Recognition of Neisseria meningitidis by the Long Pentraxin PTX3 and Its Role as an Endogenous Adjuvant

Long pentraxin 3 (PTX3) is a non-redundant component of the humoral arm of innate immunity. The present study was designed to investigate the interaction of PTX3 with Neisseria meningitidis. PTX3 bound acapsular meningococcus, Neisseria-derived outer membrane vesicles (OMV) and 3 selected meningococcal antigens (GNA0667, GNA1030 and GNA2091). PTX3-recognized microbial moieties are conserved structures which fulfil essential microbial functions. Ptx3-deficient mice had a lower antibody response in vaccination protocols with OMV and co-administration of PTX3 increased the antibody response, particularly in Ptx3-deficient mice. Administration of PTX3 reduced the bacterial load in infant rats challenged with Neisseria meningitidis. These results suggest that PTX3 recognizes a set of conserved structures from Neisseria meningitidis and acts as an amplifier/endogenous adjuvant of responses to this bacterium.     

Effects of acute hyperoxic exposure on solute fluxes across the blood-gas barrier in rat lungs

Zheng, Lu P., Rui Sheng Du, and Barbara E. Goodman.Effects of acute hyperoxic exposure on solute fluxes across the blood-gas barrier in rat lungs. J. Appl.Physiol. 82(1): 240-247, 1997.We investigatedeffects of acute hyperoxia on solute transport from air space tovascular space in isolated rat lungs. Air spaces were filled withKrebs-Ringer bicarbonate solution containing fluoresceinisothiocyanate-labeled dextran (FD-20; mol wt 20,000) and either²²Na⁺and [¹⁴C]sucrose, orD-[¹⁴C]glucoseandL-[³H]glucose.Apparent permeability-surface area products for tracers over time (upto 120 min) were calculated for isolated perfused lungs from controlrats (room air) and rats exposed to >95%O₂ for 48 or 60 h immediatelypostexposure. After O₂ exposures,mean fluxes for[¹⁴C]sucrose and FD-20were significantly higher than in room-air control lungs. However,amiloride-sensitive Na⁺ and activeD-glucose fluxes were unchangedafter hyperoxic exposure. Therefore, it is unlikely that decreases innet solute transport in this lung-injury model contributed to pulmonaryedema resulting from O₂ toxicity.Increased net solute transport shown to help resolve pulmonary edemaafter acute hyperoxic exposure must therefore begin during the recoveryperiod. In summary, our data show increases in passive solute fluxesbut no changes in active solute fluxes immediately after acutehyperoxic lung injury.

     

Spatial and temporal impacts of a diesel fuel spill on stream invertebrates

1. We assessed the effects of a 26 500 L diesel fuel spill on the macroinvertebrate fauna of a small trout stream in central New York, U.S.A. To determine the spatial extent of the spill we sampled three locations (0.7, 5.0 and 11.8 km downstream of the spill), each containing a reference site (unaffected tributary) and an impact site (downstream of spill). Sampling was repeated four times over a 15‐month period to assess temporal recovery. 2. Immediately after the spill, invertebrate density at all three locations below the spill was significantly lower than reference density. Three months after the spill, density up to 5 km below the spill was still far lower (<100 individuals per sample) than reference density (800–1200 individuals per sample). A year after the spill, density was similar between reference and impact sites, suggesting that invertebrates had recovered numerically. 3. Taxonomic richness up to 5.0 km below the spill was less than half the reference taxonomic richness and this difference persisted for at least 3 months. Some significant differences between reference and impact sites were observed after 15 months, but these differences could not be attributed to the oil spill. 4. For at least 3 months following the spill, the site immediately downstream of the spill was dominated by Optioservus, a petrochemical‐tolerant riffle beetle. Twelve to 15 months after the spill, both the reference and impact sites near the spill were dominated numerically by the mayfly Ephemerella, but the degree of dominance was twice as large at the impact site. 5. We concluded that the diesel fuel spill significantly reduced the density of invertebrates (by 90%) and taxonomic richness (by 50%) at least 5.0 km downstream, but density recovered within a year. Throughout the study, however, the fauna immediately below the spill was species poor and significantly over‐represented by a single dominant taxon, suggesting that 15 months was not sufficient for full community recovery from the oil spill.     

Occurrence of isopropylthio compounds in the aquatic ecosystem (Lake Neusiedl, Austria) as a chemical marker for Microcystis flos-aquae

Abstract Volatile organic sulfur compounds occuring during a bloom of different species of Microcystis in Lake Neusiedl, Austria, were analyzed by gas chromatography and mass spectrometry. In open water diisopropyl disulfide and diisopropyl tri-sulfide were the only sulphur compounds to be found. It was shown that Microcystis flos-aquae was the causative agent for the generation of these sulphur compounds, since high concentrations of these substances were found both in the floating scum of cyanobacteria taken from open lake and in axenic cultures of five isolated strains of M. flos-aquae . Strains isolated from colonies of Microcystis aeruginosa were not able to synthesize isopropylthio compounds. Alternatively, methylthio compounds were released. The rather unusual formation of the isopropylthio group can be used as a chemical marker to differentiate between M. flos-aquae and M. aeruginosa as two separate species which hitherto have been regarded as formae. In a canal passing through the reed belt of Lake Neusiedl where Microcystis was missing, these compounds were not detected. Different sulfur compounds (dimethyl disulfide, dimethyl trisulfide, dibutyl sulfide and bis(methylthio) methane) which in part have not yet been reported for freshwater ecosystems occurred at this site. Their origin, however, remains obscure.     

Changes in biophysical parameters of plasma membranes influence cisplatin resistance of sensitive and resistant epidermal carcinoma cells

The mechanism of resistance of cancer cells to the anticancer drug cisplatin is not fully understood. Using cisplatin-sensitive KB-3-1 and -resistant KCP-20 cells, we found that the resistant cells have higher membrane potential, as determined by membrane potential sensing oxonol dye. Electron spin resonance and fluorescence polarization studies revealed that the resistant cells have more "fluid" plasma membranes than the sensitive cells. Because of this observed difference in membrane "fluidity," we attempted modification of the plasma membrane fluidity by the incorporation of heptadecanoic acid into KB-3-1 and KCP-20 cell membranes. We found that such treatment resulted in increased heptadecanoic acid content and increased fluidity in the plasma membranes of both cell types, and also resulted in increased cisplatin resistance in the KCP-20 cells. This finding is in accord with our results, which showed that the cisplatin-resistant KCP-20 cells have more fluid membranes than the cisplatin-sensitive KB-3-1 cells. It remains to be determined whether the observed differences in biophysical status and/or fatty acid composition alone, or the secondary effect of these differences on the structure or function of some transmembrane protein(s), is the reason for increased cisplatin resistance.     

Synthesis and antibacterial activity of C6-carbazate ketolides

A novel series of ketolides containing heteroaryl groups that are linked to the erythronolide ring via a C6-carbazate functionality has been successfully synthesized. Careful modulation of the heteroaryl groups, the length and degree of saturation of the C6-carbazate linker, and the substituents present on each of the carbazate nitrogens led to compounds with potent activity against key bacterial respiratory pathogens. The best analogs of this series had in vitro and in vivo (sc dosing) profiles that were comparable to telithromycin.     

Insulin binding in the hypothalamus of lean and genetically obese Zucker rats

Recent reports have suggested that the obesity and hyperphagia of the genetically obese Zucker rat may be related to defective insulin action or binding in the hypothalamus. We used quantitative autoradiography to determine if insulin binding is altered in specific hypothalamic nuclei associated with food intake. Insulin binding was measured in the arcuate (ARC), dorsomedial (DMN), and ventromedial (VMN) hypothalamic nuclei of 3–4-month-old lean (Fa/Fa) and genetically obese (fa/fa) Zucker rats. A consistently reproducible 15% increase in the total specific binding of 0.1 nM [¹²⁵I]-insulin was found in the ARC of the obese genotype. A slight increase in insulin binding in the DMN was also found. No difference in specific insulin binding was found between genotypes in the VMN. Nonlinear least squares analysis of competitive binding studies showed that the K_d of the ARC insulin binding site was 33% higher in the lean rats than in the obese rats, indicating an increased affinity for insulin. No difference in site number (B_max) was found in the ARC, DMN or VMN, and no evidence was found for reduced insulin binding in the hypothalamus of the obese (fa/fa) genotype. The results suggest that hyperphagia and obesity of the obese (fa/fa) Zucker rat genotype may be associated with increased insulin binding in the arcuate nucleus.     

Thiol-directed immobilization of recombinant IgG-binding receptors

A genetic engineering approach is described for directed immobilization of IgG-binding receptors to a thiol-containing matrix using a single cysteine residue. The cysteine residue is introduced into the C-terminal part of receptors based on staphylococcal protein A. Receptors containing one, two or five IgG-binding domains were produced in Escherichia coli and subsequently immobilized on thiopropyl-Sepharose. A high amount, 5 mumol/ml gel (45 mg/ml), of recombinant receptor could be rapidly immobilized to the solid support and both the gel and the immobilized receptor could be regenerated by reduction and oxidation reactions. The gel was used for affinity purification of human IgG and analysis of IgG-binding capacity at different amounts of immobilized recombinant protein show the same maximal IgG-binding capacity (20-25 mg/ml) for all three immobilized receptors. However, at low substitution grade of receptors, the immobilized receptor molecules were shown to bind one (Z-Cys) and two (ZZ-Cys) IgG molecules, respectively. These results demonstrate that the immobilized protein molecules are in a functionally active form and that a two-domain receptor can bind two molecules of IgG without steric hindrance. Interestingly, the five-domain receptor (ZV-Cys), with a structure similar to native protein A, can only bind approximately two IgG molecules, despite the five-domain structure of the molecule.