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Biomedical subjects

D A Rees

Publications and source records attributed to D A Rees.

At least 19 recordsLinked to original sources
PubMed1979

Fibronectin distribution in epithelial and associated tissues of the rat.

Specific antiserum was used to investigate the distribution of the extracellular glycoprotein, fibronectin, in rat skin and tongue tissue by light and electron microscopy with immunofluorescence and immunoperoxidase techniques. We conclude that fibronectin is absent from stable, differentiated parts of tissues, such as the sebaceous glands or the matrix, medulla, cortex, and cuticles of the hair and the inner and outer root sheaths, or even in tissues in which there is some cell movement, such as the epidermis. It is, however, characteristic of sites at which cell division is occurring in contact with an extracellular scaffolding, such as basement membrane or loose connective tissue. Conspicuous examples were in the glassy membrane and connective tissue sheath associated with the follicular epithelium, the basement membrane underlying vascular endothelial cells, the connective tissues surrounding and investing nerve and muscle fibre bundles, and the dermal connective tissue where fibronectin was often associated closely with collagen fibres. At the basement membrane of the dermal/epidermal junction, fibronectin occurred at the plasma membrane of the basal cells and in the lamina lucida area. There was no correlation with specific areas of cell-substrate adhesion, such as the hemidesmosomes. The endoplasmic reticulum of fibroblasts stained strongly suggesting that these cells represent a major site of synthesis.

Animals↗
PubMed1979

Actomyosin organisation for adhesion, spreading, growth and movement in chick fibroblasts.

Examination of the actomyosin structures and their relation to adhesion, movement and growth in the first fibroblasts migrating from chick heart explants shows striking differences with fibroblasts adapted to grow in culture. The latter have focal adhesions which seem to immobilize them for anchorage-dependent growth, rather than facilitate their movement. The fibroblasts specialized for movement from the explants, though equally well spread, make contact with substratum through extensive areas of relatively unspecialized membrane, have less well developed stress fibres and a low growth rate.

Actomyosin↗
PubMed1979

Solution conformation of glycosaminoglycans: assignment of the 300-MHz 1H-magnetic resonance spectra of chondroitin 4-sulphate, chondroitin 6-sulphate and hyaluronate, and investigation of an alkali-induced conformation change.

Complete assignments are given for the 1H nuclear magnetic resonance (NMR) spectra at 300 MHz of chondroitin 4-sulphate, chondroitin 6-sulphate and hyaluronate in deuterium oxide solution, supported by spin decoupling and computer simulation. Coupling constants and chemical shifts are as expected from spectra of the model glycosides, methyl beta-D-glucopyranosiduronate, methyl 2-acetamido-2-deoxy-beta-D-glucopyranoside and methyl 2-acetamido-2-deoxy-beta-D-galactopyranoside, when allowance is made for systematic influences on chemical shifts of interglycosidic linkages and sulphate substitution. As reported elsewhere, addition of alkali causes the hyaluronate spectrum to sharpen considerably. This is taken to indicate that segmental motion is enhanced by disruption of some system of inter-residue bonding on ionisation of hydroxy groups. Concomitant changes in chemical shifts are seen mainly for H-2 of the glucuronate residue, and the CH3 and H-2 of the acetamidodeoxyglucose residue. Similar effects are not seen for chondroitin sulphates, either in line widths or chemical shifts. Comparison of the spectra of hyaluronate, chondroitin sulphates, and the model glycosides, indicates that proton chemical shifts are sensitive to the conformation differences between the polysaccharides in alkaline solution, but do not detect the differences in neutral solution that are known from NMR relaxation to be present. The altered configuration and/or substitution pattern of the acetamidodeoxyhexose residue in hyaluronate compared with chondroitin sulphates appears to have a critical influence on overall conformation in both alkaline and neutral solution.

Chondroitin↗
PubMed1979

1H-Nuclear magnetic-resonance studies on glycophorin and its carbohydrate-containing tryptic peptides.

The proton nuclear magnetic resonance (1H-NMR) spectra of glycophorin and its tryptic sialoglycopeptides were investigated. From the intensities of the assigned resonances it was concluded that all of the residues in the sialoglycopeptides are sufficiently mobile in conformation to give sharp resonances, while in glycophorin this is true for only approximately 80% of the peptide backbone. The resonances of the central sequence of some 20 of the hydrophobic residues are strongly broadened. This region is probably that of alpha-helical structure which is known to aggregate. The linewidths and intensities of the resonances are not, or only slightly, affected by changing the ionic strength, temperature or by carboxymethylation of the Met-81 residue in glycophorin. Glycophorin was found to bind about 100 mol sodium dodecylsulphate/mol protein as derived from studies on linebroadening of the latter's C-3 to C-11 methylene resonances. The bound dodecyl-sulphate probably increases the mobilities of the hydrophobic residues in the protein as these resonance intensities are increased by the binding. The carbohydrate chains in glycophorin were conformationally mobile; no evidence was found for tight carbohydrate-protein interactions. The relevance of flexible carbohydrate chains in membrane glycoproteins is discussed in relation to cell surface chemistry.

Glycopeptides↗
PubMed1979

Mechanisms of cellular adhesion. IV. Role of serum glycoproteins in fibroblast spreading on glass.

We have investigated the exogenous factors required for the transition from the round shape of suspended fibroblasts to the characteristic spread shape on serum-coated glass. Following the evidence of others that the transition is facilitated by adsorbed component(s) related to CIG/LETS (cold-insoluble globulin/large external transformation-sensitive) proteins, we have isolated 2 such preparations from chick serum. Their influence has been investigated on fibroblast adhesion, spreading and growth and they have been characterized by gel electrophoresis, immunological cross-reactivity, amino acid and carbohydrate residue analysis, sedimentation velocity behaviour and circular dichroism spectroscopy. The preparations have molecular weights of 225/215,000 and 140,000 Daltons respectively and are closely similar in composition and secondary structure. The 225/215 000 Dalton doublet is probably a product of limited proteolysis which almost certainly occurred in the avian circulation. For cells seeded on glass precoated in different ways and in different supplemented media we could detect no change in the extent of attachment but there were profound influences on cell shape following this initial adhesion. We confirm that prior adsorption of either CIG-related preparation to glass does indeed promote fibroblast spreading in the absence of other serum components and that CIG is the sole serum component with this type of activity. We now add 2 important qualifications: (i) the presence of substrate-adsorbed serum CIG is not essential for spreading when other serum components are present in the medium; and (ii) the adhesive organization shown by interference reflexion microscopy is incompletely formed unless the additional serum components are present in the medium. We therefore conclude that 16C fibroblasts have the ability when given the stimulus of soluble serum components other than CIG, but not otherwise, to synthesize all the components necessary for the highly organized contacts with glass, including endogenous CIG/LETS proteins.

Amino Acids↗
PubMed1979

Co-operativity of lectin binding to fibroblasts and its relation to cellular actomyosin.

We have investigated the binding of 125I-concanavalin A (125I-Con A) and 125I-succinyl concanavalin A (125I-s-Con A) to rat fibroblasts (16C line) as a function of the concentration of added lectin, and the alterations to this binding behaviour caused by drugs which modify the cytoskeleton. The changes in cell behaviour which occur at different levels of binding have also been studied. As shown previously for some other systems, the binding of Con A is complex and partly co-operative. Three phases can be distinguished in our system: (i) pre-nucleation binding, (ii) binding which shows a small positive slope in a Scatchard plot and a Hill coefficient greater than unity, and which therefore is incipiently co-operative, and (iii) post-co-operative binding. The co-operative phase of binding is paralleled by progressive inhibition of EGTA-mediated cell detachment from substrata, with inhibition being complete when this phase of binding is complete. Likewise, the phagocytosis of latex spheres is progressively inhibited up to a threshold which coincides with the completion of co-operative binding. Thirdly, cells pretreated with Con A round up with colchicine (10(-5) M) if co-operative finding is complete, but adopt broad epithelial shapes if it is not. s-Con A does not show cooperative binding, and correspondingly does not inhibit EGTA-mediated cell detachment, or show a distinct threshold in the inhibition of phagocytosis, or promote the 2 types of shape change with colchicine. The pattern of Con A binding is drastically altered by pretreatment of cells with cytochalasin B or azide. The Scatchard and Hill plots show that the co-operative phase remains and is complete at about the same level of binding, but that it is more readily nucleated and takes place against a changed number and/or distribution of receptors. Pretreatment of cells with colchicine causes changes in the pattern of binding which are different from those observed with cytochalasin B or azide and are more difficult to interpret. We conclude that a reciprocal relationship exists between the cellular actomyosin and the state of cell surface receptors. Perturbation of actomyosin by cytochalasin B or azide can enhance the freedom of some receptors to participate in a co-operative rearrangement which facilitates the binding of further molecules of lectin. Vice versa, the co-operative event has a feedback influence on the cellular actomyosin to cause alterations of cellular response.

Actomyosin↗
PubMed1979

The behaviour of fibroblasts migrating from chick heart explants: changes in adhesion, locomotion and growth, and in the distribution of actomyosin and fibronectin.

Fibroblasts migrating from heart explants of chick embryos at first have a high rate of locomotion but lack focal contacts or adhesions and also lack substantial actin-containing bundles. A meshwork of 7-nm filaments is present particularly in submembranous regions and is proposed to be directed towards efficient locomotion whilst maintaining a high degree of spreading. Also during the first 48 h there is little production of extracellular fibronectin and the growth rate is low. Later, these fibroblasts develop focal contacts and focal adhesions together with actomyosin bundles, with a parallel increase in fibronectin expression. We propose that progressive immobilization by the development of focal adhesions and actomyosin structures occurs to set these cells up for growth.

Actomyosin↗
PubMed1977

Rapid isolation of plasma membranes in high yield from cultured fibroblasts.

1. A method was developed which allows the rapid preparation of pure plasma membranes in high yield from cultured fibroblasts. 2. Cells are lysed in hypo-osmotic borate/EDTA and, after differential centrifugation, the membranes collected by centrifugation on a sucrose barrier. 3. Electron microscopy of the isolated material shows large membrane vesicles essentially free from contaminating organelles. 4. There is no detectable activity of the endoplasmic-reticulum enzyme marker, NADH2--lipoamide oxidoreductase (EC 1.6.4.3), and that of succinate dehydrogenase (EC 1.3.99.1), a marker for mitochondria, is substantially decreased. Chemical compositions are in good agreement with previous observations. 5. This study confirms the usefulness of applied isotopic markers for isolating plasma membranes.

Borohydrides↗