Significance of alveolar nitric oxide concentration in the airway of patients with organizing pneumonia after allogeneic hematopoietic stem cell transplantation.

Organizing pneumonia (OP) is a complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) and a manifestation of peripheral airway/alveolar inflammation. Recently, alveolar nitric oxide concentration (Calv) has been revealed as a noninvasive marker of peripheral airway inflammation; however, whether Calv levels are associated with OP and peripheral airway in patients after allo-HSCT remains unclear. Herein, we evaluated whether Calv levels could reflect the presence of OP and structural airway changes in patients after allo-HSCT. We measured the eNO levels of 38 patients (6 with OP and 32 without OP) who underwent allo-HSCT. Three-dimensional computed tomography (CT) analysis of the airway was performed in 19 patients. We found that in patients with OP, Calv levels were significantly higher than in those without OP (10.6 vs. 5.5 ppb, p < 0.01). Receiver-operating characteristic analyses revealed a Calv cut-off value for OP detection of 10.2 ppb. No significant differences in the patient characteristics, except for the presence of OP (p < 0.01), were noted between the two groups stratified by the Calv cut-off value. Three-dimensional CT images of the airway revealed gradually increasing positive correlations between Calv levels and airway wall area of the third-, fourth-, and fifth-generation bronchi (r = 0.20, 0.31, 0.38; p = 0.42, 0.19, 0.038, respectively), indicating that Calv levels are strongly correlated with the wall thickness of the distal bronchi. Our results suggest that the Calv level may be a useful noninvasive detectable marker for OP after an allo-HSCT.

Toxic metabolite profiling of Inocybe virosa.

Wild mushroom foraging involves a high risk of unintentional consumption of poisonous mushrooms which is a serious health concern. This problem arises due to the close morphological resemblances of toxic mushrooms with edible ones. The genus Inocybe comprises both edible and poisonous species and it is therefore important to differentiate them. Knowledge about their chemical nature will unambiguously determine their edibility and aid in an effective treatment in case of poisonings. In the present study, the presence of volatile toxic metabolites was verified in Inocybe virosa by gas chromatography. Methyl palmitate, phenol, 3,5-bis (1,1-dimethyl ethyl) and phytol were the identified compounds with suspected toxicity. The presence of the toxin muscarine was confirmed by liquid chromatography. The in vitro study showed that there was negligible effect of the digestion process on muscarine content or its toxicity. Therefore, the role of muscarine in the toxicity of Inocybe virosa was studied using a bioassay wherein metameters such as hypersalivation, immobility, excessive defecation, heart rate and micturition were measured. Administration of muscarine resulted in an earlier onset of symptoms and the extract showed a slightly stronger muscarinic effect in comparison to an equivalent dose of muscarine estimated in it. Further, the biological fate of muscarine was studied by pharmacokinetics and gamma scintigraphy in New Zealand white rabbits. Significant amount of the toxin was rapidly and effectively concentrated in the thorax and head region. This study closely explains the early muscarinic response such as miosis and salivation in mice. By the end of 24 h, a relatively major proportion of muscarine administered was accumulated in the liver which stands as an explanation to the hepatotoxicity of Inocybe virosa. This is one of the rare studies that has attempted to understand the toxic potential of muscarine which has previously been explored extensively for its pharmaceutical applications.

Improvement of mosquito identification by MALDI-TOF MS biotyping using protein signatures from two body parts.

BACKGROUND: Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry technology (MALDI-TOF MS) is an innovative tool that has been shown to be effective for the identification of numerous arthropod groups including mosquitoes. A critical step in the implementation of MALDI-TOF MS identification is the creation of spectra databases (DB) for the species of interest. Mosquito legs were the body part most frequently used to create identification DB. However, legs are one of the most fragile mosquito compartments, which can put identification at risk. Here, we assessed whether mosquito thoraxes could also be used as a relevant body part for mosquito species identification using a MALDI-TOF MS biotyping strategy; we propose a double DB query strategy to reinforce identification success. METHODS: Thoraxes and legs from 91 mosquito specimens belonging to seven mosquito species collected in six localities from Guadeloupe, and two laboratory strains, Aedes aegypti BORA and Aedes albopictus Marseille, were dissected and analyzed by MALDI-TOF MS. Molecular identification using cox1 gene sequencing was also conducted on representative specimens to confirm their identification. RESULTS: MS profiles obtained with both thoraxes and legs were highly compartment-specific, species-specific and species-reproducible, allowing high identification scores (log-score values, LSVs) when queried against the in-house MS reference spectra DB (thorax LSVs range: 2.260-2.783, leg LSVs range: 2.132-2.753). CONCLUSIONS: Both thoraxes and legs could be used for a double DB query in order to reinforce the success and accuracy of MALDI-TOF MS identification.

How to Wire the Diaphragm: Wholemount Staining Methods to Analyze Mammalian Respiratory Innervation.

Direct or indirect impairment of breathing in humans by diseases or environmental factors can either cause long-term disability and pain, or can ultimately result in death. Automatic respiratory centers in the brainstem control the highly structured process of breathing and signal to a specialized group of motor neurons in the cervical spinal cord that constitute the phrenic nerves. In mammals, the thoracic diaphragm separates the thorax from the abdomen and adopts the function of the primary respiratory musculature. Faithful innervation by the phrenic nerves is a prerequisite for correct functionality of this highly specialized musculature and thus, ultimately, the viability of the entire organism.To analyze the effects of diseases and genetic defects responsible for deleterious or lethal respiratory phenotypes, accurate imaging of respiratory innervation during embryonic development, e.g., in genetically modified mouse models enables the characterization of specific marker genes and pathways that underlie appropriate wiring of the diaphragm. Among the different available immunostaining techniques, wholemount staining methods provide the advantage of clear and faithful three-dimensional information about the location of the antigens of interest. In comparison to routine histological techniques, however, the researcher has to deal with technical challenges, such as antibody penetration, the stability and availability of the antigen, and clearing of the relevant tissue, and the need to be equipped with state-of-the-art microscope equipment.In this methodological chapter, we explain and share our expertise concerning wholemount processing of mouse embryos and thoracic diaphragms for the analysis of mammalian respiratory innervation.

Localization of Defensive Chemicals in Two Congeneric Butterflies (Euphydryas, Nymphalidae).

Many insect species sequester compounds acquired from their host plants for defense against natural enemies. The distribution of these compounds is likely to affect both their efficacy as defenses, and their costs. In this study we examined the distribution of sequestered iridoid glycosides (IGs) in two congeneric species of nymphalid butterfly, Euphydryas anicia and E. phaeton, and found that the pattern of localization of IGs differed between the two species. Although IG concentrations were quite high in the heads of both species, the relative concentrations in wings and abdomens differed substantially. Euphydryas anicia had relatively high IG concentrations in their abdomens and low IG concentrations in their wings, whereas the reverse was true in E. phaeton. We interpret these results in light of two current hypotheses regarding where sequestered chemicals should be localized: that they should be found in wings, which would allow non-lethal sampling by predators; and that their distribution is constrained by the distribution of tissue types to which sequestered compounds bind. We also offer the third hypothesis, that costs of storage may differ among body parts, and that the localization of compounds may reflect a cost-reduction strategy. Results from E. phaeton were consistent with all three of these non-mutually exclusive hypotheses, whereas results from E. anicia were only consistent with the notion that tissue bias among body parts plays a role in IG distribution. The finding that these two congeneric butterflies exhibit different patterns of IG localization suggests that they have been shaped by different selection regimes.

Localized air foci in the lower thorax in the patients with pneumothorax: skip pneumothoraces.

PURPOSE: To investigate the characteristics and imaging features of localized air foci in the lower thorax in patients with pneumothorax using thin-section multidetector computed tomography. MATERIALS AND METHODS: Of 10,547 consecutive CT examinations comprising the chest, the CT scans of 146 patients with ordinary pneumothoraces were identified and retrospectively evaluated. The study group included 110 male and 36 female patients (mean age, 50 years; range, 1-93 years). All examinations were performed at our institution between January 2009 and December 2009. Cause of pneumothorax was classified as traumatic or non-traumatic. Localized air foci in the lower thorax were defined as being localized air collections in the lower thorax that did not appear to be adjacent to the lung. If these criteria were met, the shape, size, location laterality, and number of foci were evaluated. Associations with trauma, sex, severity of the pneumothorax, and laterality were evaluated using the χ(2) test. All P values <0.05 were considered significant. RESULTS: Localized air foci in the lower thorax presented as slit-like or small ovoid air collections in the lowest part of the pleural space. These foci were observed in 79/146 (54.1%) patients. The traumatic pneumothoraces group showed a higher prevalence of these features than the non-traumatic group. Some foci that were situated in the anterior part mimicked the appearance of free intraperitoneal air. CONCLUSION: Patients with pneumothorax commonly had localized air foci in the lower thorax. Because such foci can mimic pneumoperitoneum, accurate recognition of them is required to avoid confusion with free intraperitoneal air, especially in traumatic cases.

Thoracic fraction of inhaled fiber aerosol.

Size-selective sampling is a health-related method to collect airborne particles based on penetration of inhaled particles into different regions of the human respiratory tract; thus, it is the most relevant sampling method to correlate health risks with occupational exposure. The current practice of sampling asbestos and other fibers is not a size-selective method. The thoracic size fraction, defined as the portion of inhaled particles that can penetrate through the larynx, has been suggested as the most relevant size-selective sampling method for fiber aerosol. The thoracic fraction is based on 1-deposition of inhaled spherical particles in the human extrathoracic airways for mouth breathing and corrected for the particle inhalability. There is no comparable information for fiber aerosols; therefore, there is no technical basis to ascertain whether the current thoracic fraction definition is suitable for fiber aerosols. No human data are available from controlled experiments of inhaled fiber aerosols for the obvious reason that most fiber materials are potentially hazardous when inhaled. Our approach was to measure penetration of fiber aerosol in realistic human oropharyngeal airway replicas and to compare that with data from spherical particles. We showed that realistic human oral airway replicas (including the oral cavity, pharynx, and larynx regions) provided useful spherical and fiber particle deposition in the human head airway. These data could be used to test the thoracic fraction curves. The spherical penetration is in agreement with human in vivo data used to establish the thoracic fraction curve. Fiber penetrations through the larynx of two human oral airway replicas were higher than those for spherical particles for the same aerodynamic diameter using the same replicas. The thoracic curve as defined for spherical particles, therefore, may not include some fibers that could penetrate to the thoracic region.

Mechanical analysis of Drosophila indirect flight and jump muscles.

The genetic advantages of Drosophila make it a very appealing choice for investigating muscle development, muscle physiology and muscle protein structure and function. To take full advantage of this model organism, it has been vital to develop isolated Drosophila muscle preparations that can be mechanically evaluated. We describe techniques to isolate, prepare and mechanically analyze skinned muscle fibers from two Drosophila muscle types, the indirect flight muscle and the jump muscle. The function of the indirect flight muscle is similar to vertebrate cardiac muscle, to generate power in an oscillatory manner. The indirect flight muscle is ideal for evaluating the influence of protein mutations on muscle and cross-bridge stiffness, oscillatory power, and deriving cross-bridge rate constants. Jump muscle physiology and structure are more similar to skeletal vertebrate muscle than indirect flight muscle, and it is ideal for measuring maximum shortening velocity, force-velocity characteristics and steady-state power generation.

Considerations in the choice of a skin donor site for harvesting keratinocytes containing a high proportion of stem cells for culture in vitro.

The treatment of severely burned patients has benefited from the grafting of skin substitutes obtained by expansion of epithelial cells in culture. The aim of this study was to evaluate whether the anatomic site chosen for harvesting skin had an impact on the quality of the derived cell cultures. Considering that hair follicles contain epithelial stem cells, we compared hairy skin sites featuring different densities and sizes of hair follicles for their capacity to generate high quality keratinocyte cultures. Three anatomic sites from adult subjects were compared: scalp, chest skin and p-auricular (comprising pre-auricular and post-auricular) skin. Keratin (K) 19 was used as a marker to evaluate the proportion of stem cells. Keratinocytes were isolated using the two-step thermolysin and trypsin cell extraction method, and cultured in vitro. The proportion of K19-positive cells harvested from p-auricular skin was about twice that of the scalp. This K19-positive cell content also remained higher during the first subcultures. In contrast to these in vitro results, the number of K19-positive cells estimated in situ on skin sections was about double in scalp as in p-auricular skin. Chest skin had the lowest number of K19-positive cells. These results indicate that in addition to the choice of an adult anatomic site featuring a high number of stem cells in situ, the quality of the cultures greatly depends on the ability to extract stem cells from the skin biopsy.

Effects of flavourzyme on yield and some biological activities of Mungoong, an extract paste from the cephalothorax of white shrimp.

The use of Flavourzyme as a processing aid for the production of Mungoong, an extract paste from the cephalothorax of white shrimp, and its effect on biological activities were investigated. Flavourzyme increased the yield of Mungoong. Mungoong prepared from raw cephalothorax with the addition of 0.15% or 0.30% (w/w) Flavourzyme showed higher yields than those prepared from the cooked counterpart with a higher formal content and nitrogen solubility index when the same levels of Flavourzyme were used (P < 0.05). Higher ammonia and amino nitrogen contents were also observed in Mungoong prepared with the aid of Flavourzyme, particularly for the raw cephalothorax (P < 0.05). All Mungoong contained protein as the major constituent. For antioxidative activity, Mungoong prepared from raw cephalothorax with 0.15% Flavourzyme (Mungoong RF 15) exhibited the highest antioxidative activity as evidenced by the highest 2,2-diphenyl-1-picryl hydrazyl and 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging activities as well as ferric reducing antioxidant power, compared with other samples. Additionally, angiotensin I converting enzyme inhibitory activity was greater in Mungoong produced with the aid of Flavourzyme, regardless of processes used (P < 0.05).

Metabolomic and flux-balance analysis of age-related decline of hypoxia tolerance in Drosophila muscle tissue.

The fruitfly Drosophila melanogaster is increasingly used as a model organism for studying acute hypoxia tolerance and for studying aging, but the interactions between these two factors are not well known. Here we show that hypoxia tolerance degrades with age in post-hypoxic recovery of whole-body movement, heart rate and ATP content. We previously used (1)H NMR metabolomics and a constraint-based model of ATP-generating metabolism to discover the end products of hypoxic metabolism in flies and generate hypotheses for the biological mechanisms. We expand the reactions in the model using tissue- and age-specific microarray data from the literature, and then examine metabolomic profiles of thoraxes after 4 h at 0.5% O(2) and after 5 min of recovery in 40- versus 3-day-old flies. Model simulations were constrained to fluxes calculated from these data. Simulations suggest that the decreased ATP production during reoxygenation seen in aging flies can be attributed to reduced recovery of mitochondrial respiration pathways and concomitant overdependence on the acetate production pathway as an energy source.

Main phospholipids and their fatty acid composition in muscle and cephalothorax of the edible Mediterranean crustacean Palinurus vulgaris (spiny lobster).

The total lipids of muscle and cephalothorax of Mediterranean lobster Palinurus vulgaris were found to be 1.0% and 2.4% of the wet tissue of which the phospholipids represented 66.5% and 47.5%, respectively. The main PhL saturated, monounsaturated and polyunsaturated fatty acids in muscle and cephalothorax were C16:0, C18:0, C18:1omega-9, C18:1omega-7, C20:4omega-6, C20:5omega-3 and C22:6omega-3. 2-OH C14:0 and cyclo-17:0 fatty acids were also identified though in low percentages. The main individual PhL in muscle were found to be phosphatidylcholine (53.5%), 72.0% of which corresponded to the structure of 1,2-diacyl-glycerocholine while the rest 28.0% to 1-O-alkyl-2-acyl-glycerocholine or 1-O-(1-alkenyl)-2-acyl-glycerocholine and phosphatidylethanolamine (19.3%), 75.0% of which corresponded to the structure of 1,2-diacyl-glyceroethanolamine and 25% to 1-O-alkyl-2-acyl-glyceroethanolamine or 1-O-(1-alkenyl)-2-acyl-glyceroethanolamine. Cephalothorax main PhL were found to be PC and PE (66.4% and 18.8%, respectively). In muscle and cephalothorax PC omega-3 fatty acids amounted 7.78% and 8.60%, while in PE amounted 30.77% and 23.65% respectively. Furthermore, in both tissues PhL, cardiolipine phosphatidylserine, phosphatidylinositol, sphingomyelin and lysophosphatidylcholine, were also found.

Magnetic material in head, thorax, and abdomen of Solenopsis substituta ants: a ferromagnetic resonance study.

Ferromagnetic resonance temperature dependence is used to study the magnetic material in smashed head, thorax, and abdomen of Solenopsis substituta ants. These three body parts present the five lines previously observed in other social insects. The magnetic material content is slightly higher in heads with antennae than in abdomen with petiole. Isolated nanoparticle diameters were estimated as 12.5 +/- 0.1 and 11.0 +/- 0.2 nm in abdomen with petiole and head with antennae, respectively. The presence of linear chains of these particles or large ellipsoidal particles are suggested. A bulk-like magnetite particle was observed in the thorax. The Curie-Weiss, the structural-electronic and ordering transition temperatures were obtained in good agreement with those proposed for magnetite nanoparticles.

Magnetic material arrangement in oriented termites: a magnetic resonance study.

Temperature dependence of the magnetic resonance is used to study the magnetic material in oriented Neocapritermes opacus (N.o.) termite, the only prey of the migratory ant Pachycondyla marginata (P.m.). A broad line in the g = 2 region, associated to isolated nanoparticles shows that at least 97% of the magnetic material is in the termite's body (abdomen + thorax). From the temperature dependence of the resonant field and from the spectral linewidths, we estimate the existence of magnetic nanoparticles 18.5+/-0.3 nm in diameter and an effective magnetic anisotropy constant, K(eff) between 2.1 and 3.2 x 10(4)erg/cm(3). A sudden change in the double integrated spectra at about 100K for N.o. with the long body axis oriented perpendicular to the magnetic field can be attributed to the Verwey transition, and suggests an organized film-like particle system.

A crustacean serotonin receptor: cloning and distribution in the thoracic ganglia of crayfish and freshwater prawn.

Serotonin (5-HT) is involved in regulating important aspects of behavior and a variety of systemic physiological functions in both vertebrates and invertebrates. These functions are mediated through binding to 5-HT receptors, of which approximately 13 have been characterized in mammals. In crustaceans, important model systems for the study of the neural basis of behaviors, 5-HT is also linked with higher-order behaviors, associated with different 5-HT receptors that have been identified at the physiological and pharmacological levels. However, no crustacean 5-HT receptors have been identified at the molecular level. We have cloned a putative 5-HT(1) receptor (5-HT(1crust)) from crayfish, prawn, and spiny lobster and have raised antibodies that recognize this protein in all three organisms. 5-HT(1crust) immunoreactivity (5-HT(1crust)ir) was observed surrounding the somata of specific groups of neurons and as punctate staining within the neuropil in all thoracic ganglia of crayfish and prawn. In the crayfish, 5-HT(1crust)ir was also found in boutons surrounding the first and second nerves of each ganglion and on the 5-HT cells of T1-4. In the prawn, 5-HT(1crust)ir was also found in axons that project across the ganglia and along the connectives. We found examples of colocalization of 5-HT(1crust) with 5-HT, consistent with the short-term modulatory role of 5-HT, as well as cases of serotonergic staining in the absence of a 5-HT(1crust) signal, which might imply that other 5-HT receptors are found at these locations. We also observed receptors that did not possess counterpart 5-HT staining, suggesting that these may also mediate long-term neurohormonal functions of serotonin.

Imaging of beta-adrenoceptors in the human thorax using (S)-[(11)C]CGP12388 and positron emission tomography.

We report positron emission tomography studies of beta-adrenoceptors in the human thorax with (S)-[(11)C]CGP12388 (4-(3-(2'-[(11)C]-isopropylamino)-2-hydroxypropoxy)-2H-benzimidazol-2-one). Beta-adrenoceptors have previously been quantified using (S)-[(11)C]CGP12177 (4-(3-tert-butylamino-2-hydroxypropoxy)-2H-benzimidazol-2[(11)C]-one), but (S)-[(11)C]CGP12388 is more easily prepared and therefore more suitable in a clinical setting. (S)-[(11)C]CGP12388 was administered to five healthy volunteers on two separate days (control and pindolol block study). Arterial plasma samples were used to determine clearance, metabolites, and protein binding of the radioligand. Heart, lung and spleen showed high uptake of radioactivity, which was strongly suppressed (68-77%) by pindolol. Plasma clearance of (S)-[(11)C]CGP12388 was rapid, binding to plasma proteins was low (53+/-4%), and the radioligand was slowly metabolized. (S)-[(11)C]CGP12388 produces high-quality images of the human thorax. Uptake of (S)-[(11)C]CGP12388 in heart, lung and spleen represents binding to beta-adrenoceptors. (S)-[(11)C]-CGP12388 seems useful for imaging of beta-adrenoceptors in a clinical setting.

Electron paramagnetic resonance study of the migratory ant Pachycondyla marginata abdomens.

Electron paramagnetic resonance was used to investigate the magnetic material present in abdomens of Pachycondyla marginata ants. A g congruent with 4.3 resonance of high-spin ferric ions and a very narrow g congruent with 2 line are observed. Two principal resonance broad lines, one with g > 4.5 (LF) and the other in the region of g congruent with 2 (HF), were associated with the biomineralization process. The resonance field shift between these two lines, HF and LF, associated with magnetic nanoparticles indicates the presence of cluster structures containing on average three single units of magnetite-based nanoparticles. Analysis of the temperature dependence of the HF resonance linewidths supports the model picture of isolated magnetite nanostructures of approximately 13 nm in diameter with a magnetic energy of 544 K. These particles are shown to present a superparamagnetic behavior at room temperature. The use of these superparamagnetic particle properties for the magnetoreception process of the ants is suggested.

Electron density of tissues and breast cancer radiotherapy: a quantitative CT study.

PURPOSE: To obtain more accurate data on the electron density of tissues to be used in the treatment planning of breast cancer patients. METHODS AND MATERIALS: Single kVp quantitative computed tomography was applied in 70 women, 20 to 77 years old, to study the electron density of the breast, the thoracic wall close and parallel to the breast, and the lung parenchyma. RESULTS: The electron density of the entire breast decreases with increasing age in premenopausal women and remains practically constant in postmenopausal women (8% less than that of water). No difference was found in the electron densities of the right and left breast. The electron density of the lung parenchyma in proximity to the breast is lower than the density in the entire lung parenchyma. CONCLUSIONS: Whenever no accurate data is available on individual patients, the electron density values to be used in treatment planning for breast and thoracic wall have to take into account both age and menstrual status. The regional differences in electron density of the lung also have to be considered.

Analysis of gunshot residues on human tissues and clothing by X-ray microfluorescence.

The analysis of gunshot residues on human tissues and clothing in suicide, homicide, suspicious death, or attempted murder events, permits the forensic scientist to confirm the possibility of an entry wound caused by a gunshot bullet from a pistol, revolver, rifle, etc. The residues to be detected are lead (Pb), antimony (Sb), barium (Ba) usually from the primer, copper (Cu) and zinc (Zn) usually from the metal jacket bullet and iron (Fe) possibly from the barrel of the gun used. The presence or absence of these elements and their relative concentrations can help in the interpretation of the event.

An on-line two-dimensional polyacrylamide gel electrophoresis protein database of adult Drosophila melanogaster.

An annotated two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) protein database of adult Drosophila melanogaster has been constructed, based on the protein patterns of heads, thoraces and abdomens of adult male and female Drosophila melanogaster. About 1200 major protein spots are catalogued. Common proteins, found in all body parts, as well as bodypart- and sex-specifically expressed proteins are reported. Of the major proteins, 91, or 7.5%, are differentially expressed in the two sexes or in different body parts, at least in part reflecting specific functional requirements. At the present time 43 proteins, or about 3.5% of the detected proteins, have been identified. These data can be accessed interactively from our World Wide Web (WWW) server through clickable inline gel images and hypertext links. Identified protein spots are cross-referenced, through hypertext links, to the SWISS-PROT annotated database of protein primary sequences and the Fly-Base database of Drosophila genomic data. Our reference gels can be used to gain immediate access to protein spot identify and to the pattern of differentially expressed proteins in Drosophila melanogaster. The work presented in this article ties together information from protein 2-D PAGE, molecular biology and genetics and offers a uniform way to access this large volume of data.