Respiratory allergy to Blomia tropicalis: Immune response in four syngeneic mouse strains and assessment of a low allergen-dose,short-term experimental model

Background

The dust mite Blomia tropicalis is an important source of aeroallergens in tropical areas. Although a mouse model for B. tropicalis extract (BtE)-induced asthma has been described, no study comparing different mouse strains in this asthma model has been reported. The relevance and reproducibility of experimental animal models of allergy depends on the genetic background of the animal, the molecular composition of the allergen and the experimental protocol.

Objectives

This work had two objectives. The first was to study the anti-B. tropicalis allergic responses in different mouse strains using a short-term model of respiratory allergy to BtE. This study included the comparison of the allergic responses elicited by BtE with those elicited by ovalbumin in mice of the strain that responded better to BtE sensitization. The second objective was to investigate whether the best responder mouse strain could be used in an experimental model of allergy employing relatively low BtE doses.

Methods

Groups of mice of four different syngeneic strains were sensitized subcutaneously with 100 μg of BtE on days 0 and 7 and challenged four times intranasally, at days 8, 10, 12, and 14, with 10 μg of BtE. A/J mice, that were the best responders to BtE sensitization, were used to compare the B. tropicalis-specific asthma experimental model with the conventional experimental model of ovalbumin (OVA)-specific asthma. A/J mice were also sensitized with a lower dose of BtE.

Results

Mice of all strains had lung inflammatory-cell infiltration and increased levels of anti-BtE IgE antibodies, but these responses were significantly more intense in A/J mice than in CBA/J, BALB/c or C57BL/6J mice. Immunization of A/J mice with BtE induced a more intense airway eosinophil influx, higher levels of total IgE, similar airway hyperreactivity to methacholine but less intense mucous production, and lower levels of specific IgE, IgG1 and IgG2 antibodies than sensitization with OVA. Finally, immunization with a relatively low BtE dose (10 μg per subcutaneous injection per mouse) was able to sensitize A/J mice, which were the best responders to high-dose BtE immunization, for the development of allergy-associated immune and lung inflammatory responses.

Conclusions

The described short-term model of BtE-induced allergic lung disease is reproducible in different syngeneic mouse strains, and mice of the A/J strain was the most responsive to it. In addition, it was shown that OVA and BtE induce quantitatively different immune responses in A/J mice and that the experimental model can be set up with low amounts of BtE.     

Tryptophan solvent exposure in folded and unfolded states of an SH3 domain by 19F and 1H NMR

The isolated N-terminal SH3 domain of the Drosophila signal transduction protein Drk (drkN SH3) is a useful model for the study of residual structure and fluctuating structure in disordered proteins since it exists in slow exchange between a folded (Fexch) and compact unfolded (Uexch) state in roughly equal proportions under nondenaturing conditions. The single tryptophan residue, Trp36, is believed to play a key role in forming a non-native hydrophobic cluster in the Uexch state, with a number of long-range nuclear Overhauser contacts (NOEs) observed primarily to the indole proton. Substitution of Trp36 for 5-fluoro-Trp36 resulted in a substantial shift in the equilibrium to favor the Fexch state. A variety of 19F NMR measurements were performed to investigate the degree of solvent exposure and hydrophobicity associated with the 5-fluoro position in both the Fexch and Uexch states. Ambient T1 measurements and H2O/D2O solvent isotope effects indicated extensive protein contacts to the 5-fluoro position in the Fexch state and greater solvent exposure in the Uexch state. This was corroborated by the measurements of paramagnetic effects (chemical shift perturbations and T1 relaxation enhancement) from dissolved oxygen at a partial pressure of 20 atm. In contrast, paramagnetic effects from dissolved oxygen revealed less solvent exposure to the indole proton of Trp36 in the Uexch state than that observed for the Fexch state, consistent with the model in which Trp36 indole belongs to a non-native cluster. Thus, although the Uexch state may be described as a dynamically interconverting ensemble of conformers, there appears to be significant asymmetry in the environment of the indole group and the six-membered ring or backbone of Trp36. This implied lack of averaging of a side chain position is in contrast to the general view of fluctuating side chains within disordered states.     

Comparison of the efficacies of disinfectants to control microbial contamination in dental unit water systems in general dental practices across the European Union

Water delivered by dental unit water systems (DUWS) in general dental practices can harbor high numbers of bacteria, including opportunistic pathogens. Biofilms on tubing within DUWS provide a reservoir for microorganisms and should be controlled. This study compared disinfection products for their ability to meet the American Dental Association's guideline of <200 CFU x ml(-1) for DUWS water. Alpron, BioBlue, Dentosept, Oxygenal, Sanosil, Sterilex Ultra, and Ster4Spray were tested in DUWS (n = 134) in Denmark, Germany, Greece, Ireland, The Netherlands, Spain, and the United Kingdom. Weekly water samples were tested for total viable counts (TVCs) on yeast extract agar, and, where possible, the effects of products on established biofilm (TVCs) were measured. A 4- to 5-week baseline measurement period was followed by 6 to 8 weeks of disinfection (intermittent or continuous product application). DUWS water TVCs before disinfection ranged from 0 to 5.41 log CFU x ml(-1). Disinfectants achieved reductions in the median water TVC ranging from 0.69 (Ster4Spray) to 3.11 (Dentosept) log CFU x ml(-1), although occasional high values (up to 4.88 log CFU x ml(-1)) occurred with all products. Before treatment, 64% of all baseline samples exceeded American Dental Association guidelines, compared to only 17% following commencement of treatment; where tested, biofilm TVCs were reduced to below detectable levels. The antimicrobial efficacies of products varied (e.g., 91% of water samples from DUWS treated with Dentosept or Oxygenal met American Dental Association guidelines, compared to 60% of those treated with Ster4Spray). Overall, the continuously applied products performed better than those applied intermittently. The most effective products were Dentosept and Oxygenal, although Dentosept gave the most consistent and sustained antimicrobial effect over time.     

Metabolic energy utilization during development of Antarctic naked dragonfish (Gymnodraco acuticeps)

We have capitalised on the availability of eggs and adults of the naked dragonfish Gymnodraco acuticeps (Sub-order Notothenioidei, F. Bathydraconidae) near McMurdo Station, Antarctica to examine metabolic energy utilization at different stages of its life cycle. Average egg respiration rates were found to increase from 2.17±1.02 nmol O₂ h⁻¹ ind⁻¹ at about 17 h post-fertilization (hpf) to 5.72±0.56 nmol h⁻¹ ind⁻¹ at about 24 hpf, during which time the eggs underwent first cleavage. The respiration rates of embryos from 2–20 days post-fertilization (dpf) averaged 4.11±1.47 nmol O₂ h⁻¹ ind⁻¹. About 10 months post-fertilization, oxygen consumption rates of 27.14±3.92 nmol O₂ h⁻¹ ind⁻¹ were recorded immediately prior to hatching, with a peak of 112.41±31.38 nmol O₂ h⁻¹ ind⁻¹ at the time of hatch. Larvae aged 46–63 days post-hatch had an average respiration rate of 64.4±15.11 nmol O₂ h⁻¹ ind⁻¹. Mass-specific respiration rates of hatched larvae (approximately 1–2 months old) were calculated using dry weights (DW) and averaged 16.1±3.4 nmol O₂ h⁻¹ mg⁻¹ DW. Adult dragonfish respiration rates (corrected for a 100 g fish and using a 0.8 scaling exponent) averaged 0.91±0.36 mmol O₂ kg⁻¹ h⁻¹ after a 48 h acclimatization period, which is not indicative of significant metabolic cold adaptation. The energy contents of dragonfish eggs and larvae were also measured by microbomb calorimetry and used, along with the respiration data, in an initial approach to estimate an energy budget. In order to balance the budget, the bulk of the available post-gastrulation respiratory energy (during 213 days of embryonic incubation) must be consumed at a relatively low average rate (7.1 nmol O₂ h⁻¹ ind⁻¹), which supports the possibility that advanced dragonfish embryos overwinter in a relatively quiescent metabolic state while awaiting a suitable stimulus (such as the return of the sun) to initiate hatching.     

Lipid-protein interactions with the Na,K-ATPase

Studies of lipid interactions with membranous Na,K-ATPase by using electron spin resonance spectroscopy in conjunction with spin-labelled lipids are reviewed. The lipid stoichiometry, selectivity and exchange dynamics at the lipid-protein interface can be determined, in addition to information on the configuration and rotational dynamics of the protein-associated lipid chains. These parameters, particularly the stoichiometry and selectivity, are related directly to the intramembranous structure of the Na,K-ATPase, and can be used to check the integrity of extensively trypsinised preparations.     

Lipid conformation in crystalline bilayers and in crystals of transmembrane proteins

Dihedral torsion angles evaluated for the phospholipid molecules resolved in the X-ray structures of transmembrane proteins in crystals are compared with those of phospholipids in bilayer crystals, and with the phospholipid conformations in fluid membranes. Conformations of the lipid glycerol backbone in protein crystals are not restricted to the gauche C1-C2 rotamers found invariably in phospholipid bilayer crystals. Lipid headgroup conformations in protein crystals also do not conform solely to the bent-down conformation, with gauche-gauche configuration of the phospho-diester, that is characteristic of phospholipid bilayer membranes. This suggests that the lipids that are resolved in crystals of membrane proteins are not representative of the entire lipid-protein interface. Much of the chain configurational disorder of the membrane-bound lipids in crystals arises from energetically disallowed skew conformations. This indicates a configurational heterogeneity in the lipids at a single binding site: eclipsed conformations occur also in some glycerol backbone torsion angles and C-C torsion angles in the lipid headgroups. Stereochemical violations in the protein-bound lipids are evidenced by one-third of the ester carboxyl groups in non-planar configurations, and certain of the carboxyls in the cis configuration. Some of the lipid structures in protein crystals have the incorrect enantiomeric configuration of the glycerol backbone, and many of the branched methyl groups in structures of the phytanyl chains associated with bacteriorhodopsin crystals are in the incorrect S-configuration.     

Azorhizobium doebereinerae sp. Nov. Microsymbiont of Sesbania virgata (Caz.) Pers

Thirty-four rhizobium strains were isolated from root nodules of the fast-growing woody native species Sesbania virgata in different regions of southeast Brazil (Minas Gerais and Rio de Janeiro States). These isolates had cultural characteristics on YMA quite similar to Azorhizobium caulinodans (alkalinization, scant extracellular polysaccharide production, fast or intermediate growth rate). They exhibited a high similarity of phenotypic and genotypic characteristics among themselves and to a lesser extent with A. caulinodans. DNA:DNA hybridization and 16SrRNA sequences support their inclusion in the genus Azorhizobium, but not in the species A. caulinodans. The name A. doebereinerae is proposed, with isolate UFLA1-100 (=BR5401, =LMG9993=SEMIA 6401) as the type strain.     

Case report: Metaplastic carcinoma presenting as a breast abscess

Metaplastic breast carcinoma (MBC) is a rare neoplasm containing a mixture of epithelial and mesenchymal elements. The epithelial component is usually ductal carcinoma but may include other variants of breast carcinomas including squamous carcinoma and osteogenic sarcoma. There is a relative paucity of data regarding such tumours. Metaplastic carcinoma carries a prognosis not dissimilar to that of comparable ductal carcinoma. This is the case of a 57 year old patient with MBC presenting with a breast abscess. A thorough literature search has not revealed any previous reports of MBC presenting as a breast abscess.     

A Metal Detector Study to Locate Inactive Small Arms Range Impact Areas

Historical records suggested the existence of .45 caliber Thompson sub-machine gun ranges near the Fort Lewis Evergreen Range; however, the precise location of the bullet impact area was not known. The site covered 20 acres, too large to cost-effectively characterize using convention grid sampling techniques. As an alternative, a study was conducted using a hand-held metal detector to locate the bullet impact areas.

The metal detector chosen, a Garrett Graphic Target Imaging? (GTI) 2500, has an LCD screen, which provides the user information on the potential size and depth of the object causing the signal. A two-man team performed the study and bullet detection was confirmed by excavation.

The metal detector proved effective at finding .45 caliber bullets. Three impact areas were successfully identified and marked using a global positioning system (GPS). The metal detecting strategy saved substantial sampling time and over $35,000.00 by decreasing the sampling area by 93 percent.     

Interindividual variation of malate dehydrogenase activity in the oyster Crassostrea virginica

Intertidal environments are dynamic, stressful niches and variation in physiological parameters may determine distribution and survival of individuals in a population. We demonstrated that mitochondria of the oyster Crassostrea virginica oxidize malate more readily than other Krebs cycle intermediates and investigated the level of interindividual variability in oyster malate dehydrogenase (MDH) activity and total protein content in muscle tissues. Both MDH activity and total protein evidenced a high level of interindividual variation in heart and adductor among a sample of more than 50 oysters. Normalization to total DNA failed to explain the variation in either MDH activity or protein content of phasic adductor and explained less than 40% of the variation in heart. This range of MDH titers defines a continuum of biochemical phenotypes important for understanding the relative selection forces operative on metabolic pathways within the muscles of the oyster.