Conjugation of meningococcal lipooligosaccharides through their lipid A terminus conserves their inner epitopes and results in conjugate vaccines having improved immunological properties

The importance of conserved inner saccharide epitopes to the immune performance of meningococcal lipooligosaccharide-protein conjugate vaccines was demonstrated in the following experiments. Two different oligosaccharides were obtained by chemical degradations of the same L7 lipooligosaccharide, and both were linked terminally to tetanus toxoid. One was a truncated oligosaccharide in which the inner epitopes were incomplete and was obtained by mild acid hydrolysis of the L7 lipooligosaccharide. This oligosaccharide was conjugated by direct reductive amination through its newly exposed terminal Kdo residue. The second, a full-length oligosaccharide, was obtained by O-deacylation of the L7 lipooligosaccharide, with subsequent removal of phosphate substituents from its lipid A moiety using alkaline phosphatase. This permitted the full-length oligosaccharide to be conjugated directly to tetanus toxoid by reductive amination through its newly exposed terminal 2-N-acyl-2-deoxy-D-glucopyranose residue. Comparison of the immune performance of the two conjugates in mice revealed, that while both were able to induce significant levels of L7-lipooligosaccharide-specific IgG antibody, the conjugate made with the full-length saccharide was able to induce antibodies with increased bactericidal activity against homologous meningococci.     

Chemically methylated and reduced pectins: preparation,characterisation by 1H NMR spectroscopy,enzymatic degradation,and gelling properties

The gelling properties of pectins are known to be closely related to the degree of methylation (DM) and the distribution of the ester groups. In order to investigate this dependency, a natural citrus pectin (DM 64%) has been methylated to pectins with higher DM or saponified to achieve pectins with lower DM. A simple method for determination of DM by 1H NMR spectroscopy is presented. New modified pectins have been prepared by treatment of pectins having different DM with NaBH(4) to reduce selectively the methyl esters to primary alcohols in the presence of free acids. The degree of reduction (DR) and the DM of the remaining carboxylic acids could likewise be determined by 1H NMR spectroscopy. The new reduced pectins are recognized by the pectin degrading enzymes polygalacturonase PGI and PGII as well as by pectin lyase, all from Aspergillus niger, but the enzymes exhibit lower specific activities as compared with unmodified pectin. The new reduced pectins exhibit high gelling properties.     

Rheological properties of sulfoacetate derivatives of cellulose

Water-soluble cellulose acetate sulfate derivatives (CAS) have been prepared through chemical reaction involving sulfuric acid as a catalyst. These CAS have been obtained from cellulosic materials of different origins (pure cellulose, wheat bran, maize bran) and their rheological behavior in salt-free aqueous solution has been estimated in dilute and semi-dilute regime using dynamic viscoelastic and viscosity measurements. Influence of concentration, temperature of solubilization and temperature of measurement has been investigated. Weak gel-like properties were exhibited at elevated concentration (typically above 7-8 g/L). These systems also exhibited thixotropic properties: the structure was partly broken down upon shearing and recovered at rest. They also displayed thermoreversibility with large hysteresis, the melting temperature being approximately 15 degrees C higher than the temperature at which gelation took place. These overall observations clearly indicate that these distinctive properties arise from intermolecular association of the macromolecular chains of the cellulose derivative.     

Immunological comparison of native and recombinant egg allergen, ovalbumin, expressed in Escherichia coli

Chicken ovalbumin is one of the major egg white allergens which causes IgE-mediated food hypersensitivity. A gene encoding for chicken ovalbumin (Gad dI) was isolated from chicken oviduct by PCR amplification and was cloned under the control of T5 promoter fused with a six-histidine tag at the N-terminal end. Escherichia coli harbouring this construct expressed high quantities of the recombinant protein in the form of soluble fraction. The protein was purified using affinity chromatography on a Ni(2+)-nitrilotriacetic acid agarose column and was further purified to homogeneity by ion exchange chromatography. Homogeneity was confirmed through SDS-PAGE, Western blot and secondary conformation analysis. The reactivity of the recombinant and native protein was tested against six egg allergic human patient's sera and the IgE and IgG binding activity was tested using both Western blot and ELISA. When compared to native ovalbumin, the recombinant protein had similar binding activity in immunoblotting, but slightly increased activity by ELISA. Circular dichroism revealed that the recombinant protein had a slightly less compact structure than the native form. Both antigens exhibited a similar immunogenicity in mice.     

Volatile constituents of Aristolochia argentina

The essential oils from leaves, aerial stems and underground organs of Aristolochia argentina Gris., a medicinal plant popularly known as "charrúa", were obtained by hydrodistillation and analyzed by GC and GC/MS. Forty-three components were identified in the oils. All parts of the plant afforded volatile oils characterized by high levels of argentilactone (57-89%) and the presence of undecatriene isomers (0.3-4.0%), these latter compounds providing the essential oils and extracts with an intense particular odor. Terpenes account for the remaining portion of the essential oils (5-29%). Bicyclogermacrene predominates in the aerial parts of the plant, whereas ishwarane is the main terpene of the subterranean organs. Argentilactone, a suspected carcinogenic compound, was also identified in a medicinal commercial tincture of A. argentina.     

Structural characterization of diC14-amidine, a pH-sensitive cationic lipid used for transfection

The structure of N-t-butyl-N'-tetradecyl-3-tetradecylaminopropionamidine (diC(14)-amidine) cationic vesicles, used for transfection, was investigated at different pH values and ionic strengths, through the analysis of the electron spin resonance (ESR) spectra of spin labels. Phospholipid derivatives, spin labeled at the 5th and 16th C-atoms along the hydrocarbon chain, incorporated in diC(14)-amidine bilayers, show that the bilayer structure is highly sensitive to the pH value of the medium, due to the two titratable groups present in the amphiphile. Compared with samples at higher pH values, the double charged diC(14)-amidine at pH 3 presents a rather non-organized bilayer gel phase, and a much lower gel-fluid temperature transition, in accord with a strong headgroup electrostatic repulsion. In addition, the structure was found to be highly dependent on the ionic strength of the medium. However, pH 3 diC(14)-amidine bilayer, in the fluid phase, was found to be slightly more closely packed than those at pH 7.4 or 9.0, which are less charged. Parallel to that, the larger isotropic hyperfine splitting measured for nitroxides in the center of the pH 3 diC(14)-amidine bilayer suggests a higher membrane polarity for the highly charged low pH sample.     

Cardiac specification of embryonic stem cells

Over the past decade, cell transplantation has been recognized as a mean of repairing infarcted myocardium. Both adult stem cells and differentiated cells have yielded encouraging results with regard to engraftment into postinfarction scars. However, these cells now feature serious restrictions. Asan alternative, embryonic stem (ES) cells are particularly attractive, because of their plasticity and the subsequent possibility to drive them towards a cardiomyogenic phenotype after exposure to appropriate growth factors. An additional theoretical advantage of ES cells is their expected immune privilege. In this article, we summarize the findings obtained in cell therapy using ES cells and discuss the molecular mechanisms of cardiac specification of the cells.     

Erythrinaline alkaloids from the flowers and pods of Erythrina lysistemon and their DPPH radical scavenging properties

Fourteen different erythrinaline alkaloids have been isolated from the flowers and pods of Erythrina lysistemon with four being reported for the first time in nature and five for the first time in this species and the rest having been re-isolated. The new compounds are (+)-11beta-hydroxyerysotramidine (1), (+)-11beta-methoxyerysotramidine (2), (+)-11beta-hydroxyerysotrine N-oxide (4) and (+)-11beta-hydroxyerysotrine (8). (+)-11alpha-Hydroxyerysotrine N-oxide (3), earlier misidentified as erythrartine N-oxide (beta-hydroxyerysotrine N-oxide 4), was also re-isolated along with four other alkaloids. Correct identification of compounds 4 and 8 was aided by the fact that the two sets of C-11 epimers 3, 4 and 8, 9 were both isolated in this study thus making it easier to identify and assign the individual epimers. (+)-Erythristemine (14) was found distributed in most of the plant parts investigated. Preliminary work on the crude chloroform/methanol (1:1) showed moderate toxicity to brine shrimp (LC50 23 ppm) and moderate (IC50 86 microg/ml) radical scavenging properties against stable 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical. The DPPH radical scavenging properties of the isolated compounds were assessed using TLC autographic and spectrophotometric assays whereupon only compounds 11 (1 microg; 90 microg/ml) and 12 (0.1 microg; 160 microg/ml) showed any notable activity. It appears the two compounds are slow reacting and do not reach steady state conditions within the standard half an hour time frame but only seemed to have reached steady state conditions after 4 h.     

Food procurement and tooth use in two sympatric lemur species

This study of two lemur species (Lemur catta and Propithecus v. verreauxi) in Madagascar combines observations of food procurement and initial food placement in the mouth with comparisons to food toughness and external properties. Food toughness was hypothesized to play a decisive role in determining food placement during ingestion. It was found that tougher foods are generally ingested on the postcanines for all foods eaten. However, when leaves and fruits are analyzed separately, food size and shape, represented here by mass and food type, are more reliable predictors of initial food placement. Larger leaves and bulkier fruits and stalks are ingested posteriorly. Leaf toughness is not related to leaf size, though the toughness and size of the most commonly eaten fruits are correlated. Furthermore, ingestive food toughness, which is the maximum toughness, and "average" food toughness may make different mechanical demands on the masticatory apparatus that have consequences for jaw morphology.