bluuebunny/crossref_metadata_2025_split huggingface dataset:
Microarray data mining with visual programming
Authors: Curk Tomaz , Demsar Janez , Xu Qikai , Leban Gregor , Petrovic Uros , Bratko Ivan , Shaulsky Gad , Zupan Blaz 
Published: August 2004

Abstract
Summary: Visual programming offers an intuitive means of combining known analysis and visualization methods into powerful applications. The system presented here enables users who are not programmers to manage microarray and genomic data flow and to customize their analyses by combining common data analysis tools to fit their needs.
Availability: http://www.ailab.si/supp/bi-visprog
Contact: blaz.zupan@fri.uni-lj.si
Supplementary information: http://www.ailab.si/supp/bi-visprog
Evaluation of rapid volume changes of substrate‐adherent cells by conventional microscopy 3D imaging
Authors: BOUDREAULT F. , GRYGORCZYK R. 
Published: August 2004

SummaryPrecise measurement of rapid volume changes of substrate‐adherent cells is essential to understand many aspects of cell physiology, yet techniques to evaluate volume changes with sufficient precision and high temporal resolution are limited. Here, we describe a novel imaging method that surveys the rapid morphology modifications of living, substrate‐adherent cells based on phase‐contrast, digital video microscopy. Cells grown on a glass substrate are mounted in a custom‐designed, side‐viewing chamber and subjected to hypotonic swelling. Side‐view images of the rapidly swelling cell, and at the end of the assay, an image of the same cell viewed from a perpendicular direction through the substrate, are acquired. Based on these images, off‐line reconstruction of 3D cell morphology is performed, which precisely measures cell volume, height and surface at different points during cell volume changes. Volume evaluations are comparable to those obtained by confocal laser scanning microscopy (ΔVolume ≤ 14%), but our method has superior temporal resolution limited only by the time of single‐image acquisition, typically ∼100 ms. The advantages of using standard phase‐contrast microscopy without the need for cell staining or intense illumination to monitor cell volume make this system a promising new tool to investigate the fundamentals of cell volume physiology.
MLJ Reviews
Authors: Liskin‐Gasparro Judith E. 
Published: August 2004

Books reviewed in this article:Bowker, Lynne, & Jennifer Pearson. Working with Specialized Language.Brown, James D., & Theodore S. Rodgers. Doing Second Language Research.De Kock, Josse, & Carmen Gómez Molina. Lingüística aplicada. La lengua: meta, materia y referencia en investigación, enseñanza y estilística.Granger, Sylviane, Joseph Hung, & Stephanie Petch‐Tyson. (Eds.). Computer Learner Corpora, Second Language Acquisition, and Foreign Language Teaching.Gubbins, Paul, & Mike Holt. (Eds.). Beyond Boundaries: Language and Identity in Contemporary Europe.Haverkate, Henk. The Syntax, Semantics and Pragmatics of Spanish Mood.Johnson, Karen E., & Paula R. Golombek. Teachers' Narrative Inquiry as Professional Development.Kikuchi, Masato. A Systematic Approach to Business Japanese.Lazaraton, Anne. A Qualitative Approach to the Validation of Oral Language Tests.Leaver, Betty Lou, & Boris Shekhtman. (Eds.). Developing Professional‐Level Language Proficiency.Medgyes, Peter. Laughing Matters: Humour in the Language Classroom.Miquel, Lourdes, & Neus Sans. ¿Eres tú, María?Miquel, Lourdes, & Neus Sans. La llamada de La Habana.Miquel, Lourdes, & Neus Sans. Lejos de casa.Miquel, Lourdes, & Neus Sans. Poderoso caballero.Miquel, Lourdes, & Neus Sans. Por amor al arte.Miquel, Lourdes, & Neus Sans. Una nota falsa.Miquel, Lourdes, & Neus Sans. Vacaciones al sol.Nakajima, Kazuko. (Ed.). Learning Japanese in the Network Society.Salaberry, M. Rafael, & Yasuhiro Shirai. (Eds.). The L2 Acquisition of Tense‐Aspect Morphology.Sampaio, João, & Barbara Mcintyre. Colloquial Portuguese: The Complete Course for Beginners.Schleppegrell, Mary J., & M. Cecilia Colombi. (Eds.). Developing Advanced Literacy in First and Second Languages: Meaning with Power.Sullivan, Joann Hammadou. (Ed.). Research in Second Language Learning, Vol. 1: Literacy and the Second Language Learner.
We See in the Dark
Authors: Sorensen Roy 
Published: August 2004

AbstractDo we need light to see? I argue that the black experience of a man in a perfectly dark cave is a representation of an absence of light, not an absence of representation. There is certainly a difference between his perceptual knowledge and that of his blind companion. Only the sighted man can tell whether the cave is dark just by looking. But perhaps he is merely inferring darkness from his failure to see. To get an unambiguous answer, I switch the focus from perceptual knowledge to non‐epistemic seeing. My conclusion is that we see even in the limiting case of absolute darkness – regardless of whether we believe we are seeing. We see little of pratical interest. But in terms of basic information, we see about as much as we do when the lights are on. Depending on what has gone before and after, we may even see ordinary objects.
Presence of immunoglobulin M antibodies around the glomerular capillaries and in the mesangium of normal and passive Heymann nephritis rats
Authors: Barabas Arpad Z. , Cole Chad D. , Barabas Arpad D. , Cowan Jord M. , Yoon Chang Soon , Waisman David M. , Lafreniere Rene 
Published: August 2004

SummaryDiffuse distribution of small, faintly staining, beaded deposits of rat immunoglobulin M (IgM) around the glomerular capillary blood vessels, and a more intensely staining larger deposition in the mesangium, were observed on the kidney sections of normal rats. As glomerular‐fixed nephritogenic antigens are known to be present on the epithelial aspect of the glomerular basement membrane (GBM), especially at the soles of foot processes and at the slit pores, it was assumed that the IgM antibodies were directed against these antigens. Investigation by immunofluorescent antibody double‐staining techniques of rat kidney sections obtained from normal and rabbit anti‐FX1A‐injected rats stained for the nephritogenic antigen showed that a number of antigenic sites in the glomeruli and in the mesangium shared antibody hits by heterologous rabbit IgG and autologous rat IgM antibodies. Most sites in the glomeruli stained specifically for rat IgM or rabbit IgG, but preferentially for the latter. The intensely fluorescent mesangial deposits stained mainly for rat IgM, indicating that at these sites the antigenic material was virtually saturated, while areas at the entry to the mesangial space also stained for rabbit IgG, indicating that at these locations free nephritogenic epitopes were still available for reaction with the anti‐FX1A antibody. Western blot analysis have shown that the rabbit anti‐rat FX1A IgG and the rat anti‐rat KF3 IgM antibodies are directed against the same renal tubular‐derived antigen with a molecular weight of 70,000. These experimental findings collectively demonstrate that the heterologous IgG and autologous IgM antibodies are directed against the same nephritogenic antigen, which is found in the glomeruli, the mesangium and the proximal convoluted tubules. Thus, the IgM autoantibody has a possible physiological role but, in addition, there is evidence of active immunophagocytic events, manifested in a rapid and continuous entrapment and expulsion of macromolecules after their processing by the mesangial cells of normal and passive Heymann nephritis rats.
Conformations of cell surface oligosaccharides and recognition by lectins from pathogens
Authors: Imberty Anne 
Published: August 2004

Pseudomonas aeruginosa is essentially a saprophytic bacterium that can turn into an aggressive pathogen when in contact with damaged and inflamed tissues such as burns and lungs of cystic fibrosis (CF) patients. In all of these cases, the host cell surfaces present altered glycosylations. The most studied example is that of CF where point mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, results in altered ion movements and also influences the N‐glycosylation of the CFTR protein and other cell surface glycoproteins. Increased fucosylation and decreased sialylation with higher levels of Lewis x and Lewis a epitopes correlate to P. aeruginosa‐specific binding. In addition, both the mucus covering the airway epithelia of CF patients and their salivary mucins display higher levels of sialylated and sulfated Lewis oligosaccharides that attract P. aeruginosa.The galactose‐ and fucose‐binding (PA‐IL and PA‐IIL) lectins of P. aeruginosa contribute to the virulence of this pathogenic bacterium, which is a major cause of morbidity and mortality in CF patients. Database searching in newly sequenced genomes indicates that the presence of these lectins is characteristics of high environmental adaptability of some opportunistic bacteria. The crystal structures of PA‐IL and PA‐IIL lectins have been solved in complex with the corresponding monosaccharides. Molecular modelling studies were used in order to predict the binding mode of PA‐IIL with complex oligosaccharides. Combination of theoretical studies with binding assays give clear indication that the histo‐blood group epitope Lewis a could be the biological ligand present in the lung of CF patients.
Phage display‐derived antibodies: new tools to study heparan sulfate diversity
Authors: Van Kuppevelt Toin 
Published: August 2004

Heparan sulfates comprise a heterogeneous group of polysaccharides involved in many basic cellular phenomena including proliferation, migration and differentiation. Their biosynthesis allows for the formation of a large number of different structures. Specific HS structures are likely involved in the binding of HS to proteins, such as growth factors. Anti‐HS antibodies, obvious tools to study structural heterogeneity, are difficult to obtain due to the poor immunogenicity of HS. To circumvent this, we have adopted antibody phage display technology, because this system allows for the generation of antibodies against self‐antigens.A human semisynthetic antibody phage display library (Nissim et al. 1994) and a semisynthetic library in which the CDR 3 region of the antibodies contained an XBBXBX amino acid sequence (a consensus heparin‐binding site) were used to select antibodies to HS using biopanning strategies.A large number of antibodies were selected, sequenced and characterized. A number of chemical groups in HS, important for antibody recognition, were identified. Antibodies revealed a large and regulated domain diversity of HS in a number of tissues (Dennissen et al. 2002). The up/down‐regulation of specific HS structures in various pathologies including cancer and nephropathies were identified (Jenniskens et al. 2003; Smetsers et al. 2003). Transfection of cells with cDNAs encoding the antibodies could be used to block HS in vitro and to study its physiological relevance.It is concluded that phage display‐derived antibodies to HS may provide new tools to study the role of HS in health and disease.
Modulation of heparan sulfate/chemokine interactions using RNA interference
Authors: Arzoumanian Hrant , Bennet Terry , Watson Zara , Carter Noel 
Published: August 2004

Introduction  The glycosaminoglycan heparan sulfate (HS) on the luminal surface of the vascular endothelium plays a vital role in promoting leucocyte migration during inflammation via specific interaction chemokines. HS is modified by sulfation and this appears to regulate chemokine binding. We previously observed that under pro‐inflammatory conditions [tumour necrosis factor α (TNF‐α) and interferon‐γ (IFN‐γ)], the endothelial cell line HMEC‐1 showed an increased sulfation HS. This promoted a significant increase binding of the chemokine RANTES and transendothelial migration of leucocytes. TNF‐α and IFN‐γ clearly modulated HS biosynthesis, in part by up‐regulating the NDST 1 and 2 genes responsible for N‐sulfation of the glycan, resulting in structural changes that favour HS/chemokine interactions (Carter et al. 2003). We have attempted to down‐regulate the expression of the gene NDST1 by RNA interference in order to investigate the structural and functional effects of reduced HS N‐sulfation.Materials and methods  A double‐stranded interference RNA was designed and synthesized in vitro to target the sulfotransferase NDST‐1 and used to tranfect the HMEC‐1 cell line. A number of assays measured its effect on HS structure and function. Cell surface N‐sulfation was measured by flow cytometry using the antibody F58 10E4 (Seikagaku, Japan). Cell surface binding of the chemokine MCP‐1 was visualized by immunohistochemistry. The functional effect of MCP‐1 binding was measured using a trans‐endothelial migration assay.Results  Surprisingly, the transfected cells appeared to show a small but marked increase in cell surface N‐sulfation. This was consistent with previous results transfecting a full‐length NDST‐1 antisense construct into the cell line, A549 (unpublished). The increase in N‐sulfation observed was supported functional date. Namely, transfected cells show an increased binding of the chemokine MCP‐1 and this promoted a statistically significant increase in transendothelial migration of leucocytes.Discussion  The results observed were contrary to expectations. It would appear that the interference RNA molecule is in some way interfering in the regulation of the NDST‐1 expression leading to an increase not decrease in gene expression. This has yet to be verified but we are beginning real time expression studies to validate this. We have hypothesized two possible reasons for this observation; endothelial cells may be able to sense a reduction in NDST activity and up‐regulate gene expression. Or the presence of the interference RNA itself promotes NDST‐1 expression. The latter is supported by growing evidence that short double‐stranded RNAs are thought to up‐regulate the pro‐inflammatory STAT transcription factor family (Shuey et al. 2002).
Identification of major heparin‐binding proteins in plasma using electrophoresis and mass spectrometry
Authors: Killeen Ruth , Wait Robin , Begum Shajna , Gray Elaine , Mulloy Barbara 
Published: August 2004

Introduction  The anticoagulant glycosaminoglycan heparin binds to the plasma protein antithrombin, potentiating its activity and so inhibiting the coagulation cascade. Among other therapeutic applications, it is used to prevent and treat deep vein thrombosis. As heparin binding is an attribute shared by many proteins other than antithrombin, sequestration by plasma proteins could potentially reduce the available concentration of therapeutically administered heparin, and thus modulates its biological activity. Although the heparin‐binding ability of some abundant plasma proteins is well characterized, the binding potential of the vast majority of plasma constituents is unknown. We have therefore initiated a systematic study of the heparin‐binding plasma proteome, using affinity capture on immobilized heparin, followed by salt elution and protein identification by mass spectrometry.Materials and methods  Five platelet‐depleted human plasma pools were fractionated by affinity chromatography on HiTrap heparin columns. Bound material was eluted stepwise with 0.5 m, 0.7 m, 1.2 m and 2.2 m NaCl. The approximate protein concentration of each plasma pool and fraction was determined by optical density reading at 280 nm. The fractions were separated by SDS‐PAGE and protein bands were visualized by staining with Coomassie blue or silver nitrate. Stained bands were excised, digested in gel with trypsin, and the liberated peptides were analysed by tandem electrospray mass spectrometry using a Micromass Q‐Tof. Proteins were identified by correlation of the uninterpreted tandem mass spectra to entries in the UniProt database.Results  Over 50 proteins were identified in whole plasma. No systematic differences in protein composition were observed between the different plasma pools. Heparin‐binding proteins recovered in salt washes of 0.7 m or above included fibronectin, histidine‐rich glycoprotein, apolipoproteins, complement factors C3 and C4b, and antithrombin III. Antithrombin was the major protein eluting at 1.2 m NaCl or above.Discussion  The rationale of this study was to investigate plasma for the presence of proteins potentially able to compete with antithrombin for therapeutically administered heparin. Of over 50 different plasma proteins identified in this preliminary study, only antithrombin remained bound to heparin above 1.2 m NaCl. We anticipate that use of larger volumes of albumin‐ and immunoglobulin‐depleted plasma will reveal low concentrations of additional high‐affinity binding proteins, but these are unlikely to bind significant amounts of heparin. However, in the present study, we detected several abundant proteins, which bound to immobilized heparin at 0.7 m NaCl, and these may reduce the bioavailability of heparin by nonspecific binding.
Heparan sulfate is essential for neuron migration and axon outgrowth in Caenorhabditis elegans
Authors: Kinnunen Tarja , Townsend Joanne , Turnbull Jeremy 
Published: August 2004

Introduction  The importance of heparan sulfate proteoglycans (HSPGs) has been highlighted by a number of human genetic disorders associated with mutations in genes encoding for HSPG protein cores or biosynthetic enzymes required for heparan sulfate assembly.Materials and methods  We have studied the role of HS and HSPGs in the nervous system development using the nematode Caenorhabditis elegans as a model organism. Our focus has been on the single C. elegans homologues of the vertebrate syndecans, sdn‐1 and heparan 2‐O‐sulfotransferase, hst‐2.Results and discussion  C. elegans syndecan, sdn‐1 and heparan 2‐O‐sulfotransferase, hst‐2, expressions coincide with the start of morphogenesis in mid‐embryonic stages after most cell divisions have occurred. Sdn‐1 is expressed primarily in neuronal cells, whereas hst‐2 is more widely expressed in neurons, hypodermis (epidermis), pharynx and the gonad leader cells (DTC). A deletion mutant of sdn‐1 which lacks the putative heparan sulfate (HS) attachment sites has specific neuron migration and axon pathfinding defects. Hst‐2 mutants have partially overlapping yet diverse neuronal migration defects. In both sdn‐1 and hst‐2 mutants, the serotonergic hermaphrodite‐specific neurons (HSNs) fail to migrate to their correct position in the vicinity of the vulva and hence fail to innervate the vulval muscles causing sdn‐1 and hst‐2 mutants to become egg‐laying defective (egl) and accumulate embryos within the mother. Canal‐associated neurons (CANs) that, like the HSNs, have a stereotyped long‐distance postembryonic migration pattern also fail to migrate to their correct position in sdn‐1 mutants, whereas in hst‐2 mutants the CANs migrate normally. These results suggest that specific 2‐O‐sulfation of sdn‐1 HS is essential for HSN migration, whereas it is dispensable for CAN migration.Hst‐2 mutants have additional cell migration defects that are not detected in sdn‐1 mutants suggesting that other HSPGs such as perlecan/unc‐52 and glypican/gpn‐1 may be responsible for these defects.
Studying the role of heparin in the formation of FGF1‐FGFR2 complexes using gel chromatography
Authors: Robinson Christopher J. , Harmer Nicholas J. , Blundell Tom L. , Gallagher John T. 
Published: August 2004

Introduction  The fibroblast growth factors (FGFs) are a family of at least 24 proteins which have diverse roles in regulating cell proliferation, migration and differentiation during embryogenesis, angiogenesis and tissue repair. Four FGF receptor tyrosine kinases (FGFRs) have been identified, each of which binds a specific subset of the FGF family. FGF binding induces dimerization of the FGFRs leading to their activation through transphosphorylation. Heparin or heparan sulfate (HS) is essential for the activity of the FGFs, because they form an integral part of the signalling complex on the cell surface. Our group has been using gel filtration FPLC to study the role that heparin plays in the formation of FGF1‐FGFR2 complexes.Methods  A Superdex 200 HR 10/30 FPLC column was calibrated using protein standards and used to estimate the size of protein complexes which form in the presence of heparin. The column was run in 150 mm NaCl, 50 mm HEPES, pH 7.2 buffer and complexes could be visualized by protein UV absorbance at 280 nm.Results  In agreement with published data (Pellegrini et al. 2000), we found that 2 : 2 : 1 complexes formed spontaneously in solution between FGF1, FGFR2IIIc and heparin decasaccharides (dp10). Although heparin dp6–8 fragments could bind FGF1 and FGFR2 simultaneously, only dp10 and larger fragments efficiently formed 2 : 2 : 1 complexes. These complexes appear to be relatively stable, as they remain associated following prolonged incubation with heparinase. FGF1 was able to bind heparin saccharides strongly even in the absence of receptor. In contrast, FGFR2 had very little affinity for either FGF1 or heparin dp12 separately, but bound efficiently to the FGF1‐heparin complex. Dimerization of FGF1 appears to be essential for subsequent dimerization of FGFR2. Selectively desulfated heparin saccharides could only bind FGF1 singly and formed 1 : 1 : 1 complexes with FGFR2, which did not then associate with one another.Discussion  Our data supports the idea that heparin plays a dual role in the formation of FGF1 signalling complexes, firstly by mediating interactions between FGF1 and FGFR2 and secondly by acting as a template for the dimerization of FGF1‐FGFR2 subunits. This technique could be easily adapted to study other heparin/HS‐mediated ligand–receptor interactions and could provide new insights into the mechanisms of formation of multiprotein signalling systems on the cell surface.
The analysis of the fibroblast growth factor ligand‐receptor complex using a quartz crystal microbalance‐dissipation
Authors: Rudd Tim , Duchesne Laurence , Fernig David G. , Nichols Richard J. 
Published: August 2004

Introduction  The quartz crystal microbalance‐dissipation (QCM‐D) is a device that measures both the mass of adsorbates upon a surface and the energy dissipated by the surface. The latter quantity, known as dissipation (D), gives information about the viscoelastic properties of species adsorbed to the surface leading to insights into their shape and conformation.Methods  The fibroblast growth factor (FGF) receptor‐ligand signalling complex is simulated within the QCM‐D by the immobilization of heparin‐derived oligosaccharides to a gold‐sputtered QCM‐D crystal surface, and then the surface is interrogated with various growth factors and a recombinant receptor. Fibroblast growth factor receptor 1 (FGFR1) extracellular domain (immunoglobulin loops II and III and the acid box of the FGFR1IIIc), FGF‐1, FGF‐2, FGF‐7 and hepatocyte growth factor/scatter factor (HGF/SF) all bind to immobilized heparin‐derived oligosaccharides with different changes in D relative to mass for the same surface.Results  For example, complexes of FGF‐1 and octasaccharide are, on a mass basis, far more dissipative than complexes of HGF/SF. One interpretation of these results is that octasaccharides are relatively flexible, or at least contain hinge regions and that different FGFs have a preference for binding above the hinge (so more dissipative) or on the hinge (so more rigid). Binding of FGFR1 produced complexes that were less dissipative than the FGF‐1 complexes. Intriguingly, deglycosylation of the FGFR1 increased the amount of binding to heparin tetrasaccharide, and this complex on a mass basis is more dissipative.Conclusion  By using the QCM‐D, we are able to gain insights into novel aspects of these protein‐glycosaminoglycan complexes.