bluuebunny/crossref_metadata_2025 huggingface dataset:
Extreme sunbathing: Three weeks of small total O3 columns and high UV radiation over the southern tip of South America during the 2009 Antarctic O3 hole season
Authors: de Laat A. T. J. , van der A R. J. , Allaart M. A. F. , van Weele M. , Benitez G. C. , Casiccia C. , Paes Leme N. M. , Quel E. , Salvador J. , Wolfram E.
Published: July 2010

This paper presents an analysis of satellite and ground‐based measurements of total O3 columns and the UV index of a unique event during the 2009 Antarctic O3 hole season. From 11 to 30 November 2009 the Antarctic vortex was located just south of the southern tip of South America rather than at its climatological position over Antarctica. Analysis of 30 years of assimilated total O3 column and UV index measurements shows that this 20‐day event was unique in the history of the ozone hole for these latitudes. During this period, small total O3 columns and large UV index values were observed over the southern tip of South America. Comparison of ground‐based and satellite measurements of total O3 columns and satellite based calculations of the UVI index – never designed nor validated for such extreme Southern Hemisphere conditions – show excellent agreement.
Synovial expression of cell adhesion molecules in polymyalgia rheumatica
Authors: MELICONI R , PULSATELLI L , MELCHIORRI C , FRIZZIERO L , SALVARANI C , MACCHIONI P , UGUCCIONI M , FOCHERINI M C , FACCHINI A
Published: July 2010

Polymyalgia rheumatica (PMR) is a common disorder of the elderly: the pathogenesis of the syndrome is still debated, though active synovitis of the shoulder has recently been confirmed. To investigate the pathogenesis of this synovitis we evaluated cell adhesion molecule (CAM) expression in shoulder synovial tissue from patients with PMR, correlated synovial expression with the serum levels of soluble forms, and assessed the changes associated with corticosteroid treatment. Arthroscopic synovial biopsies were obtained from 12 untreated and seven corticosteroid (CS)-treated cases. CAM expression was evaluated by MoAb staining on frozen sections and computerized image analysis. Soluble CAM were quantified by ELISA. Endothelial cells expressed intercellular adhesion molecule-1 (ICAM-l), E- and P-selectins. Infiltrating cells were ICAM-1 and β1-integrin-positive, while L-selectin expression was limited to intravascular leucocytes. Synovial lining cells strongly expressed vascular cell adhesion molecule-1 (VCAM-1), and less intensely ICAM-1. Only the soluble form of ICAM-1 (sICAM-1) was elevated in untreated patients. CS treatment was associated with a decrease in ICAM-1, VCAM-1 and E- and P-selectin expression. sICAM-1 levels were in the normal range in treated patients. VLA-5 and 6 expression was widely distributed among cell types, and was not CS-sensitive. Active shoulder synovitis is associated with different CAM expression in PMR. ICAM-l expression is widely distributed and correlates with elevated levels of the soluble form; it is significantly lower in CS-treated asymptomatic cases.
Peripheral blood T cell expansions in patients with Behc¸et's disease
Authors: ESIN S , GÜL A , HODARA V , JEDDI-TEHRANI M , DILSEN N , KONIÇE M , ANDERSSON R , WIGZELL H
Published: July 2010

Behçet's disease (BD) is a chronic multisystemic inflammatory disorder characterized mainly by recurrent oral and genital aphthous ulcerations and uveitis. Etiology and pathogenesis of BD remain unknown. T cell receptor (TCR) Vα/Vβ gene product expression as well as Jβ gene segment expression in peripheral blood of BD patients were analysed to investigate the possible role of T lymphocytes in the etiopathogenesis of BD. Flow cytometry with 12 TCR V-specific MoAbs was used for TCRV analyses. Jβ gene segment usage by T cell populations expressing certain Vβs was determined by polymerase chain reaction (PCR) technique with Vβ- and Cβ-specific primers, Southern blotting of PCR products, and subsequent hybridization with radiolabelled Jβ gene segment-specific probes. Although 13 of the 23 BD patients exhibited increases in expression of one or more TCR V-gene products, only expansions among the CD4+ T cell subset were significantly more frequent in BD patients (7/23) compared with healthy controls (0/15) (P = 0.019). Six out of eight cases followed for up to 20 months had at least one expansion correlated with disease activity. A strict preference for particular Jβ gene segments implicating clonality was apparent in all analysed T cell expansions and correlated well with disease activity. These results suggest a possible involvement of antigen-specific T lymphocytes in the pathogenesis of BD.
Involvement of β2-glycoprotein I and anticardiolipin antibodies in oxidatively modified low-density lipoprotein uptake by macrophages
Authors: HASUNUMA Y , MATSUURA E , MAKITA Z , KATAHIRA T , NISHI S , KOIKE T
Published: July 2010

Anticardiolipin antibodies (aCL) in the sera of patients with antiphospholipid syndrome (APS) recognize an altered structure of β2-glycoprotein I (β2-GPI) interacting with solid-phase negatively charged phospholipids. β2-GPI bound to Cu2+-oxidized plasma lipoproteins, i.e. oxidized very low-density lipoprotein (oxVLDL), oxidized low-density lipoprotein (oxLDL), or oxidized high-density lipoprotein (oxHDL). β2-GPI inhibited in vitro uptake, i.e. cell surface binding, cellular association, and proteolytic degradation of oxLDL by murine macrophage J774A.1 cells. The binding of oxLDL to the macrophages was inhibited by the addition of polyinosinic acid (poly (I)), a competitor of the scavenger receptor, but not by another polyanionic acid, polycytidylic acid (poly (C)). Conversely, the binding of oxLDL was significantly increased by the simultaneous addition of human β2-GPI and monoclonal aCL derived from NZW × BXSB F1 (WB F1) mice, an animal model of APS, or anti-β-GPI antibodies from BALB/c mice immunized with human β2-GPI. These findings indicate that β2-GPI may be an anti-atherogenic protein and that the autoimmune response against β2-GPI may have a role in the development of atherosclerosis in APS.
Stomatal uptake and stomatal deposition of ozone in isoprene and monoterpene emitting plants
Authors: Fares S. , Loreto F. , Kleist E. , Wildt J.
Published: July 2010

Volatile isoprenoids were reported to protect plants against ozone. To understand whether this could be the result of a direct scavenging of ozone by these molecules, the stomatal and non‐stomatal uptake of ozone was estimated in plants emitting isoprene or monoterpenes. Ozone uptake by holm oak (Quercus ilex, a monoterpene emitter) and black poplar (Populus nigra, an isoprene emitter) was studied in whole plant enclosures (continuously stirred tank reactors, CSTR). The ozone uptake by plants was estimated measuring ozone concentration at the inlet and outlet of the reactors, after correcting for the uptake of the enclosure materials. Destruction of ozone at the cuticle or at the plant stems was found to be negligible compared to the ozone uptake through the stomata. For both plant species, a relationship between stomatal conductance and ozone uptake was found. For the poplar, the measured ozone losses were explained by the uptake of ozone through the stomata only, and ozone destruction by gas phase reactions with isoprene was negligible. For the oak, gas phase reactions of ozone with the monoterpenes emitted by the plants contributed significantly to ozone destruction. This was confirmed by two different experiments showing a) that in cases of high stomatal conductance but under low CO2 concentration, a reduction of monoterpene emission was still associated with reduced O3 uptake; and b) that ozone losses due to the gas phase reactions only can be measured when using the exhaust from a plant chamber to determine the gas phase reactivity in an empty reaction chamber. Monoterpenes can therefore relevantly scavenge ozone at leaf level contributing to protection against ozone.
The Characteristics of Small Island Tourist Economies
Authors: McElroy Jerome L. , Parry Courtney E.
Published: July 2010

This study presents a review of the literature and develops three related empirical analyses to determine the characteristics of small island tourist economies (SITEs). With a sample of 39 small islands, the first means difference analysis contrasts patterns in the Caribbean versus Pacific and Indian Ocean islands. The second, a Tourism Penetration Index, identifies the most successful SITEs along Butler's life cycle. The third is a regression analysis used to explain the uneven spread of island tourism across the globe. Results suggest the importance of adequate infrastructure, a degree of modernization, favorable location, uncrowded ambience and dependent political status in SITE success. However, only the last three are statistically significant influences.
Advertising and Tourist Arrivals: Evidence from Jamaica
Authors: Williams Densil A. , Spencer Andrew
Published: July 2010

The major tourist generating countries for Jamaica's tourism industry were the most heavily hit by the global financial crisis. With home foreclosure and job losses on the rise in these supplier markets, luxury goods such as travel are likely to be significantly reduced. The response of the Jamaican government has been to intensify advertising campaigns. Although this may on the face appear to generate interest, there has been very little empirical evidence to suggest that advertising increases arrivals. To address this gap, the current work assesses the impact of advertising, along with a number of economic variables (for example, exchange rate, inflation), on tourist arrivals in Jamaica. The data for this research were generated from a database which comprises 30 years of secondary time series data on the variables for Jamaica. These were analysed using multiple regression analysis. The results revealed that it is exchange rate, not advertising that is most influential in stimulating tourist arrivals to Jamaica.
Influence of lanthanum doping on the morphotropic phase boundary of lead zirconate titanate
Authors: Hinterstein Manuel , Schoenau Kristin A. , Kling Jens , Fuess Hartmut , Knapp Michael , Kungl Hans , Hoffmann Michael J.
Published: July 2010

A phase diagram for morphotropic (Pb0.985La0.01)(Zr1−xTix)O3 is proposed based on a combination of X-ray and neutron powder diffraction experiments and complemented by transmission electron microscopy. Dependent on composition three regions are characterized. The stability range of tetragonal microdomains for high Ti contents, the stability range of rhombohedral microdomains for low Ti contents, and an intermediate stability range of nanodomains. All three regions exhibit a corresponding low temperature configuration. Temperature dependent diffraction revealed that lanthanum doping reduces the sensitivity of the structure to changes in temperature and composition. A continuous transition from pseudorhombohedral to tetragonal symmetry with an intermediate two-phase region at the morphotropic phase boundary is observed. A similar transition of low temperature superstructure phases from pseudorhombohedral to pseudotetragonal with an intermediate monoclinic phase governed by a continuous change in the oxygen octahedral tilt system from a−a−a− over a−a−c− to a0a0c− is identified.
Complex phase compositions in nanostructured coatings as evidenced by photoelectron spectroscopy: The case of Al–Si–N hard coatings
Authors: Pélisson-Schecker Aude , Hug Hans Josef , Patscheider Jörg
Published: July 2010

The chemical state evolution of the Al–Si–N thin films at various Si contents is investigated by x-ray photoelectron spectroscopy (XPS). The detailed evolution of the Al 2p, Si 2p, and N 1s photoelectrons line positions and widths are used to identify different chemical environments as the Si content is changed. The results are compared to x-ray diffraction (XRD) data that indicate the formation of a two-phase Al1−xSixN/SiNy composite when the solubility limit of 6 at. % of Si in AlN is exceeded. In contrast to XRD data, no particular effect is observed in the XPS data at the solubility limit of Si. Instead, two compositional regions can be identified that are separated by a distinct change in the evolution of core level binding energy differences and chemical shifts at about 10–15 at. % of Si. This silicon concentration is identified as the onset of the formation of a SiNy intergranular phase that is a few monolayers thick, having a chemical bonding similar to that in bulk silicon nitride. The observed changes in the XPS data coincide well with the structural changes in the material at different silicon contents. The unambiguous identification of phases, especially of minority phases from XPS data, is, however, not possible.
The effect of annealing on the photoconductivity of carbon nanofiber/TiO2 core-shell nanowires for use in dye-sensitized solar cells
Authors: Rochford Caitlin , Li Zhuang-Zhi , Baca Javier , Liu Jianwei , Li Jun , Wu Judy
Published: July 2010

Electrical transport properties and photoresponse of individual TiO2-coated carbon nanofibers were studied in an attempt to elucidate the limiting factors of core-shell nanowire-based dye-sensitized solar cells (DSSC). The role of the semiconductor shell microstructure was investigated by comparing as grown and thermally annealed samples. Steady state I-V and transient photoconductivity measurements suggest that improving the microstructure leads to reduced resistivity and contact resistance, a decrease in charge traps, improved surface stoichiometry for dye adsorption, and reduced absorption of visible light by the semiconductor, all of which may improve nanowire-based DSSC performance.
Ferroelectric and antiferroelectric properties of AgNbO3 films fabricated on (001), (110), and (111)SrTiO3 substrates by pulsed laser deposition
Authors: Sakurai Hiroyuki , Yamazoe Seiji , Wada Takahiro
Published: July 2010

We fabricated 001, 110, and 111 oriented AgNbO3 (AN) films on (001)SrTiO3 (STO), (110)STO, and (111)STO substrates by pulsed laser deposition, respectively. Scanning electron microscope images showed that the surface textures of the AN films on the (001), (110), and (111)STO substrates took the form of a lattice-shaped pattern, a striped pattern, and trigonal pyramid-like structures, respectively. The AN film on the (001)STO had the largest relative dielectric constant of 748 in all films. The P-E hysteresis showed that the AN film on (001)STO exhibited antiferroelectricity and the other AN films deposited on (110) and (111)STO showed a ferroelectric property.
Flux pinning with a magnetic nanorod array
Authors: Kim K. , Ozmetin A. E. , Naugle D. G. , Lyuksyutov I. F.
Published: July 2010

We report measurements of the phase diagram and critical current of a PbBi superconducting film with an embedded square array of Ni nanorods. Strong hysteresis of the superconducting properties and an increase in the critical current which varies from several times to several orders of magnitude depending on the magnetic field and temperature is found.
Giant magnetoimpedance-based microchannel system for quick and parallel genotyping of human papilloma virus type 16/18
Authors: Yang Hao , Chen Lei , Lei Chong , Zhang Ju , Li Ding , Zhou Zhi-Min , Bao Chen-Chen , Hu Heng-Yao , Chen Xiang , Cui Feng , Zhang Shuang-Xi , Zhou Yong , Cui Da-Xiang
Published: July 2010

Quick and parallel genotyping of human papilloma virus (HPV) type 16/18 is carried out by a specially designed giant magnetoimpedance (GMI) based microchannel system. Micropatterned soft magnetic ribbon exhibiting large GMI ratio serves as the biosensor element. HPV genotyping can be determined by the changes in GMI ratio in corresponding detection region after hybridization. The result shows that this system has great potential in future clinical diagnostics and can be easily extended to other biomedical applications based on molecular recognition.
Sandra Schmid: Collaring endocytosis
Authors: Sedwick Caitlin
Published: July 2010

Schmid explores how the dynamin GTPase pinches off endocytic vesicles.
Ras-mediated activation of the TORC2–PKB pathway is critical for chemotaxis
Authors: Cai Huaqing , Das Satarupa , Kamimura Yoichiro , Long Yu , Parent Carole A. , Devreotes Peter N.
Published: July 2010

In chemotactic cells, G protein–coupled receptors activate Ras proteins, but it is unclear how Ras-associated pathways link extracellular signaling to cell migration. We show that, in Dictyostelium discoideum, activated forms of RasC prolong the time course of TORC2 (target of rapamycin [Tor] complex 2)-mediated activation of a myristoylated protein kinase B (PKB; PKBR1) and the phosphorylation of PKB substrates, independently of phosphatidylinositol-(3,4,5)-trisphosphate. Paralleling these changes, the kinetics of chemoattractant-induced adenylyl cyclase activation and actin polymerization are extended, pseudopodial activity is increased and mislocalized, and chemotaxis is impaired. The effects of activated RasC are suppressed by deletion of the TORC2 subunit PiaA. In vitro RasCQ62L-dependent PKB phosphorylation can be rapidly initiated by the addition of a PiaA-associated immunocomplex to membranes of TORC2-deficient cells and blocked by TOR-specific inhibitor PP242. Furthermore, TORC2 binds specifically to the activated form of RasC. These results demonstrate that RasC is an upstream regulator of TORC2 and that the TORC2–PKB signaling mediates effects of activated Ras proteins on the cytoskeleton and cell migration.
Modification of hERG1 channel gating by Cd2+
Authors: Abbruzzese Jennifer , Sachse Frank B. , Tristani-Firouzi Martin , Sanguinetti Michael C.
Published: July 2010

Each of the four subunits in a voltage-gated potassium channel has a voltage sensor domain (VSD) that is formed by four transmembrane helical segments (S1–S4). In response to changes in membrane potential, intramembrane displacement of basic residues in S4 produces a gating current. As S4 moves through the membrane, its basic residues also form sequential electrostatic interactions with acidic residues in immobile regions of the S2 and S3 segments. Transition metal cations interact with these same acidic residues and modify channel gating. In human ether-á-go-go–related gene type 1 (hERG1) channels, Cd2+ coordinated by D456 and D460 in S2 and D509 in S3 induces a positive shift in the voltage dependence of activation of ionic currents. Here, we characterize the effects of Cd2+ on hERG1 gating currents in Xenopus oocytes using the cut-open Vaseline gap technique. Cd2+ shifted the half-point (V1/2) for the voltage dependence of the OFF gating charge–voltage (QOFF-V) relationship with an EC50 of 171 µM; at 0.3 mM, V1/2 was shifted by +50 mV. Cd2+ also induced an as of yet unrecognized small outward current (ICd-out) upon repolarization in a concentration- and voltage-dependent manner. We propose that Cd2+ and Arg residues in the S4 segment compete for interaction with acidic residues in S2 and S3 segments, and that the initial inward movement of S4 associated with membrane repolarization displaces Cd2+ in an outward direction to produce ICd-out. Co2+, Zn2+, and La3+ at concentrations that caused ∼+35-mV shifts in the QOFF-V relationship did not induce a current similar to ICd-out, suggesting that the binding site for these cations or their competition with basic residues in S4 differs from Cd2+. New Markov models of hERG1 channels were developed that describe gating currents as a noncooperative two-phase process of the VSD and can account for changes in these currents caused by extracellular Cd2+.
CTX-M–Type Fluoroquinolone-Resistant Escherichia coli: Analysis of the Colonization of Residents and Inanimate Surfaces 1 Year after a First Case of Urinary Tract Infection at a Nursing Home in France
Authors: van der Mee-Marquet Nathalie , Savoyen Philippe , Domelier-Valentin Anne-Sophie , Mourens Chantal , Quentin Roland ,   Réseau des Hygiénistes du Centre Study Group
Published: July 2010

At a nursing home in France, 1 year after a first case of a cluster of urinary tract infections caused by an extended-spectrum β-lac-tamase-producing strain of Escherichia coli, 9 (22%) of 49 residents were colonized with the outbreak strain. Colonization was associated with poor state of health (P = .04), incontinence (P = .007), and recent treatment with fluoroquinolones and/or cephalosporins (P = .003).
A Simple Educational Intervention to Decrease Incidence of Central Line–Associated Bloodstream Infection (CLABSI) in Intensive Care Units with Low Baseline Incidence of CLABSI
Authors: Parra Alfonso Pérez , Menárguez María Cruz , Granda María Jesús Pérez , Tomey María Jesús , Padilla Belén , Bouza Emilio
Published: July 2010

After an educational intervention in 3 intensive care units, 34 central line-associated bloodstream infections occurred in 11,582 central venous catheter [CVC]-days, compared with 45 episodes in 10,661 CVC-days before intervention (4.22 vs 2.94 episodes per 1,000 CVC-days [30.9% reduction]; P = .03, Wilcoxon rank sum test; P = .11, Poisson regression analysis).
The use of orthogonal projection to visualize mosaic domains from topographic data collected on protein crystals
Authors: Lovelace Jeffrey J. , Borgstahl Gloria E. O.
Published: July 2010

Owing to the relatively low intensity of diffraction and resulting low contrast topographic images, and also the richly detailed domain structure, it can be difficult to extract individual mosaic domains (size and shape) for protein crystals. Here, orthogonal projection was tested and found to be a superior approach over previous methods to extract mosaic domain information from a fine-sliced topographic sequence of images. The topographic sequence was collected at room temperature from a lysozyme crystal with a low mosaicity. Orthogonal projection can be applied to image sequences that are spatially invariant and composed of linearly additive image formation processes. The domains were determined by using a particle swarm optimizer to fit Gaussians to the integrated intensity profile of the reflection as a function of angle, although any basis function could have been used. This optimization method was more amenable to automation and converged to the global minimum more efficiently. The number of domains can normally be determined by visually inspecting the integrated intensity profile; this only needs to be done once for each crystal, because the number of domains remains constant during collection, and not for each reflection. The results can be used to provide a picture of the internal structure of the crystal and may be useful to aid in the improvement of crystal growth techniques.
Remarkable precision of the 90-year-old dynamic diffraction theories of Darwin and Ewald
Authors: Agamalian Michael , Carpenter John M. , Treimer Wolfgang
Published: July 2010

The reflectivity functions calculated by Bonse and Hart for a multi-bounce channel-cut single crystal, following the 90-year-old theories of Darwin and Ewald, exhibit extremely narrow nearly rectangular profiles. This feature provides ultra-high angular resolution and sensitivity for the double-crystal diffractometers now widely used for the observation of large-scale structures in condensed matter. However, the experimental results are several orders of magnitude poorer than the theoretical prediction, the `wings problem'. The reason for this discrepancy has remained unidentified for more than 40 years, creating problems for both theory and practice. A solution to this problem for neutron diffraction in Si channel-cut crystals is presented here. The results enable nearly theoretical functions of multiple reflectivity to be obtained, demonstrate the remarkable precision of the Darwin and Ewald theories in the range of the wings, and give rise to much improved sensitivity for the next generation of Bonse–Hart double-crystal diffractometers.
Structure ofStaphylococcus aureusadenylosuccinate lyase (PurB) and assessment of its potential as a target for structure-based inhibitor discovery
Authors: Fyfe Paul K. , Dawson Alice , Hutchison Marie-Theres , Cameron Scott , Hunter William N.
Published: July 2010

The medium-resolution structure of adenylosuccinate lyase (PurB) from the bacterial pathogenStaphylococcus aureusin complex with AMP is presented. Oxalate, which is likely to be an artifact of crystallization, has been modelled in the active site and occupies a position close to that where succinate is observed in orthologous structures. PurB catalyzes reactions that support the provision of purines and the control of AMP/fumarate levels. As such, the enzyme is predicted to be essential for the survival ofS. aureusand to be a potential therapeutic target. Comparisons of this pathogen PurB with the enzyme fromEscherichia coliare presented to allow discussion concerning the enzyme mechanism. Comparisons with human PurB suggest that the close similarity of the active sites would make it difficult to identify species-specific inhibitors for this enyme. However, there are differences in the way that the subunits are assembled into dimers. The distinct subunit–subunit interfaces may provide a potential area to target by exploiting the observation that creation of the enzyme active site is dependent on oligomerization.
Validation of crystallographic models containing TLS or other descriptions of anisotropy
Authors: Zucker Frank , Champ P. Christoph , Merritt Ethan A.
Published: July 2010

The use of TLS (translation/libration/screw) models to describe anisotropic displacement of atoms within a protein crystal structure has become increasingly common. These models may be used purely as an improved methodology for crystallographic refinement or as the basis for analyzing inter-domain and other large-scale motions implied by the crystal structure. In either case it is desirable to validate that the crystallographic model, including the TLS description of anisotropy, conforms to our best understanding of protein structures and their modes of flexibility. A set of validation tests has been implemented that can be integrated into ongoing crystallographic refinement or run afterwards to evaluate a previously refined structure. In either case validation can serve to increase confidence that the model is correct, to highlight aspects of the model that may be improved or to strengthen the evidence supporting specific modes of flexibility inferred from the refined TLS model. Automated validation checks have been added to thePARVATIandTLSMDweb servers and incorporated into theCCP4iuser interface.
Structural basis of the histidine-mediated vitamin D receptor agonistic and antagonistic mechanisms of (23S)-25-dehydro-1α-hydroxyvitamin D3-26,23-lactone
Authors: Kakuda Shinji , Ishizuka Seiichi , Eguchi Hiroshi , Mizwicki Mathew T. , Norman Anthony W. , Takimoto-Kamimura Midori
Published: July 2010

TEI-9647 antagonizes vitamin D receptor (VDR) mediated genomic actions of 1α,25(OH)2D3in human cells but is agonistic in rodent cells. The presence of Cys403, Cys410 or of both residues in the C-terminal region of human VDR (hVDR) results in antagonistic action of this compound. In the complexes of TEI-9647 with wild-type hVDR (hVDRwt) and H397F hVDR, TEI-9647 functions as an antagonist and forms a covalent adduct with hVDR according to MALDI–TOF MS. The crystal structures of complexes of TEI-9647 with rat VDR (rVDR), H305F hVDR and H305F/H397F hVDR showed that the agonistic activity of TEI-9647 is caused by a hydrogen-bond interaction with His397 or Phe397 located in helix 11. Both biological activity assays and the crystal structure of H305F hVDR complexed with TEI-9647 showed that the interaction between His305 and TEI-9647 is crucial for antagonist activity. This study indicates the following stepwise mechanism for TEI-9647 antagonism. Firstly, TEI-9647 forms hydrogen bonds to His305, which promote conformational changes in hVDR and draw Cys403 or Cys410 towards the ligand. This is followed by the formation of a 1,4-Michael addition adduct between the thiol (–SH) group of Cys403 or Cys410 and theexo-methylene group of TEI-9647.
Crystallographic binding studies with an engineered monomeric variant of triosephosphate isomerase
Authors: Salin Mikko , Kapetaniou Evangelia G. , Vaismaa Matti , Lajunen Marja , Casteleijn Marco G. , Neubauer Peter , Salmon Laurent , Wierenga Rik K.
Published: July 2010

Crystallographic binding studies have been carried out to probe the active-site binding properties of a monomeric variant (A-TIM) of triosephosphate isomerase (TIM). These binding studies are part of a structure-based directed-evolution project aimed towards changing the substrate specificity of monomeric TIM and are therefore aimed at finding binders which are substrate-like molecules. A-TIM has a modified more extended binding pocket between loop-7 and loop-8 compared with wild-type TIM. The A-TIM crystals were grown in the presence of citrate, which is bound in the active site of each of the two molecules in the asymmetric unit. In this complex, the active-site loops loop-6 and loop-7 adopt the closed conformation, similar to that observed in liganded wild-type TIM. Extensive crystal-soaking protocols have been developed to flush the bound citrate out of the active-site pocket of both molecules and the crystal structure shows that the unliganded open conformation of the A-TIM active site is the same as in unliganded wild-type TIM. It is also shown that sulfonate compounds corresponding to the transition-state analogue 2-phosphoglycolate bind in the active site, which has a closed conformation. It is also shown that the new binding pocket of A-TIM can bind 3-phosphoglycerate (3PGA; an analogue of a C4-sugar phosphate) and 4-phospho-D-erythronohydroxamic acid (4PEH; an analogue of a C5-sugar phosphate). Therefore, these studies have provided a rationale for starting directed-evolution experiments aimed at generating the catalytic properties of a C5-sugar phosphate isomerase on the A-TIM framework.
Structural and functional analysis of Rv0554 fromMycobacterium tuberculosis: testing a putative role in menaquinone biosynthesis
Authors: Johnston Jodie M. , Jiang Ming , Guo Zhihong , Baker Edward N.
Published: July 2010

Mycobacterium tuberculosis, the cause of tuberculosis, is a devastating human pathogen against which new drugs are urgently needed. Enzymes from the biosynthetic pathway for menaquinone are considered to be valid drug targets. The protein encoded by the open reading frameRv0554has been expressed, purified and subjected to structural and functional analysis to test for a putative role in menaquinone biosynthesis. The crystal structure of Rv0554 has been solved and refined in two different space groups at 2.35 and 1.9 Å resolution. The protein is dimeric, with an α/β-hydrolase monomer fold. In each monomer, a large cavity adjacent to the catalytic triad is enclosed by a helical lid. Dimerization is mediated by the lid regions. Small-molecule additives used in crystallization bind in the active site, but no binding of ligands related to menaquinone biosynthesis could be detected and functional assays failed to support possible roles in menaquinone biosynthesis.
Enhancing MADFAdata for substructure determination
Authors: Xu Hongliang
Published: July 2010

Heavy-atom substructure determination is a critical step in phasing an unknown macromolecular structure. Dual-space (Shake-and-Bake) recycling is a very effective procedure for locating the substructure (heavy) atoms usingFAdata estimated from multiple-wavelength anomalous diffraction. However, the estimatedFAare susceptible to the accumulation of errors in the individual intensity measurements at several wavelengths and from inaccurate estimation of the anomalous atomic scattering correctionsf′ andf′′. In this paper, a new statistical and computational procedure which merges multipleFAestimates into an averaged data set is used to further improve the quality of the estimated anomalous amplitudes. The results of 18 Se-atom substructure determinations provide convincing evidence in favor of using such a procedure to locate anomalous scatterers.
Promoting crystallization of antibody–antigen complexesviamicroseed matrix screening
Authors: Obmolova Galina , Malia Thomas J. , Teplyakov Alexey , Sweet Raymond , Gilliland Gary L.
Published: July 2010

The application of microseed matrix screening to the crystallization of antibody–antigen complexes is described for a set of antibodies that include mouse anti-IL-13 antibody C836, its humanized version H2L6 and an affinity-matured variant of H2L6, M1295. The Fab fragments of these antibodies were crystallized in complex with the antigen human IL-13. The initial crystallization screening for each of the three complexes included 192 conditions. Only one hit was observed for H2L6 and none were observed for the other two complexes. Matrix self-microseeding using these microcrystals yielded multiple hits under various conditions that were further optimized to grow diffraction-quality H2L6 crystals. The same H2L6 seeds were also successfully used to promote crystallization of the other two complexes. The M1295 crystals appeared to be isomorphous to those of H2L6, whereas the C836 crystals were in a different crystal form. These results are consistent with the concept that the conditions that are best for crystal growth may be different from those that favor nucleation. Microseed matrix screening using either a self-seeding or cross-seeding approach proved to be a fast, robust and reliable method not only for the refinement of crystallization conditions but also to promote crystal nucleation and increase the hit rate.
Energy‐dispersive X‐ray Analysis of the Mitochondria of Sideroblastic Anaemia
Authors: Grasso Joseph A. , Myers Thomas J. , Hines John D. , Sullivan Albert L.
Published: July 2010

Summary. Energy‐dispersive X‐ray analysis has been performed on marrow sideroblasts obtained from 10 patients with sideroblastic anaemias or erythroleukaemia (six primary refractory sideroblastic anaemia, two pyridoxine‐responsive, one secondary sideroblastic anaemia, two erythroleukaemia). Irrespective of the nature of the disorder associated with the presence of sideroblasts, X‐ray analysis of siderotic mitochondria consistently revealed the presence of iron and phosphorus with the average Fe/P intensity ratio measuring 1.4–1.5. Other elements variably detected within siderotic mitochondria included calcium, lead, potassium and zinc. Variation in the presence of these latter elements was detected not only between different patients, but also within different samples taken at different times from a single patient, and even among different cells of the same sample. Despite the detection of lead in siderotic mitochondria of a significant number of patients (five out of seven), there was no clinical evidence of lead toxicity. The elemental composition of the intramitochondrial deposits in sideroblasts was distinct from that of ferritin or haemosiderin and probably consists of ferric phosphate, possibly ferric orthophosphate (FePo4).
Subclasses of IgG Antibodies in Immune Thrombocytopenic Purpura (ITP)
Authors: Rose W. F. , Adams J. P. , Yount W.J.
Published: July 2010

Summary. The subclasses of platelet‐associated IgG were measured on the platelets of patients with immune thrombocytopenic purpura (ITP) by modification of the antiglobulin consumption test. All subclasses were found. In most patients, one subclass was predominant; this was usually IgG1. The platelet‐associated IgG in normal donors is predominantly IgGl and IgG2.
β‐Thalassaemia in Campania: DNA polymorphism analysis in βA and βthat chromosomes and its usefulness in prenatal diagnosis
Authors: Carestia C. , Pagano L. , Ioretti G. F , Mastrobuoni A.
Published: July 2010

In order to evaluate the feasibility of first trimester prenatal diagnosis of β‐thalassaemia by restriction fragment length polymorphism (RFLP) in Campania, one of the most affected regions in Southern Italy, DNA polymorphism analysis was performed on 40 unrelated patients, affected with homozygous β‐thalassaemia, and on their parents. Frequency of the presence of the Hinc II ε, Hind III Gγ and Aγ, Hinc II SPSβ and SPSβ, Ava II SPSβ, Ava II β and Bam HI 3′β sites have been determined in the βA and βthat chromosome samples. In 31 families (over 75%). RFLPs enabled tracing the β‐thalassaemia mutations in both father and mother (100% diagnosis). In the remaining nine families, RFLPs enabled tracing only one of the two mutations (50% diagnosis) because the other parent was found to be homozygous in all the analysed polymorphic sites. Restriction haplotypes, assembled on the basis of linkage analysis, were most heterogeneous, hence a wide heterogeneity of mutations is expected.
Peripheral blood and bone marrow abnormalities in patients with HIV related disease
Authors: Treacy Marilyn , Lai Leslie , Costello Christine , Clark Andrew
Published: July 2010

Between February 1983 and April 1986 we studied peripheral blood and bone marrow samples from 20 patients with human immunodeficiency virus (HIV) related disease. 14 patients had AIDS, three had ARC, two had PGL and one had ITP as a sole manifestation of HIV related disease.

Peripheral blood abnormalities included marked anisocytosis and poikilocytosis, rouleaux formation, neutropenia, lymphopenia, monocytopenia, a left shift in the granulocyte series and, in the patients with AIDS, vacuolated monocytes.

The most frequent bone marrow abnormalities were reticuloendothelial iron block, dyserythropoiesis, megaloblastic change and erythroid hypoplasia. Excess histiocytes were noted in four marrows, one exhibiting haemophagocytosis. None of the bone marrows showed lymphopenia. Eight of the 20 marrows were difficult or impossible to aspirate. None of the trephine biopsies showed increased reticulin.

The causes of these abnormalities are probably multiple and include opportunistic infections, drug therapy, immune mechanisms and possibly direct insult by the HIV virus.
Interactions between prostacyclin analogue ZK 36374 and heparin in their effects upon platelet function
Authors: Yardumian D. A. , O'Flynn K. , Linch D. C. , Machin S. J.
Published: July 2010

. Prostacyclin and heparin can be administered to patients concurrently, with the aim of maximizing anticoagulant effect. It has been suggested that the two agents may interact in their effects upon platelet function. We have therefore studied this parameter in the presence of ZK 36374, a chemícally stable analogue of prostacyclin and heparin, alone and in combination. Platelet responses to ADP and thrombin have been studied by standard aggregometry, and intra‐platelet calcium mobilization has simultaneously been recorded after loading the cells with the fluorescent calcium indicator, Quin 2. ZK 36374 alone consistently inhibited platelet aggregation and calcium mobilization in response to ADP and thrombin, in a dose‐dependent fashion. Heparin alone did not signiflcantly alter the responses to ADP but consistently reduced those to thrombin. Pre‐mixture of ZK36374, with heparin, was shown to interfere with the ability of the prostacyclin analogue to inhibit platelet aggregation and calcium flux in response to ADP. In contrast, ZK 36374 inhibition of platelet responses was markedly enhanced when thrombin was the agonist.
Carrier detection strategy in haemophilia A: the benefits of combined DNA marker analysis and coagulation testing in sporadic haemophilic families
Authors: Lillicrap David , Holden Jeanette J. A. , Giles Alan R. , White Bradley N. ,   The Ontario Haemophilia Study Group
Published: July 2010

A province wide study of carrier detection methods in haemophilia A is reported. The principal objective of this project was to compare the relative merits of coagulation testing and DNA marker analysis in carrier diagnosis for a large unselected haemophilic population. Factor VIII:C (F.VIII:C) and von Willebrand factor antigen (vWf:Ag) were measured on plasma samples sent to a central laboratory. Coagulation results were analysed by a logistic regression model of discrimination. Of 91 potential carriers tested, 15% had indeterminate coagulation test carrier probabilities. Two restriction fragment length polymorphisms were analysed in 123 women (42 obligate carriers and 81 potential carriers). The BcII polymorphism within the F.VIII gene and the locus DXS 52, approximately 5 cM (centimorgan) from F.VIII were used as DNA markers. Of the 81 potential carriers tested with RFLP analysis, a carrier diagnosis was achieved in 52%. Studies with the F.VIII intragenic BglI polymorphism in 23 of these families gave no additional information. Thirty‐nine potential carriers remained undiagnosed after DNA marker analysis. Twenty‐seven of these women were from families with a sporadic case of haemophilia. In this group of 27 women, 14 were found to have high probability carrier estimates derived from their coagulation tests. Combined coagulation and RFLP data was available in 42 potential carriers. Disagreement between DNA and coagulation carrier diagnoses was found in four instances. In each case, the coagulation data resulted in a carrier probability of indeterminate value. This study emphasizes some of the limitations associated with DNA marker linkage analysis as it pertains to haemophilia A carrier detection. Where a previous family history exists and appropriate females are informative for the DNA markers, this type of analysis is very productive. However, large numbers of potential carriers from ‘sporadic’haemophilia families were a factor in this project. With this in mind, an optimal service for haemophilia A carrier diagnosis must continue to offer reliable coagulation test probabilities in addition to DNA marker studies.
Dietary fish lipids do not diminish platelet adhesion to subendothelium
Authors: Owens Michael R. , Cave William T.
Published: July 2010

Clinical trials suggest that dietary fish lipids offer protection against pathologic thrombosis. Measurements of platelet aggregation and bleeding time indicate that these protective effects are mediated in part through alterations of platelet function. We studied the effects of supplementary fish lipids (MaxEPA) on platelet adhesion to arterial subendothelium utilizing a modified Baumgartner flow chamber. Template bleeding times were also performed, and platelet lipid composition was determined by gas chromatographic analysis. The results showed that platelet lipid composition was altered significantly during the study. However, total platelet adhesion to arterial subendothelium was not changed by ingestion of fish lipids during the study period. Bleeding times showed some prolongation in most subjects (range: ‐1.0 to +6.75 min), with a mean change of +0.75 min. We conclude that, in individuals maintained on ad libitum diets, the supplemental amount of fish oil used in this study was enough to induce a significant alteration in platelet lipid composition, but these changes were unaccompanied by any impairment in the ability of platelets to adhere to subendothelium in our model system.
Altered expression of the retinoblastoma susceptibility gene in chronic lymphocytic leukaemia
Authors: Neubauer Andreas , Kant Eric de , Rochlitz Christoph , Laser Jutta , Zanetta Ana Maria , Gallardo Jorge , Oertel Joachim , Herrmann Richard , Huhn Dieter
Published: July 2010

Summary. The pathogenesis of chronic lymphocytic leukaemia (CLL) is unknown. One of the most frequent cytogenetic abnormalities in CLL is a deletion within the long arm of chromosome 13, the region to which the retinoblastoma (Rb) gene has been mapped. Lack of Rb expression has been linked to the carcinogenic process in many human tumours. We therefore sought to investigate the role of Rb gene inactiva‐tion in CLL using differential polymerase chain reaction on reverse transcribed RNA. The result of the PCR was quanti‐tated using HPLC. 5/39 patients revealed a lack or significantly impaired expression of the Rb gene upon differential PCR analysis. In addition, immunocytochemical studies were performed using the Rb‐specific monoclonal antibody PMG245. 10/56 patients showed a weak or absent expression upon immunocytochemical analysis compared to monocytes or granulocytes. The samples lacking Rb were from both early and late stage CLL. Our results indicate that inactivation of the Rb protein occurs in a fraction of CLL cases and can be found in early and late stages of the disease.
Coexistence of hereditary spherocytosis (HS) due to band 3 deficiency and β‐thalassaemia trait: partial correction of HS phenotype
Authors: Giudice Emanuele Miraglia del , Perrotta Silverio , Nobili Bruno , Pinto Luciano , Cutillo Luisa , Iolascon Achille
Published: July 2010

Summary. A kindred with hereditary spherocytosis and β‐thalassaemia trait was identified. Detailed studies of the red cell membrane proteins on polyacrylamide gels with sodium dodecyl sulphate (SDS‐PAGE) demonstrated the presence of band 3 (anion transporter) deficiency in all HS subjects (20–25% reduction) whereas spectrin content was in the normal range. The molecular defect of β thalassaemia in this kindred was due to a β° codon 39 (C‐T) mutation, as assessed by β globin gene amplification and ASO‐probe hybridization. Seven subjects of this family were studied: two were normal, two had HS alone, two co‐inherited HS and β‐thalassaemia trait, and one had β‐thalassaemia trait only. The two subjects with HS alone had a typical clinical form of spherocytosis with anaemia, reticulocytosis and increased red cell osmotic fragility. The two with both HS and β‐thalassaemia trait were not anaemic and showed a small, well‐compensated haemoIysis. Hence the finding of red cells with abnormalities of both HS and β‐thalassaemia indicates that β‐thalassaemic trait ‘silences’ HS caused by band 3 deficiency.
Granulocyte colony‐stimulating factor (G‐CSF) downregulates its receptor (CD114) on neutrophils and induces gelatinase B release in humans
Authors: Jilma Bernd , Hergovich Nicole , Homoncik Monika , Jilma‐Stohlawetz Petra , Kreuzer Claudia , Eichler Hans‐Georg , Zellner Maria , Pugin Jerome
Published: July 2010

Despite the increasing use of granulocyte colony‐stimulating factor (G‐CSF) for the mobilization of stem cells and neutrophils, its pharmacodynamic actions are not fully understood. Because of the roles of G‐CSF and gelatinase B in leucokinetics, we set out to characterize the interaction of G‐CSF with its receptor in humans and its effects on gelatinase B release. G‐CSF was infused at bolus doses of 1 μg/kg and 5 μg/kg, and compared to placebo and dexamethasone (1 mg/kg b.i.d), which enhances the plasma levels of endogenous G‐CSF. The study was randomized, double‐blind, four‐way crossover, in eight healthy male volunteers. G‐CSF dose‐independently induced profound neutropenia (> 95%) within minutes and downregulated its own receptor (CD114) on neutrophils by 75%. The G‐CSF/CD114 interaction dose‐independently induced degranulation of neutrophils as evidenced by a 300–400% increase in CD11b expression. Degranulation induced up to a 10‐fold increase in plasma levels of gelatinase B, an enzyme known to precipitate neutropenia and subsequent neutrophilia in animals. In this study, it was shown that G‐CSF downmodulates CD114 expression on the surface of neutrophils in humans and the consequent degranulation enhances gelatinase B release into plasma, which may contribute to mobilization of neutrophils or stem cells.
Two novel factor VII gene mutations in a Chinese family with factor VII deficiency
Authors: Au W. Y. , Lam C. C. K. , Chan E. C. , Kwong Y. L.
Published: July 2010

We report two novel factor VII (FVII) gene mutations in a Chinese family with FVII deficiency. The proband, a 55‐year‐old woman, was incidentally found to have right shoulder arthritis consistent with chronic haemophilic arthropathy. FVII studies showed a FVII activity of 0·02 iu/ml and a FVII antigen of 49%. Molecular analysis showed a double heterozygous state, with an exon 4 nonsense mutation (C6003→A; Cys61→Term) and an exon 8 missense mutation (T10902→G; Cys329→Gly) that disrupted a Cys310/Cys329 disulphide bond. The genotypes and phenotypes were correlated in the patient's daughters. Two daughters were heterozygous for the Cys61→Term mutation and showed a type 1 FVII gene mutation phenotype consistent with a nonsense mutation. One daughter was heterozygous for the Csy329→Gly mutation and showed a type 2 mutation phenotype consistent with a missense mutation. These are the first reported FVII gene mutations in the Chinese people.