the primitive streak. In vivo and in vitro investigations suggest that TGF-beta may be involved in the regulation of phenotypic transformation, matrix deposition, and cell proliferation. Tumor necrosis factor-alpha and its two receptors are also located with distributions that suggest important involvement for this pleiotropic factor in early morphogenetic processes. Tumor necrosis factor-alpha is found in several cell populations from the time of gastrulation onwards, including the lens. In vitro investigations, using tissue from the gastrulating embryo as well as from the lens, suggest that this factor may be associated with the extensive cell death that occurs throughout the first 6 d of development, and with nuclear degeneration in the lens. We hypothesize that TNF-alpha, acting in a paracrine or autocrine fashion, may be involved in the signalling pathways that effect the regulation of cell death in development.

Animals↗
PubMed1996

Ultrastructural identification of apoptotic nuclei using the TUNEL technique.

We describe an ultrastructural adaptation of the method of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL) for the identification of DNA fragmentation. Thin sections of tissue embedded in hydrophilic resin were nick end labelled with biotinylated dUTP which was subsequently labelled with avidin conjugated to gold particles. The technique was validated by labelling the nuclei of L929-8 cells treated with tumour necrosis factor alpha. These cells are known to respond to treatment with the factor by undergoing apoptosis. The method was then used on tissue from the chick embryo which is known to be undergoing programmed cell death. This tissue was from the neural tube and the posterior necrotic zone of the limb bud, where cells can be identified as undergoing apoptosis based on the morphology of their nuclei. The method specifically labelled heterochromatin adjacent to the nuclear envelope as well as that associated with the nucleolus of cells from regions of the embryo where programmed cell death was expected. In addition to labelling the nuclei of cells that were clearly undergoing apoptosis, the method also identified nuclei of apparently normal cells. This method, used in conjunction with corroborating techniques, provides a means for the early detection of cells undergoing DNA fragmentation, before the onset of gross apoptotic morphology, and in cells that do not show classical apoptotic characteristics.

Animals↗
PubMed1996

Yellow fever: an emerging threat for Kenya and other east African countries.

Yellow fever (YF) is a well known disease that had plagued the tropics relentlessly until an effective vaccine was developed. Although the yellow fever vaccine is relatively affordable and one dose protects for over ten years, its use has predominantly been for known endemic areas of the world and international travellers. Eastern and southern African states, have hitherto been free of epidemic yellow fever, hence routine YF vaccination is not a policy in these countries. The sudden emergence of YF in the Rift Valley in Kenya in 1992-1993, introduces new dimensions into the challenges of YF to eastern and southern African states. Isolation of a virus deemed to be native of the area is discussed in this article in the context of YF policy issues confronting the region. A case has been argued for the establishment of a network of active surveillance systems in the region backed by adequate laboratory YF expertise locally, regionally, and internationally.

Africa, Eastern↗
PubMed1995

Sodium channel distribution in a spider mechanosensory organ.

A site-directed antibody was used immunocytochemically to measure the distribution of sodium channels in the tissues of a spider mechanoreceptor organ. The VS-3 slit sense organ contains 7-8 pairs of bipolar sensory neurons; these neurons are representative of a wide range of arthropod mechanoreceptors. Sensory transduction is thought to occur at the tips of the dendrites and to cause action potentials that are regeneratively conducted to the cell bodies, although it has not been possible to confirm this by direct intracellular recordings from the dendrites. Wholemount preparations were labelled by immunofluorescence and thin sections were immunogold labelled, using an antibody to the highly conserved SP19 sequence of the voltage-activated sodium channel. Labelling for sodium channels was found in the neurons and in their surrounding glial cells. Both cytoplasm and membranes were labelled, but immunogold particles were clearly aligned along cell membranes, indicating that the majority of labelling represented membrane-bound sodium channels. Channel density in the dendrites was similar to the axons and higher than in the cell bodies, supporting the idea of active conduction in the sensory dendrites. Labelling in glial cell membranes was indistinguishable from the neighboring neurons, suggesting a significant role for sodium channels in the functions of these supporting cells.

Action Potentials↗
PubMed1995

Transport of serum transthyretin into chicken oocytes. A receptor-mediated mechanism.

Transthyretin (TTR) is involved in the transport of thyroid hormones and, due to its interaction with serum retinol-binding protein, also of vitamin A. The importance of both ligands in vertebrate embryonic development has prompted us to investigate the molecular details of TTR transport function in a powerful germ cell system, the rapidly growing chicken oocytes. Yolk TTR is derived from the circulatory system, since biotinylated TTR was recovered by immunoaffinity chromatography of yolk obtained from a hen previously infused with in vitro biotinylated chicken serum proteins. In concordance with the intraoocytic localization in an endosomal compartment, ligand blotting and chemical cross-linking experiments revealed the presence of a approximately 115-kDa TTR-binding oocyte membrane protein. This putative TTR receptor was not detected in chicken ovarian granulosa cells or embryonic fibroblasts and was different from the previously described oocyte-specific receptor for two estrogen-induced chicken serum lipoproteins, vitellogenin and very low density lipoprotein (Barber, D. L., Sanders, E. J., Aebersold, R., and Schneider, W. J. (1991) J. Biol. Chem. 266, 18761-18770). Furthermore, in contrast to the serum levels of the yolk precursor lipoproteins, those of TTR were not significantly changed by estrogen; thus, TTR represents a newly defined, estrogen-independent class of yolk precursor proteins. These data strongly suggest that oocytic TTR is derived from the circulation, where it is a constitutive component, and deposited into yolk as a result of endocytosis mediated by a specific receptor.

Animals↗
PubMed1995

Glycoconjugates in normal and abnormal secondary neurulation.

In chick embryos, the anterior greater portion of the neural tube develops by the folding, apposition, and fusion of the neuroectoderm. The smaller caudal portion that forms the secondary neural tube (lumbosacral and coccygeal regions) is derived from the tail bud, an aggregate of mesenchymal cells located at the caudal limit of the body. Tail bud mesenchyme, arranged in a solid cord, undergoes mesenchymal-epithelial transformation to form the secondary neural tube. Previous evidence suggests that this transformation is accompanied by modulation of cell surface glycoconjugates in the differentiating tissues. In this study, we show by lectin histochemistry and lectin blotting of proteins isolated by SDS-PAGE, that Datura stramonium agglutinin (DSA) binds preferentially to differentiating tail bud cells. This lectin is specific for beta 1-4-linked N-acetylglucosamine oligomers, such as the oligosaccharides of the poly-N-acetyllactosamine series that have been previously implicated in cell differentiation. Ultrastructural lectin cytochemistry indicates that at least some of the proteins binding DSA are localized extracellularly. The use of DSA as a teratogen resulted in embryos showing a variety of neural tube and notochord defects. We have also examined the binding of DSA to embryos that were treated with teratogenic doses of retinoic acid by sub-blastodermal injection, and find that the DSA-binding patterns are perturbed. Analysis of DSA-treated embryos using the TUNEL technique indicated that cell death was not a factor in DSA teratogenesis. This strongly suggests that the glycoconjugates of the cell surface have a role in the normal differentiation of tail bud mesenchyme into the neuroepithelium of the secondary neural tube. Perturbations of glycoconjugate activity results in defects of the secondary neural tube and associated tail bud derivatives.

Abnormalities, Drug-Induced↗
PubMed1995

Potential roles for tumour necrosis factor alpha during embryonic development.

This paper reviews the evidence indicating possible roles for tumour necrosis factor-alpha (TNF alpha) in development. It is proposed that TNF alpha may have essentially three major roles during embryonic development, which may be analogous to its roles in the immune system and during inflammation: a role in programmed cell death; a role as a cellular growth and differentiation factor; and also a role in the remodelling of extracellular matrix, and the regulation of cell adhesion molecules and integrins. The concept of the existence of a cytokine array during embryogenesis, analogous to that occurring in inflammation, is discussed, as well as potential roles for TNF alpha in the induction of ubiquitin; protective mechanisms embryonic cells may employ against TNF alpha-mediated cytotoxicity; and a consideration of the role TNF alpha may play in a "free radical theory of development".

Animals↗
PubMed1995

Programmed cell death in development.

Although cell death has long been recognized to be a significant element in the process of embryonic morphogenesis, its relationships to differentiation and its mechanisms are only now becoming apparent. This new appreciation has come about not only through advances in the understanding of cell death in parallel immunological and pathological situations, but also through progress in developmental genetics which has revealed the roles played by death in the cell lineages of invertebrate embryos. In this review, we discuss programmed cell death as it is understood in developmental situations, and its relationship to apoptosis. We describe the morphological and biochemical features of apoptosis, and some methods for its detection in tissues. The occurrence of programmed cell death during invertebrate development is reviewed, as well as selected examples in vertebrate development. In particular, we discuss cell death in the early vertebrate embryo, in limb development, and in the nervous system.

Animals↗
PubMed1995

Chicken yolk contains bona fide high density lipoprotein particles.

Lipoproteins, the major nutrient source for developing embryos in egg-laying species, are thought to be transported from the circulation of the hen to the yolk of growing oocytes. In order to fully understand the contribution of the different lipoprotein species to oocyte growth, yolk formation, and embryo development, we have started to elucidate the relationships between the high density lipoproteins (HDL) in serum with the hitherto uncharacterized yolk HDL fraction. Immunoblotting with antibodies against apolipoprotein (apo) A-I, the major protein moiety of circulating HDL, revealed, for the first time, significant amounts of this protein in yolk. Importantly, yolk apoA-I was an integral component of bona fide lipoprotein particles: i) the apoA-I-containing particles could be purified by ultracentrifugal flotation and immunoaffinity chromatography on immobilized anti-apoA-I IgG; ii) the particles resembled serum HDL in ultrastructural, chemical, and biochemical aspects; and iii) in particular, these particles contained another major apolipoprotein, apo II. To date, apo II has been assumed to be unique to the very low density lipoprotein (VLDL) and HDL fractions of laying hen serum. Its residence on yolk HDL particles, together with the other results, strongly implies that yolk HDL, at least to a large part, is derived from serum. This implication is supported by the presence of apoA-I in oocytic coated vesicles. However, an oocyte plasma membrane receptor for the transport of HDL could not be identified; furthermore, immunoelectron microscopy demonstrated that yolk HDL particles do not colocalize with VLDL, known to be endocytosed via a specific receptor. Thus, these studies have revealed that HDL particles are taken up into the oocyte from the serum of the laying hen, and are deposited into the yolk by a mechanism distinct from that involved in the uptake of other yolk lipoproteins.

Animals↗
PubMed1994

Expression of TGF beta 1/beta 3 during early chick embryo development.

We have used an antibody against a TGF beta peptide fragment to localize this growth factor in the early chick embryo from laying to the ten-somite stage of development. Western blotting showed that the antibody reacted with both mammalian TGF beta 1 and chicken TGF beta 3. By immunocytochemistry we find that at the earliest developmental stage (stage X of Eyal-Giladi and Kochav) immunoreactivity to this antibody is primarily located in the cells of the area opaca and marginal zone, as well as in the most peripheral edge cells of the blastoderm. The yolk is non-reactive, except in a highly localized region subjacent to the edge cells. This pattern persists at stage XII, and at both stages individual isolated cells in the epiblast and hypoblast are also reactive. By the time of gastrulation, reactivity in the epiblast is polarized to the ventral extremity of the cells, and again some isolated cells in this layer are intensely immunoreactive. At this stage also, the endoderm cells, particularly those underlying the primitive streak, are positive, as are the mesoderm cells lateral to the streak. At somite stages, the neuroepithelium is not reactive but the ectoderm lateral to it is strongly positive. At the caudal primitive streak levels of early somite embryos, the ectoderm and endoderm are immunoreactive while the mesoderm loses the reactivity it showed at the early gastrulation stages. The neuroepithelial cells later show reactivity at their apical poles, and, as at the earlier stages, individual cells show intense labelling. These results indicate that TGF beta 1 and/or TGF beta 3 immunoreactivity is developmentally regulated from very early stages of morphogenesis in the chick, and together with data from earlier functional studies, suggest that this factor has roles in embryonic axis formation and in blastoderm expansion.

Animals↗
PubMed1994

Distribution of TNF alpha-like proteins correlates with some regions of programmed cell death in the chick embryo.

Early chick embryos have previously been shown to express tumor necrosis factor-alpha-cross-reactive proteins (TNF alpha-CRPs) in a developmentally regulated manner, thus implicating these proteins in programmed cell death and in tissue remodeling. In this study, cells undergoing DNA fragmentation have been identified, using terminal deoxynucleotide transferase (TdT) mediated dUTP-biotin nick-end-labeling (TUNEL), during the embryonic development of the chick, between stages 18 and 29. DNA fragmentation is indicative of cells undergoing programmed cell death. TUNEL-positive cells were identified in several well documented areas of programmed cell death, including the limb buds, the heart, spinal motoneurons, dorsal root ganglia, and the ventral horn of the neural tube. In addition, other areas of cell death were identified including the floor plate and the mesonephros. In several locations, a close correlation was noted between the presence of TUNEL-positive cells and regions of TNF alpha-immunoreactivity. These regions included the ventral horn and marginal zone of the neural tube, spinal motoneurons, paravertebral ganglia, parts of the myotome, mesenchyme of the body wall, and the mesonephros. In addition, using the TNF alpha-sensitive L929-8 bioassay it was shown that homogenate of stage 18 chick embryos is cytotoxic to L929-8 cells and that this toxicity can be reduced using neutralizing antibodies to mouse TNF alpha. This bioassay allowed us to estimate the mean concentration of TNF alpha-like activity in embryo homogenate, which is within the range of physiological (pg/ml) levels of TNF alpha found in other systems. These results suggest that proteins with TNF alpha-like activity may have a role in programmed cell death in some tissues during early chick embryo development.

Animals↗
PubMed1994

Guidance of filopodial extension by fibronectin-rich extracellular matrix fibrils during avian gastrulation. A study using confocal microscopy.

We have used double-label confocal microscopy to examine the relationships between the orientation of filopodial extension in mesoderm cells and the orientation of fibronectin-rich extracellular matrix fibrils during chick embryo gastrulation. We fluorescently labeled mesoderm tissue dissected from donor embryos by immersion in carboxyfluorescein and then grafted it into unlabeled host embryos at the same stage of gastrulation. After further incubation, the host embryos were fixed, the endoderm removed, and the extracellular matrix was immunostained with antibodies to fibronectin conjugated to Texas Red. We found that both the general shape of the mesoderm cells and the orientation of filopodial extension were influenced by the surrounding matrix fibrils. Elongated shape was associated with individual fibrils which impinge on only one side of the cell. Similarly, filopodial extension followed a single fibronectin-rich fibril, although filopodia were also observed to be channeled between pairs of parallel fibrils. Cells attached to non-aligned regions of substratum showed no polarity. The mesoderm cells themselves apparently synthesize their own fibronectin, and deposit this on the cell surface not attached to the substratum. We conclude that individual fibronectin-rich substratum fibrils, in the size range 0.7-2.8 microns, are able to exert contact guidance on the mesoderm cells, despite the production of endogenous fibronectin by the cells themselves. These results support the contention that contact guidance is a physiological mechanism influencing the orientation and directionality of cells during the morphogenetic movements of embryogenesis.

Animals↗
PubMed1994

Expression of the galactose-binding lectins during the formation of organ primordia in the chick embryo.

Early chick embryos contain two beta-galactoside-binding lectins of 16 kDa and 14 kDa. using several antisera to these proteins, we have studied lectin expression at embryonic stages when the segregation and early differentiation of organ primordia are taking place. With antisera to the 16 kDa lectin that display similar immunoreactivity in immunoblot analysis, we show that these antisera exhibit varying immunoreactivity in embryo sections. One antiserum reacts preferentially with a matrix form of lectin while another detects mainly a cellular form of this protein. During early development, galactoside-binding lectins of the matrix type are expressed in the vitelline membrane, the outer and inner limiting membranes of the neural tube, the surface of the notochord and the coelomic surface of the cardiac rudiments. The cellular form of the lectin occurs in the intracellular yolk of early embryos, in the primordial germ cells, the myocardium, in the early myotome, and in a cohort of cells which are presumed to belong to the neural crest. Our results indicate that, although all of the antisera recognize the intracellular lectin of the extraembryonic endoderm, some antisera to the 16 kDa lectin exhibit preferential reactivity with different lectin isoforms. The extracellular matrix form of lectin is transiently expressed during early development at the stages when the segregation of organ primordia is occurring. It's expression could be related to the acquisition of polarity in developing epithelia. Results also suggest that various versions of the same protein may perform distinct developmental roles in the embryo.

Animals↗
PubMed1993

Expression of tumor necrosis factor-alpha (TNF alpha)-cross-reactive proteins during early chick embryo development.

We have investigated the expression of tumor necrosis factor-alpha (TNF alpha)-cross-reactive proteins during the early development of the chick embryo from day 1 to day 6 (H-H stages 5-29) using a polyclonal antibody and two monoclonal antibodies to recombinant mouse TNF alpha. We have confirmed the cross-reactivity of the antibodies with chicken tissue in Western blotting studies. Proteins of 50 kDa and 70 kDa, showing anti-TNF alpha cross-reactivity, have been identified during early chick development. In addition, both monoclonal antibodies recognize a 120 kDa protein. These molecules probably represent cytosolic or transmembrane TNF-alpha-like proteins, similar to those previously identified on the surface of cytotoxic T-lymphocytes. We show by ultrastructural cytochemistry that immunoreactivity can be detected at the surfaces of some cells, suggesting that at least some of the antigen is membrane-associated. The proteins are shown to have a widespread tissue distribution during this period of development. Immunoreactivity is first detected in the gastrulating embryo, in the mesoderm and the endoderm. By day 2, expression is confined to the ectoderm and the endoderm, while at day 3 expression appears in the myotome, the notochord, and in nervous tissue. At day 4 the distribution of reactivity is more extensive and includes the notochord, the sclerotome, and the myotome, while the cranial and spinal nerves also become intensely immunoreactive. Also at this stage, neural tube reactivity becomes localized to the marginal neuroepithelial zone, and the lens fibers become positive. This distribution of staining then persists until 6 days of development. We hypothesize that the expression of TNF alpha-cross-reactive proteins in early development could be indicative of a role for them in programmed cell death (apoptosis) during differentiation of the notochord, the lens, and the nervous system, and in tissue remodeling.

Animals↗
PubMed1993

Immunocytochemical localization of sodium channels in an insect central nervous system using a site-directed antibody.

Antibodies to channel proteins and specific peptide sequences have been previously used to localize voltage-activated sodium channels in the rat brain. Here we describe the first localization of sodium channels in an insect nervous system using a site-directed antibody. The mesothoracic ganglion of the cockroach was stained with an antibody to the highly conserved SP19 sequence. Antibody labelling was visualized by light microscopy using the avidin/biotin method on wax sections, and transmission electron microscopy of immunogold-labelled thin sections. Central ganglia of insects contain clearly separated regions of cell bodies, synaptic neuropil, axon tracts, and nerves. Antibody staining by light microscopy was limited to neurons, and was intense in axons throughout the ganglion and nerves. Staining was also strong in the cytoplasm, but not the nuclei, of many neuronal cell bodies. Neuropil regions were relatively lightly labelled. These findings can be correlated with the known electrophysiology of the ganglion. Electron microscopy detected sodium channels in areas surrounding axons, probably including axon membranes and enveloping glial cell membranes. Axonal mitochondria were also heavily labelled, suggesting a sodium channel transport function for these organelles.

Amino Acid Sequence↗
PubMed1993

The involvement of TGF beta 1 in early avian development: gastrulation and chondrogenesis.

We examined the effects of transforming growth factor-beta 1 (TGF beta 1) and a neutralizing monoclonal antibody on two phases of early chick embryo development: gastrulation and chondrogenesis. We carried out experiments in vivo and in vitro on mesoderm cells from the gastrulating embryo at day 1, and on sclerotome cells from day 3 embryos, having previously shown that this factor is present among these cells at these stages of development. Addition of the antibody to cultures of these cells produced a dose-dependent decrease in cell out-growth and spreading and concomitantly reduced fibronectin deposition. In vivo studies of the effects of TGF beta 1 on mesoderm during gastrulation were carried out by grafting beads carrying this agent into gastrulating embryos. We used beads of ion-exchange resin as well as hydrolysed polyacrylamide, and found that the grafts produced an accumulation of mesoderm cells around the implant and, at later stages, the formation of enlarged somites. There was no effect on embryonic axis formation. Studies of bromodeoxyuridine (BrdU) incorporation indicated that the mesoderm accumulation was due, at least in part, to an increase in cell proliferation. However, examination of the effect of TGF beta 1 on BrdU incorporation by mesoderm during gastrulation and sclerotome cells in vitro indicated in inhibition of cell proliferation, an inconsistency explained in terms of the variation between the in vivo and in vitro conditions. We conclude that TGF beta 1 is both appropriately located, and is able, to influence cell proliferation among the mesodermal cell populations during early development, and that this effect contributes to the overall control of mesodermal morphogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗