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P Thorogood

Publications and source records attributed to P Thorogood.

At least 37 records · Page 2Linked to original sources
PubMed1994

Patterns of cell behaviour underlying somitogenesis and notochord formation in intact vertebrate embryos.

We have made a detailed analysis of cell behaviour using high resolution time-lapse microscopy of the earliest cellular interactions taking place during morphogenesis of the notochord and somites in intact teleost embryos. Notochord formation is typified by active intercalation of paraxial mesenchyme cells into the lateral surfaces of the primordium. Following this recruitment phase, complete immiscibility develops between cells of the notochord and the presomitic mesenchyme. Dorso-ventral and rostro-caudal expansion of the notochord is characterised by translocation of cells within dorso-ventral planes of section and is supported by elongation of the remaining cells and reduction in width across its latero-medial axis. A lateral palisading of paraxial mesenchyme against the lateral aspects of the notochord precedes overt segmentation. Intersomitic furrows form by localised de-adhesion at small foci at the nascent intersomitic planes, which are consolidated by coalescence of such areas by de-adhesion to produce the interface. It is not possible to predict precisely where cells would initiate de-adhesion since there is a stochastic element to the phenomenon. Once formed, boundaries between somites are stable and provide no opportunity for mixing, except across the first formed furrow, which disintegrates at the 4-6 somite stage. The first ten somites form at a constant rate of 2.3 somites/hr, during which time we recorded constant relative displacement of the segmental plate against the rostro-caudally elongating notochord. Unlike teleost epiboly and gastrulation, no large-scale movements of individual cells can be detected during elaboration of the embryonic axis.

Animals↗
PubMed1994

Behaviour of fibroblasts during initial attachment to a glass-ceramic implant material in vitro: a time-lapse video-micrographic study.

The in vitro behaviour of 3T3 fibroblasts plated on a glass-ceramic implant material, Apoceram, and on tissue culture polystyrene (TCP) was monitored using phase-contrast, time-lapse video microscopy and scanning electron microscopy. Significant differences were observed in the short-term cellular response to TCP and Apoceram in the timing of the fibroblasts' morphological changes, with earlier spreading and stabilization of the cells occurring on TCP. Cell surface ruffling activity was greater on the glass-ceramic, reflecting weaker adhesions to the substratum. The experimental method used facilitates direct analysis of the dynamic activity of cells attaching to artificial substrata.

Animals↗
PubMed1994

Distribution of, and a putative role for, the cell-surface neutral metallo-endopeptidases during mammalian craniofacial development.

Endopeptidase-24.11 (neutral endopeptidase, neprilysin, 'enkephalinase', EC 3.4.24.11) and endopeptidase-24.18 (endopeptidase-2, meprin, EC 3.4.24.18) are cell-surface zinc-dependent metallo-endopeptidases able to cleave a variety of bioactive peptides including growth factors. We report the first study of the cellular and tissue distribution of both enzymes and of the mRNA for NEP during embryonic development in the rat. Endopeptidase-24.11 protein was first detected at E10 in the lining of the gut and, at E12, the enzyme was present on the notochord, medial and lateral nasal processes, otocyst, mesonephros, heart and neuroepithelium. In contrast, at this time endopeptidase-24.18 was present only on the apical surface of the neuroepithelial cells. By E14 and E16, NEP was also detected in a wide range of craniofacial structures, notably the palatal mesenchyme, the choroid plexus, tongue and perichondrium. The distribution of endopeptidase-24.18 at these stages was restricted to the inner ear, the nasal conchae, and ependymal layer of the brain ventricles and the choroid plexus. Although endopeptidase-24.11 had been detectable in the craniofacial vasculature at E12 and E14, this was no longer apparent at E16. Significantly, the distribution of endopeptidase-24.11 mRNA closely matched the immunolocalization of the protein at all stages investigated. In order to explore the functional role of these enzymes, inhibition studies were carried out using two selective inhibitors of endopeptidase-24.11, phosphoramidon and thiorphan. E9.5 and E10.5 embryos exposed to either inhibitor displayed a characteristic, asymmetric abnormality consisting of a spherical swelling, possibly associated with a haematoma, predominantly on the left side of the prosencephalon, and the severity of this defect appeared to be a dose-dependent phenomenon. This study suggests that these enzymes play previously unrecognized roles during mammalian embryonic development.

Animals↗
PubMed1994

Regenerative capability of upper and lower jaws in the newt.

The regenerating amphibian jaw represents an important model for studying pattern formation and the mechanisms underlying regeneration of facial structures. We have studied regeneration of upper and lower jaws in the urodele amphibian, Notophthalmus viridescens, using whole mount preparations stained for bone and cartilage, scanning electron microscopy and immunocytochemistry to further characterize these regenerating systems. In addition, we have investigated whether lower jaws of adults and larvae display similar regenerative ability. Although in adult animals the original shape of both the lower and upper jaws is rather faithfully reproduced following amputation, and the teeth and oral mucosa with its specialized sensory organs fully regenerate, significant differences in the regenerative ability of the various skeletal elements are observed. In fact, only tooth-bearing skeletal elements ossify, while the other elements of the regenerated skeleton remain cartilaginous for as long as 5 months after amputation. In contrast, a regenerated lower jaw in the larva is indistinguishable from an unamputated one at the same stage of development. Interestingly, regenerating adult jaws form directly bicuspid teeth, which are the type of teeth normally found in the adult, rather than the monocuspid teeth characteristic of larval jaws, indicating that jaw regeneration is not a recapitulation of development, in that an adult jaw blastema directly regenerates an adult jaw. Finally, we have studied the expression of tissue specific markers in normal and regenerating upper and lower jaws to establish whether the blastemal cells, which will form the missing part of the jaw, express any of these markers of the differentiated state, or are undifferentiated as suggested by their morphological appearance. Under our experimental conditions, no expression of markers of the differentiated state, such as those for muscle, cartilage and glands is detectable in early regenerates. On the contrary, the mesenchymal marker 22/31, whose expression in normal jaws is restricted to dermal fibroblasts and the dental pulp, is expressed in at least a half of the blastemal cells. The significance of these observations in relation to the origin of blastemal cells in the jaw will be discussed.

Animals↗
PubMed1992

Heads and tales: recent advances in craniofacial development.

In the last few years, our understanding of the complex series of events governing craniofacial morphogenesis has significantly increased. A variety of experimental approaches, such as cell lineage marking, monoclonal antibodies, organ culture and recombinant DNA techniques, have been fundamental to this progress. The neural crest contribution to the head and face of the embryo has been mapped, the significance and mechanisms of epithelial-mesenchymal interactions are better understood, and a number of regulatory molecules, which are likely to be causally involved in mediating the morphogenesis of the different craniofacial regions, have been identified. Although the information presently available on the molecules involved is far from complete, and the experimental strategies need further refinement, the existence of specific combinatorial gene-codes (ie the homeobox-containing gene code) has become evident. These findings are proving invaluable to our understanding of the mechanisms underlying craniofacial development, and of certain syndromes affecting craniofacial structures in humans.

Animals↗
PubMed1992

Harderian gland hyperplasia in c-mos transgenic mice.

Transgenic mice carrying the mouse mos proto-oncogene linked to a retroviral LTR develop hyperplasia of the Harderian glands. Enlargement of the glands is evident as early as 18 weeks after birth, with glands reaching up to 10 times their normal weight. Approximately 65% of the cases of hyperplasia occur bilaterally, and the majority of mice affected are male (66%). Elevated levels of mos expression are found in all Harderian glands of mice from the affected transgenic line, but not in glands of normal mice or a non-affected transgenic line, indicating that hyperplasia is dependent on mos expression. Histological examination of the tissue reveals a general involvement of the entire gland epithelium in hyperplastic growth, with no evidence of focal or malignant tumours. These observations show that in addition to neu, myc, ras and ret transgenes, mos, a member of the protein-serine/threonine kinase family of oncogenes, can induce Harderian gland hyperplasia, thus revealing an unusual response by this organ to various classes of oncogenes. Analysis of fos, jun, myc and ets oncogene RNA in mos-induced hyperplastic Harderian glands shows that there are no consistent changes in the level of expression of these oncogenes, suggesting that mos acts via a mechanism other than by increasing the expression of these genes.

Animals↗
PubMed1992

Monoclonal antibodies raised against pre-migratory neural crest reveal population heterogeneity during crest development.

In order to address the problem of when heterogeneity arises within premigratory and early migratory neural crest cell populations, mouse monoclonal antibodies were raised against quail premigratory neural crest. Due to the limited availability of immunogen an intrasplenic route for immunization was used. Three monoclonal antibodies (referred to as LH2D4, LH5D3 and LH6C2) were subsequently isolated which recognized subpopulations in 24 h cultures of both quail and chick mesencephalic and trunk neural crest in immunocytochemical studies. Subsequent investigations using a range of six antibodies, including LH2D4, LH5D3 and LH6C2, showed that population heterogeneity (which was not cell cycle related) could be detected as early as 15 h following mesencephalic crest explantation, a stage at which all the neural crest cells were morphologically identical. However, premigratory neural crest from the same axial level of origin was homogeneous, as judged by immunoreactivity patterns with these antibodies. Significant differences were found in the proportion of immunoreactive cells between populations of mesencephalic and trunk neural crest cultures. Double immunofluorescence studies revealed the existence of at least four separate cell populations within individual crest cultures, each identified by their unique antibody reactivity pattern, thus providing some insight into the underlying complexity of subpopulation composition within the neural crest. Immunocytochemical studies on quail embryos from stages 7-22 showed that the epitopes detected by LH2D4, LH5D3 and LH6C2 were not necessarily confined to the neural crest or to cells of crest derivation. All three epitopes displayed a spatiotemporal regulation in their expression during early avian ontogeny. Since the differential epitope expression described in this investigation was detectable as early as 15 h after premigratory neural crest explantation, took place in vitro in the absence of any other cell type and changed progressively with time, we conclude that a certain degree of population heterogeneity can be generated very early in neural crest ontogeny and independently of the tissue interactions that normally ensue in vivo.

Animals↗
PubMed1992

Hypermedia in biomedical education: a case study.

The origination, development and application of a hypermedia system for use in the life sciences is described. Implemented in the Apple Macintosh HyperCard environment, the project has been designed to make use of commercially produced laser discs as visual archives and has included the production of authoring and navigational tools. We report here on our experience in authoring stacks for undergraduate teaching, monitoring of student use and assessment of effectiveness as a self-teaching resource. Future developments and applications within the biomedical sciences are discussed.

Biological Science Disciplines↗
PubMed1991

The transient expression of type II collagen at tissue interfaces during mammalian craniofacial development.

Using immunocytochemical techniques, the spatiotemporal distribution of the major collagen isoform of cartilage, type II collagen, has been investigated during early craniofacial development in the mouse embryo. Early and transient expression was associated with the otic and optic vesicles, the ventrolateral surfaces of the developing brain, olfactory conchi, endocardial and mesocardial tissues, the lateral and basal surfaces of the pharyngeal endoderm and beneath the ectoderm of the branchial arches. A number of these locations are sites of epithelial-mesenchymal tissue interaction believed to generate the component parts of the chondrocranium; here, type II collagen appears transiently in advance of overt chondrogenesis in the mesenchyme. At such sites, immunofluorescence is typically localised along the basal surface of the epithelial partner, with the strongest reaction detected between the basal aspects of the otic and rhombencephalic epithelia. Immunoelectron microscopy, using pre-embedding immunostaining and a protein G-gold technique, reveals that the type II collagen is adjacent to, but not integral with, the basal laminae. Gold particles are clearly associated with 10-15 nm fibrils of the extracellular matrix in the reticulate lamina region. The pattern of type II collagen expression in the mouse closely correlates with that demonstrated previously in the quail, indicating a high degree of phylogenetic conservation between these two vertebrate species. These findings are consistent with the hypothesis that the pattern of epithelial secretion of type II collagen, or a coexpressed matrix molecule, constitutes a morphogenetic signal, realised as a matrix-mediated tissue interaction, and specifying the form of the vertebrate chondrocranium. Three-dimensional reconstruction of early type II collagen distribution, and of the subsequent chondrocranial cartilages, reveals that chondrocranial form can be derived from a 'pre-pattern' of epithelially derived type II collagen expressed at epithelial-mesenchymal tissue interfaces.

Animals↗
PubMed1988

The developmental specification of the vertebrate skull.

The initial form of the embryonic bony skull is determined in two ways; cranially, by the relative growth of the developing brain, and facially, by the chondrocranium. Both are essentially acting as structural templates around which the bony components of the skull are assembled. Assuming, therefore, that the specification of form and pattern in the facial skeleton occurs at the formation of the chondrocranium, this paper will focus on precisely how the chondrocranium forms. Any acceptable explanation of chondrocranial morphogenesis must satisfy at least two prerequisites. First, given the constancy of chondrocranial form in vertebrates, any model proposed should be equally applicable to all vertebrates. Second, it should enable us to answer questions of homology concerning the skull and, in particular, provide explanation for those instances where 'homologous' structures have a different (lineage) composition. From studies limited to a small number of amphibian, avian and mammalian species, it is apparent that chondrogenesis in the vertebrate skull is largely, if not entirely, elicited by epitheliomesenchymal tissue interactions. Analysis of such interactions (and of those promoting osteogenesis) reveals that these are matrix-mediated and, recently, the expression of certain 'relevant' matrix components has been shown to be developmentally regulated in a fashion that correlates with the location and timing of these interactions. From these, and related, observations a morphogenetic model, the so-called 'Flypaper Model', has been proposed to explain the specification of chondrocranial form. A number of predictions arising from that model are currently being tested experimentally and the current status of the model is reviewed. Finally, the ability of this model to satisfy the prerequisites defined above is assessed.

Animals↗
PubMed1987

Autonomy of differentiation in avian branchial somites and the influence of adjacent tissues.

This study investigates the differentiative abilities of avian brachial somites at stages of development before, during and after the migration of somitic cells into the wing primordium. These somites are the source of cells that migrate into the forelimbs and there give rise exclusively, and totally, to the skeletal muscle lineage of the wing and yet show no morphological evidence of commitment to that fate when they leave the somites. The aim of the study was to see if the brachial somitic cells are committed to particular developmental pathways at these stages. The brachial somites were isolated from HH stage-12, -15 and -18 chick embryos, either by microdissection or enzymatic dissociation, and grown in organ culture, in explant culture on different substrata or on the chorioallantoic membrane (CAM) of host chicks, either alone or in combination with adjacent tissues. Myogenesis and chondrogenesis occurred in all stage-18 enzymatically separated somites, regardless of the growth environment. Myogenesis was reduced in stage-15 somites and unobservable in stage-12 somites; however, recombination of stage-12 somites with epithelium or neural tube increased the incidence of myogenesis at this stage. The incidence of chondrogenesis was also less in the younger explants. Unlike its effect on myogenic expression, recombination with epithelium resulted in a dramatic decrease in chondrogenesis in both stage-12 and -15 somites. The recombination experiments suggest that conditions that maintain the normal spatial relationships within isolated somites permit expression of a preexisting specification to a particular fate. They also show that the overlying epithelium can inhibit chondrogenesis in these somites. Overall, the results suggest that by the time migration of somitic cells into wing regions is finishing, brachial somitic cells have become stabilized in their ability to undergo both myogenesis and chondrogenesis for they will do so under a variety of growth conditions and independently of adjacent tissues. However, immediately before (stage 12) and shortly after (stage 15) the onset of migration, both myogenic and chondrogenic expression by brachial somitic cells are still under the influence of interactions with adjacent tissues.

Animals↗
PubMed1987

Analysis of in vivo cell movement using transparent tissue systems.

The embryos of certain teleost species are transparent and cell behaviour within the intact embryo can be observed and recorded using Nomarski microscopy coupled with time-lapse video recording or time-lapse cine filming. In this report we review some of our recent analyses of cell behaviour patterns underlying key morphogenetic events. (1) Contact-guided cell migration through a structurally ordered extracellular matrix during fin development; (2) movement of tissue layers during epibolic overgrowth; and (3) cell 'social' behaviour during the establishment of the body axis (i.e. notochord formation and somitogenesis). These results, on cell behaviour correlated with normal morphogenesis, provide a baseline for further work in which hypotheses concerning subcellular and molecular controls of cell behaviour can be tested by experimental perturbation in vivo.

Animals↗
PubMed1986

Transient expression of collagen type II at epitheliomesenchymal interfaces during morphogenesis of the cartilaginous neurocranium.

In the avian embryo a matrix-mediated tissue interaction between retinal pigmented epithelium and neural crest-derived periocular mesenchyme leads to the differentiation of (scleral) cartilage. The composition of the extracellular matrix at the interface between these two tissues has been examined immunohistochemically, both during and after the interaction has taken place. Of the matrix components studied (fibronectin, laminin, and collagen types I, II, IV, and V) only collagen type II displayed a dramatic change in distribution between the two stages. During the interaction, at stage 15, type II was present in the extracellular compartment basal to the epithelium. After completion of the interaction, collagen type II was no longer detectable at the interface even though it was readily detectable in the vitreous humor, cornea, and perinotochordal sheath, and subsequently will be expressed by the chondrogenic tissue itself as overt differentiation commences. These results suggest that collagen type II might be causally involved in this particular epitheliomesenchymal interaction. Examination of the spatial and temporal patterns of collagen type II expression elsewhere in the developing craniofacial complex revealed a hitherto unreported pattern of distribution. In addition to its predictable locations (i.e., cornea, vitreous, and perinotochordal sheath) it was found to be present at certain other sites, for example, at the basal surfaces of some neuroepithelia. These additional locations are all known to be sites of chondrogenesis-promoting tissue interactions which result in the formation of the elements of the cartilaginous neurocranium (e.g., otic vesicle). Furthermore this spatial distribution exhibits a changing temporal pattern in that it is detectable at the time that the interactions are known to be taking place, but subsequently is no longer detectable by the immunohistochemical means employed. This definable pattern of transient collagen type II expression, occurring at very early stages of craniofacial development, is interpreted as reflecting one level of morphogenetic specification of chondrocranial/skull form in the developing vertebrate head.

Age Factors↗
PubMed1986

Specific thromboxane synthetase inhibition and retinopathy in insulin-dependent diabetics.

We have previously reported that specific thromboxane synthetase inhibition may be associated with a reduction in albumin excretion rate in insulin-dependent diabetics (IDD). We now report studies of retinal morphology assessed by serial fluorescein angiography in 25 insulin-dependent diabetics during a 16-week double-blind, randomized, placebo-controlled study of the specific thromboxane synthetase inhibitor UK-38,485. Assessment of angiograms was by, computerized image analysis indicating the percentage area of vascularization and, panel scoring by 4 ophthalmologists who scored the posterior pole in each of the 4 angiograms "blind" according to a prearranged system. There was no significant change in percentage area of vascularization in either placebo or "active" group during the study. Panel scoring, however, suggested that some patients deteriorated whilst others remained unchanged or improved. There was, however, no significant difference in these parameters between those subjects on UK-38,485 and those on placebo. We conclude that specific thromboxane synthetase inhibition is not associated with any significant change in diabetic retinopathy over a 4-month period of study.

Adult↗