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Biomedical subjects

N A Ratcliffe

Publications and source records attributed to N A Ratcliffe.

At least 37 records · Page 2Linked to original sources
PubMed1993

Detection, isolation and characterization of multiple lectins from the haemolymph of the cockroach Blaberus discoidalis.

Three agglutinins (lectins), designated BDL1, BDL2 and BDL3, were identified in the haemolymph of the cockroach Blaberus discoidalis by erythrocyte cross-adsorption and sugar inhibition tests. With the use of (NH4)2SO4 fractionation, anion-exchange and affinity chromatography, BDL1 and BDL2 have been purified to homogeneity, and BDL3 has been partially purified to three bands on SDS/PAGE. BDL1 has a molecular-mass estimate of 390 kDa by gel filtration and approx. 158 kDa by SDS/PAGE under non-reducing conditions, further reduced to subunits of 36 kDa under reducing conditions. BDL2 has a molecular mass of approx. 140 kDa and is composed of subunits of 67 kDa which can be further reduced to identical subunits of 23 kDa. Isoelectric focusing in agarose gels revealed that BDL1 and BDL2 both focused as single bands at pH 6.0 and pH 5.2 respectively. The purified forms of BDL1 and BDL2 were stained by the periodic acid/Schiff's reagent showing that both lectins are glycoproteins. In addition, BDL1 was deglycosylated by endo-beta-N-acetylglucosaminidase H. Immunological tests showed that these three lectins are not structurally related. All three lectins bind galactose but have different specificities for binding other sugars and for a range of vertebrate erythrocytes. BDL1 is specifically inhibited by D-(+)-glucose, D-(+)-mannose and N-acetyl-D-mannosamine, but not by N-acetyl-D-glucosamine, and BDL2 is inhibited by N-acetyl-D-glucosamine, but not by D-(+)-glucose, D-(+)-mannose or N-acetyl-D-mannosamine. BDL3 is strongly inhibited by N-acetyl-D-galactosamine, but not by any of the other above-mentioned sugars. Erythrocyte specificities showed that BDL1 is more specific for rabbit than mouse erythrocytes, whereas BDL2 and BDL3 are more specific for mouse than rabbit erythrocytes. The haemagglutinating activities of both the serum and isolated lectins are Ca(2+)-dependent. Localization of BDL1 and BDL2 with fluorescein isothiocyanate-labelled antibodies showed that both lectins are associated with the granules and other areas of the cytoplasm of all blood cell types.

Animals↗
PubMed1993

Purification of the pro-phenol oxidase enzyme from haemocytes of the cockroach Blaberus discoidalis.

Pro-phenol oxidase was purified from the haemocytes of the cockroach Blaberus discoidalis by Blue Sepharose chromatography, hydrophobic-interaction chromatography on a Phenyl-Superose column and, finally, gel filtration on a Superose 6 column. Results suggest that the molecule exists as a polymer of identical 76 kDa monomeric units. The enzyme is a glycoprotein with pI of 5.2 and can be converted by trypsin into phenol oxidase.

Animals↗
PubMed1991

Activation of the prophenoloxidase cascade and initiation of nodule formation in locusts by bacterial lipopolysaccharides.

The activation of the prophenoloxidase (proPO) system of the locusts, Schistocerca gregaria and Locusta migratoria, by several bacterial lipopolysaccharides (LPS) is described. Activation of proPO by LPS occurred only in the presence of whole blood homogenates and not with hemocyte lysate preparations alone. Levels of phenoloxidase generated by the different LPSs in vitro were also correlated with numbers of nodules formed in vivo by injection of these LPSs. This further strengthens the evidence for the involvement of proPO activation in the insect cellular defenses. Finally, the wisdom in using anticoagulants in order to stabilize fragile hemocytes in studies on the proPO system is discussed.

Animals↗
PubMed1991

The prophenoloxidase system and in vitro interaction of Trypanosoma rangeli with Rhodnius prolixus and Triatoma infestans haemolymph.

The presence of the prophenoloxidase (proPO) system in the haemolymph of Rhodnius prolixus and Triatoma infestans and the role played by Trypanosoma rangeli in the in vitro activation of proPO were tested. Both R. prolixus and T. infestans whole blood preparations showed a very active ProPO system. The proPO cascade of the two insect species were differentially activated by microbial-derived extracts: laminarin was a better activator of T. infestans haemolymph than of R. prolixus blood, and lipopolysaccharides from Shigella flexneri or Pseudomonas aeroginosa caused significant proPO activation of T. infestans haemolymph but not of R. prolixus preparations. For the two insect species, neither T. rangeli from culture nor parasite lysates were able to trigger proPO activation. The presence of the parasite in R. prolixus haemolymph/laminarin assays, however, significantly reduced the level of proPO activation to that of spontaneous activating controls. The immobilization of T. rangeli in vitro in haemolymph preparations occurred in both insect species and was dependent on the proPO activation intensity. Our results suggest that the susceptibility of R. prolixus to T. rangeli haemocoel infection may be explained, at least in part, by the suppression of the insect immune defence system i.e., inhibition of proPO in the presence of this protozoan parasite.

Animals↗
PubMed1990

Direct binding and lectin-mediated binding of erythrocytes to haemocytes of the insect, Extatosoma tiaratum.

Treatment of Extatosoma haemocytes on a monolayer with fixed rabbit erythrocytes (RE) in the presence of Ca2+ results in ca. 11% plasmatocytes (PL) and 58% spreading granular (SG) cells rosetting. Pretreatment of haemocytes with lactose, D-galactose, or asialofetuin reduces rosetting, suggesting that the membrane-associated receptors involved may be lectins. The percentage of rosetting PL and SG cells is increased by a number of heterologous lectins (indicating the presence of D-galactose, mannose, and Glu Nac receptors on these haemocytes) and a D-galactose specific lectin isolated from the serum of the insect. The latter acts as a bridging molecule and requires Ca2+ to bind to specific receptors on both haemocytes and fixed RE.

Agglutination↗
PubMed1990

Activity and preliminary characterization of Branchiostoma lanceolatum agglutinin.

The physical and chemical properties of a hemagglutinin from whole-body homogenates of the cephalochordate, Branchiostoma lanceolatum, are described. The hemagglutinin is proteinaceous since it is precipitated by trichloroacetic acid and ammonium sulphate, and all activity is lost at 60 degrees C or by treating with proteases. Carbohydrate moieties are probably also present since activity is lost after incubation with sodium metaperiodate. Activity is stable over pH 6-10. The agglutinin does not require Ca++ or Mg++, and a reduced titer after treatment with 2-mercaptoethanol and urea suggests the presence of both disulphide bonds and noncovalent linkages. Haemagglutination inhibition experiments with 17 saccharides and glycoproteins failed to show clear-cut carbohydrate specificity, with only mucin and fetuin having any inhibitory effect, so that the binding may be complex. Finally, cross-adsorption experiments suggest that only one lectin is present. The function of this lectin, especially in an immunobiological context, remains speculative.

Animals↗
PubMed1989

Effect of exposure of Pieris brassicae larvae to 2,4,5-trichlorophenoxyacetic acid on the natural antibacterial activity of serum.

Larvae of the cabbage white butterfly, Pieris brassicae, were reared on a semisynthetic diet with or without 20 ppm of the herbicide 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and using three assays the sera were subsequently tested for natural antibacterial activity against Bacillus cereus, Escherichia coli K12, and Micrococcus luteus. These assays showed that exposure of larvae to 2,4,5-T lowered the antibacterial activity of the serum against E. coli and M. luteus compared with control animals. Spectrophotometric tests for the presence of a lysozyme-like principle in the serum also revealed similar trends with a significant loss of enzyme activity in 2,4,5-T-treated insects. Overall total serum protein levels of control and 2,4,5-T-treated insects were similar, suggesting a specific effect of the herbicide on certain serum components such as lysozyme. The possible mode of action of the herbicide on production of antibacterial factors is discussed.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMed1989

The biological significance of immunity.

The classical definition of immunity as the resistance of the body to disease, views the immune system in simple mechanistic terms. In this brief overview, possible reasons and consequences of the presence of an effective immune system across a range of invertebrate and vertebrate animals are considered. Topics discussed include the forces favouring the development of immunity, such as the acquisition of the colonial habit and terrestrial mode of life by many primitive animals, the constant threats of microbial and macrobial invasion, and the need to eliminate somatic mutations. The consequences of immunity in terms of the development of autoimmunity and hypersensitivity reactions, as well as the interaction of the immune system with the brain, the neuroendocrine organs and environmental factors, are also examined. Finally, the possibilities that the delicate balance maintained between many parasites and the immune systems of their hosts results in the spread of disease, and may also determine the maintenance of sexual reproduction and the choice of mate, are discussed.

Animals↗
PubMed1989

The evolution of blood cells: facts and enigmas.

Although the origin and evolution of vertebrate blood cells is conjectural and controversial, it is, nevertheless, of great interest, particularly to comparative immunologists seeking an underlying trend in the phylogenetic development of immunity. This review examines possible relationships between invertebrate and vertebrate blood cells and describes some of the original concepts put forward to account for blood cell evolution.

Animals↗
PubMed1989

Marine invertebrate mucus--agglutinating and antibacterial activity, with emphasis on Metridium senile.

The agglutinating and antibacterial activities of mucus from marine invertebrates are described. Mucus from eleven species was tested for the presence of haemagglutinins using a range of vertebrate erythrocytes. Mucus from Metridium senile, Ophiocomina nigra and Branchiostoma lanceolatum yielded positive results and was investigated further. The haemagglutinin of M. senile was purified, characterized and shown to be mannose-specific. Mucus from this animal was subsequently assayed for antibacterial activity using seawater isolates and verified marine and terrestrial cultures. Only two test organisms were inhibited, while the majority showed significantly enhanced growth. Possible functions of the mucus barrier in marine invertebrates are discussed.

Agglutinins↗
PubMed1987

Activity and preliminary characterisation of a hemagglutinin from the hemichordate Saccoglossus ruber.

Examination of whole body homogenates of the hemichordate, Saccoglossus ruber, has revealed the presence of a hemagglutinin with specificity for carbohydrates containing a D-galactose configuration. An analysis of the chemical and physical properties of the agglutinin showed it to be a non-dialysable, heat-labile protein, susceptible to the action of pronase E, but resistant to trypsin. Oxidation by periodate had no effect on agglutination and activity was not dependent upon the divalent cations, Ca2+ or Mg2+. The agglutinin was associated with the proboscis, collar and trunk regions, and was also detected in mucus covering the body surface. A possible role for the agglutinin in the defence system of the animal is discussed.

Animals↗
PubMed1983

The effects of pulsed magnetic fields on chick embryo cartilaginous skeletal rudiments in vitro.

The biological response of cultured 7-day embryonic chick tibiae to small alternating currents induced by pulsed magnetic fields (PMFs) was investigated. It was found that continuous exposure to PMFs over 7 days did not alter the overall DNA content of rudiments nor the incorporation of 3H-thymidine when compared with control tibiae. The overall collagen content of explants was slightly reduced by PMF exposure whilst the incorporation of 3H-proline was significantly suppressed. The synthesis of sulphated glycosaminoglycans was also measured in terms of 35SO4--incorporation, but PMF treatment failed to alter the levels of isotope incorporation. These results suggest that, whereas noncollagenous, and possibly collagenous, protein synthesis is affected by PMF exposure, glycosaminoglycan synthesis is not. Histological and electron microscopical observations substantiated this interpretation and revealed selective inhibition of matrix secretion in the periphery of the proliferative epiphyseal zones in experimental explants. High-power electron microscope examination of these zones showed that PMF-exposed matrix was sparsely invested with fibrous protein while elements of the stellate reticulum had formed large aggregates which were often clumped about the cell membrane. The results are discussed in terms of the possible role of naturally occurring potentials in the development and maintenance of connective tissue systems such as cartilage and bone.

Animals↗