bluuebunny/crossref_metadata_2025 huggingface dataset:
After the War Comes Peace? An Examination of the Impact of the Northern Ireland Conflict on Young People
Authors: Gallagher Tony
Published: August 2004

When the conflict in Northern Ireland erupted over 30 years ago many voiced concern at the implications for children and young people. In the early years many looked to U.S. experience in order to frame questions, problems, and solutions. This article examines the strands of psychological research that developed and their link to the articles in the present collection. The article critiques the consensus among psychologists in the 1970s and 1980s that young people coped reasonably well in difficult circumstances. One theme of the current collection is to highlight the difficulties of dealing with the legacies of the past thirty years and hence the constraints facing the attempt to establish a peaceful and settled society.
Volunteerism and Social Problems: Making Things Better or Worse?
Authors: Penner Louis A.
Published: August 2004

Volunteerism is described and defined and then a model of the decision to volunteer is presented. Data from an archival analysis of volunteering after the September 11, 2001 attacks and an on‐line survey of volunteers are presented in support of the model. Finally, the implications of increasing volunteerism for the solution of social problems are considered.
The Competitive Advantage of Foreign Languages and Cultural Knowledge
Authors: Grosse Christine Uber
Published: August 2004

This study presents the results of an electronic survey of 2,500 randomly selected alumni from the graduating classes of 1970 through 2002 of Thunderbird, The American Graduate School of International Management. Of the 2,500 alumni, 581 responded. Thunderbird required a minimum of 4 semesters of foreign language for graduation. The survey concerned whether or not the alumni had received a competitive advantage in their careers from their foreign language skills and cultural knowledge. The vast majority of the respondents acknowledged that both foreign language skills and cultural knowledge had benefited them in their professional lives. Slightly more of them reported receiving an edge from their cultural skills (89%) than from their foreign language abilities (82%). The research results provide empirical data on how members of the U.S. and international business communities perceive the value of foreign language and cultural knowledge to their work.
Computer Anxiety and Students' Preferred Feedback Methods in EFL Writing
Authors: Matsumura Shoichi , Hann George
Published: August 2004

Computer‐mediated instruction plays a significant role in foreign language education. The incorporation of computer technology into the classroom has also been accompanied by an increasing number of students who experience anxiety when interacting with computers. This study examined the effects of computer anxiety on students' choice of feedback methods and academic performance in English as a foreign language (EFL) writing. The study included 207 university‐level Japanese students in EFL writing classes, who received both face‐to‐face teacher feedback and online teacher and peer feedback while revising an essay writing assignment. The students were free to choose their preferred feedback method. The results of multiple regression analysis revealed that the students' choices of feedback method varied as a function of the level of their computer anxiety and that providing the choice of using or not using computers helped both high‐ and low‐anxiety students improve their essay writing. The findings reveal the importance of recognizing computer anxiety and creating a learning environment in which students who are highly computer anxious are not disadvantaged.
Tiny Publics: Small Groups and Civil Society
Authors: Fine Gary Alan , Harrington Brooke
Published: August 2004

It has been conventional to conceptualize civic life through one of two core images: the citizen as lone individualist or the citizen as joiner. Drawing on analyses of the historical development of the public sphere, we propose an alternative analytical framework for civic engagement based on small-group interaction. By embracing this micro-level approach, we contribute to the debate on civil society in three ways. By emphasizing local interaction contexts—the microfoundations of civil society—we treat small groups as a cause, context, and consequence of civic engagement. First, through framing and motivating, groups encourage individuals to participate in public discourse and civic projects. Second, they provide the place and support for that involvement. Third, civic engagement feeds back into the creation of additional groups. A small-groups perspective suggests how civil society can thrive even if formal and institutional associations decline. Instead of indicating a decline in civil society a proliferation of small groups represents a healthy development in democratic societies creating cross-cutting networks of affiliation.
The Tripartite Structure of Social Science Analysis
Authors: Jasso Guillermina
Published: August 2004

The goal of sociology, and all social science, is to produce reliable knowledge about human behavioral and social phenomena. To reach that goal, we undertake three kinds of activities: theoretical work, empirical work, and, even more basic, we develop frameworks that assemble the fundamental questions together with the fundamental tools that will be used to address them. This article examines the three sets of activities and their interrelations. Both deductive and nondeductive theory are highlighted, as are three kinds of empirical work—testing the predictions of deductive theories, testing the propositions produced by nondeductive theories, and extratheoretical measurement and estimation. Illustrations are drawn from the fields of status, justice, and migration.
Sociology and the Second Darwinian Revolution: A Metatheoretical Analysis
Authors: Machalek Richard , Martin Michael W.
Published: August 2004

Sociologists tend to eschew biological explanations of human social behavior. Accordingly, when evolutionary biologists began to apply neo-Darwinian theory to the study of human social behavior, the reactions of sociologists typically ranged from indifference to overt hostility. Since the mid-1960s, however, neo-Darwinian evolutionary theory has stimulated a “second Darwinian revolution” in traditional social scientific conceptions of human nature and social behavior, even while most sociologists remain largely uninformed about neo-Darwinian theory and research. This article traces sociology's long-standing isolation from the life sciences, especially evolutionary biology, to divergence in the metatheoretical assumptions that typify conventional sociological thought versus contemporary evolutionary biology. We conclude with a discussion of the recent emergence of a nascent “evolutionary sociology” that integrates sociobiological reasoning with contemporary sociological thought.
Illusory Causation: Why It Occurs
Authors: Lassiter G. Daniel , Geers Andrew L. , Munhall Patrick J. , Ploutz-Snyder Robert J. , Breitenbecher David L.
Published: August 2004

Considerable evidence indicates that people overattribute causality to a given stimulus when it is salient or the focus of their attention—the so-called illusory-causation phenomenon. Although illusory causation has proved to be quite robust and generalizable, a compelling explanation for it has not been empirically documented. Four social-attribution studies were conducted to test the hypothesis that illusory causation occurs because salient information is initially registered, or perceptually organized, differently than nonsalient information. The results provide considerable support for the notion that people's literal point of view affects how they initially perceive, or extract, information from an observed interaction, which in turn affects their judgments regarding the causal influence exerted by each interactant.
Herpes simplex virus‐based vectors
Authors: Lachmann Robin
Published: August 2004

Herpes simplex virus (HSV)‐based vectors have primarily been developed for neuronal gene delivery, taking advantage of the virus' natural neurotropism. Two types of vector are available: replication defective viruses, whose cytotoxicity has been abolished by deleting viral gene products, and amplicon vectors, which are plasmids packaged into HSV particles with the aid of a helper virus. In this review I discuss how the cytotoxicity of the wild‐type virus has been abolished, the progress which has been made toward defining promoter elements capable of directing long‐term transgene expression form the latent viral genome and some of the potential clinical uses of these versatile vectors.
Bone marrow cells carrying the env‐pX transgene play a role in the severity but not prolongation of arthritis in human T‐cell leukaemia virus type‐I transgenic rats: a possible role of articular tissues carrying the transgene in the prolongation of arthritis
Authors: Abe Asami , Ishizu Akihiro , Ikeda Hitoshi , Hayase Hiroko , Tsuji Takahiro , Miyatake Yukiko , Tsuji Muneharu , Fugo Kazunori , Sugaya Toshiaki , Higuchi Masato , Matsuno Takeo , Yoshiki Takashi
Published: August 2004

Transgenic rats carrying the env‐pX gene of human T‐cell leukaemia virus type‐I (env‐pX rats) were immunized with type II collagen (CII), and chronological alterations of arthritis were compared with findings of collagen‐induced arthritis (CIA) in wildtype Wistar–King–Aptekman–Hokudai (WKAH) rats. Arthritis induced by CII in env‐pX rats was more severe and persisted longer than CIA in WKAH rats. To determine whether the phenomenon is caused mainly by the transgene‐carrying lymphocytes or articular tissues, we immunized lethally irradiated env‐pX and WKAH rats with reciprocal bone marrow cell (BMC) transplantation. A severe but transient arthritis was induced by CII in WKAH rats reconstituted by env‐pX BMC (w/tB/CII rats). On the other hand, in env‐pX rats reconstituted by WKAH BMC, arthritis persisted longer than in w/tB/CII rats, although the degree was less at an early phase after CII immunization. These findings suggest that articular tissues rather than the BMCs carrying the env‐pX transgene play a role in the prolongation of arthritis in env‐pX rats, although BMCs carrying the transgene are associated with the severity of arthritis. When inflammatory cytokines in synovial cells isolated from env‐pX rats before they developed arthritis were examined, interleukin‐6 (IL‐6) was detected at a higher level than in synovial cells from WKAH rats, thus suggesting the critical role of IL‐6 in env‐pX arthritis.
Immunohistochemical analysis of heme oxygenase‐1 in preneoplastic and neoplastic lesions during chemical hepatocarcinogenesis§
Authors: Caballero Fabiana , Meiss Roberto , Gimenez Alejandra , Batlle Alcira , Vazquez Elba
Published: August 2004

Heme oxygenase (HO) breaks down the pro‐oxidant heme into carbon monoxide, iron and the antioxidant biliverdin. The isoform HO‐1 plays an effective role to counteract oxidative damage and to control inflammation. Prolonged cellular damage due to chronic inflammation is one of the reasons leading to the development of tumours. The aim of this work was to investigate HO‐1 expression and localization along the different stages of chemically induced hepatocarcinogenesis (HCC) and the occurring morphological changes. To provoke sustained oxidative stress and chronic inflammation, CF1 mice received dietary p‐dimethylaminoazobenzene (DAB, 0.5%, w/w) during a whole period of 14 months. HO‐1 expression increased along the experimental trial in morphologically normal hepatocytes in DAB‐treated animals. HO‐1 expression diminished in altered hepatic foci (AHF) and oval cells and early preneoplastic lesions. Otherwise, marked HO‐1 overexpression was detected in Kupffer cells and macrophages surrounding necrotic and nodular areas. Adenomas showed decreased HO‐1 immunostaining. In hepatocellular carcinomas, an inverse relationship was found between the immunohistochemical expression of HO‐1 and the degree of tumour differentiation, being negative in poorly differentiated tumours. In our experimental model, down modulation of HO‐1 expression correlated with malignancy progression. Thus, our data point to activation of HO‐1 as a potential therapeutic tool.
Regulation of signal transduction by glycosylation
Authors: Haltiwanger Robert S.
Published: August 2004

The incredible diversity found in cell‐surface glycoconjugate structures led researchers over 30 years ago to propose that complexity in carbohydrates must play a role in cellular communication. Recent studies from a number of laboratories have confirmed this hypothesis, demonstrating that cell‐surface glycoconjugates play significant roles in signal transduction events. One striking example is the effect of O‐fucose modifications on the Notch‐signalling pathway. Notch is a cell‐surface receptor that is essential for proper development. The extracellular domain of Notch contains up to 36‐tandem epidermal growth factor‐like (EGF) repeats, many of which are predicted to be modified at putative consensus sequences with O‐fucose and O‐glucose saccharides. Genetic alterations (by knockout or RNAi methodologies) in the enzyme responsible for the addition of O‐fucose to Notch, protein O‐fucosyltransferase‐1, result in severe, embryonic lethal phenotypes resembling Notch mutants. Thus, O‐fucosylation appears to be essential for proper Notch function. Elongation of the O‐fucose monosaccharide by the β1,3‐N‐acetylglucosaminyltransferase, Fringe, modulates Notch function, either increasing or decreasing response from ligands depending on context. Although it is now clear that O‐fucose modifications affect Notch signalling, the molecular mechanism by which this occurs is not known. As an initial step in understanding how O‐fucose glycans affect Notch function, we are mapping O‐fucose modifications to specific sites on Notch. Already, we have demonstrated that O‐fucose modifies one of the EGF repeats involved in ligand binding, suggesting that the sugars may play a role in Notch–ligand interactions. Experiments to test the role of O‐fucose modifications at specific sites are in progress. We have also found that Fringe modifies O‐fucose on some EGF repeats but not others. Our initial analyses suggest that the basis of this specificity is encoded within the sequences of the individual EGF repeats. Site mapping has also confirmed the presence of O‐glucose saccharides on Notch. The evolutionarily conserved, predicted O‐glucose sites on Notch are as numerous as those for O‐fucose, suggesting that the O‐glucose modifications will play an equally important role in Notch biology. We have recently identified an enzymatic activity capable of catalyzing the addition of O‐glucose to EGF repeats. Purification of the protein O‐glucosyltransferase is underway. These and other results will be discussed.
MS strategies for high throughput glycomics and glyco‐proteomics
Authors: Dell Anne , Sutton‐Smith Mark , Panico Maria , Chalabi Sara , Wong Nyet‐Kui , Hitchen Paul , Jang‐Lee Jihye , North Simon , Ledger Victoria , Parry Simon , Haslam Stuart , Morris Howard
Published: August 2004

Ultra‐high sensitivity mass spectrometric strategies for defining the primary structures of highly complex mixtures of glycopolymers are revolutionizing structural glycobiology in the postgenomic era. MS strategies incorporating MALDI‐MS and ES‐MS/MS enable very complex mixtures from biological extracts and/or glycopolymer digests to be screened, thereby revealing the types of glycans present and, importantly, providing clues to structures that are likely to be functionally important.

This lecture will overview strategies employed in our laboratory which enable the glycome of cells, tissues and organs to be examined and the glycoforms of individual glycoproteins to be identified. The types of structural problems that can be solved with current technology will be exemplified by some of our recent research in the following areas:

Development and application of glycomics screening strategies for examining glycosylation changes in cells and tissues of knockout mice.

Defining changes in glycosylation of murine T and B cells upon cytokine activation.

Characterization of the ovarian cancer antigen, CA125.

Sequencing and site analysis of N‐ and O‐glycans in murine ZP3, the putative sperm receptor.

Characterization of mucin glycosylation in cystic fibrosis‐related research.

Glyco‐proteomic investigations of changes in O‐glycosylation of CD8 during T‐cell development.

Glyco‐proteomic analyses of novel glycosylation of bacterial proteins.
Hyaluronan: a simple polysaccharide with structural plasticity and functional diversity
Authors: Day Anthony J.
Published: August 2004

Hyaluronan (HA) is a high‐molecular weight glycosaminoglycan that is involved in an extracellular matrix (ECM) organization and cell adhesion, essential to a wide range of normal physiological processes (e.g. development, immunology and reproduction). Its diverse biological activities may seem surprising for a simple linear polysaccharide composed solely of a repeating disaccharide of glucuronic acid and N‐acetylglucosamine. However, HA is likely to be able to take up many different conformational states, which are transient and rapidly interchanging in solution (Day & Sheehan 2001). It has been suggested that the interaction of HA with different HA‐binding proteins (hyaladherins) may stabilize particular conformations of the polysaccharide leading to HA‐protein complexes with distinct architectures and unique biological properties (Day & Sheehan 2001). Consistent with this hypothesis, recent studies on TSG‐6 (Blundell et al. 2003) and CD44 (Teriete et al. 2004) and molecular modelling of other members of the Link module superfamily reveal significant structural diversity in the HA‐binding domains in these related hyaladherins. Furthermore, there is evidence emerging that the complexes formed between polymeric HA and proteins can exhibit a wide range of higher order structures depending on the hyaladherin(s) involved. In some cases, these complexes may play mainly a structural role in ECM (e.g. the link protein/aggrecan/HA aggregates that provide load bearing function to cartilage), while in other cases they act as cell binding/activation sites (e.g. HA cables formed during inflammation). There is evidence that the function of HA can be modulated by its interaction with proteins. For example, preincubation of HA with TSG‐6 enhances/induces the binding of HA to cell surface CD44 on constitutive/inducible lymphocyte backgrounds (Lesley et al.); these activities are probably caused by the formation of cross‐linked HA fibrils that lead to receptor clustering. HA can also become modified by the covalent attachment of heavy chains (HCs), derived from the interα‐inhibitor, where TSG‐6 plays an essential role in this process (Mukhopadhyay et al.); this reaction is critical for the formation of a HA‐rich ECM required for successful ovulation and fertilization. HC‐HA complexes are also formed during inflammation but their function is not yet clear.
Biosynthesis of heparan sulfate – how regulated does it need to be?
Authors: Lindahl Ulf
Published: August 2004

Heparan sulfate proteoglycans (HSPGs) are ubiquitous in animals. The functional significance of the structural variability exhibited by the constituent HS chains remains a fundamental question in cell and developmental biology. The polysaccharides influence cell behaviour, by modulating the activities of signalling proteins or through interactions in the extracellular matrix. HS is synthesized as a linear polymer of alternating glucuronate (GlcA) and N‐acetylglucosamine (GlcNAc) units. The resultant [GlcA‐GlcNAc]n polymer is partially modified through a series of N‐deacetylation/N‐sulfation, GlcA C5‐epimerization and O‐sulfation reactions (Esko & Lindahl 2001) and may be further modified by selective removal of O‐sulfate groups (Ai et al. 2003). Owing to the variable extent of these reactions in different cells and tissues, and at different developmental stages, the mature HS products display different structures, typically expressed through sulfated domains of variable length and composition (Esko & Lindahl 2001; Casu & Lindahl 2001). These domains provide binding sites for protein ligands and thus constitute the structural basis for the diverse functional roles of HS. The regulated diversity of HS structure is demonstrated through compositional analysis of HS from various organs as well as by immunohistochemical recognition of HS epitopes (van Kuppevelt, T. H et al. 1998). So far, a precise structure–function relation has been defined only for the antithrombin‐binding pentasaccharide sequence (Casu & Lindahl 2001).

HS is essential throughout development, as mouse embryos devoid of HS‐polymerizing enzymes fail in gastrulation (Gallagher 2001). By contrast, targeted disruption of genes encoding polymer‐modifying enzymes is associated with surprisingly mild phenotypes. Embryos lacking N‐deacetylase/N‐sulfotransferase‐1, 2‐O‐sulfotransferase (Gallagher 2001) or GlcA C5‐epimerase (Li et al. 2003), thus show major defects incompatible with postnatal life, but also apparently normal organ systems known to rely on HS‐binding morphogens or growth factors for their development. Moreover, transgenic mice overexpressing heparanase appear essentially normal (Zcharia et al. 2004), in spite of an overall drastic reduction in HS chain size. How stringent are structure/function relations in HS–protein interactions?
An ultrastructural investigation of dermatopontin‐knockout mouse corneas
Authors: Cooper L.J. , Bentley A.J. , Nieduszynski I.A. , Fullwood N.J. , Ellis T.S. , Thomson A. , Utani1 A. , Sinkai H. , Brown G.M.
Published: August 2004

Introduction  The corneal stroma is composed of a network of heterotypic collagen fibrils, proteoglycans and matrix proteins. Transparency of the tissue principally requires the uniformity of fibril diameters and interfibrillar distances and the presence of a quasi‐hexagonal lattice arrangement of parallel fibrils. Keratan sulfate proteoglycans (KSPGs) have a crucial role and the KS chains are clearly required for the maintenance of transparency. Undersulfation of corneal KS results in tissue opacity and the lumican (a KSPG) knockout mouse shows corneal opacity and the disruption of collagen fibril diameters and interfibrillar distances (Chakravarti et al. 1998). Biochemical analysis has shown that dermatopontin is an abundant component of the extracellular matrix and that it interacts with KSPGs via the KS chains. This study aims to determine whether dermatopontin has a direct role in corneal matrix organization by investigating the corneal ultrastructure of dermatopontin‐null (dpt–/–) mouse corneas.

Materials and methods  Conventional light microscopy was used to compare the corneal thickness of dpt–/– mice (Takeda et al. 2002) with that of the wild‐type. Collagen fibril distribution was studied using transmission electron microscopy and the datasets analysed using SIS‐pro image analysis software to determine fibrillar volume, shape factor, fibril diameter and spacing.

Results  Light microscopy demonstrated that dpt–/– corneas show a 24% reduction in average stromal thickness compared to wild‐type (P < 0.001). The epithelium and Descemet's membrane appear normal. Examination of dpt–/– stroma by transmission electron microscopy indicates a significant disruption to lamellar organization in the posterior region while the central and anterior regions appear largely unaffected compared to wild‐type. The collagen fibrils in dpt–/– stroma show a pronounced increase in interfibrillar spacing as well as exhibiting a lower fibril volume fraction. There is no apparent difference in fibril diameter between dpt–/– and wild‐type mice.

Discussion  Collectively, these data suggest that dermatopontin plays a key role in collagen fibril organization and deposition. Like the cornea from lumican‐knockout mice (Chakravarti et al. 1998), the defects in collagen organization in dpt–/– cornea appear to be most severe in the posterior stroma. It is likely that dermatopontin interacts with the KS chains on lumican and this interaction is involved in the maintenance of stromal architecture.
PVA combined with 0.8% alginate: a potential 3‐dimensional vehicle to deliver and retain cells during autologous repair of connective tissues?
Authors: Gargiulo B. , Menage J. , Evans H. , Urban J.P.G. , Caterson B. , Curtis C. , Eisenstein S.M. , Roberts S.
Published: August 2004

Introduction  Autologous chondrocyte implantation (ACI) is routinely used for the repair of articular cartilage defects. A similar method may be employed to treat degenerate intervertebral discs or other connective tissues. A system in which cells could not only be delivered, but also retained would offer advantages compared to ACI. Such a vehicle would also allow a homogeneous distribution of cells throughout the defect and enhance nutrient penetration to the seeded cells.

Materials and methods  Bovine nucleus cells were isolated via enzyme digestion and expanded in number to passage 3. The cells were resuspended in 0.8% alginate and loaded into polyvinyl alcohol (PVA) cubes. These constructs were placed into a solution of calcium chloride to ‘gel’ the alginate. Constructs were cultured in DMEM + 10% FBS within 15‐ml conical tubes rotated at 37 °C for up to 28 days. Cell distribution/morphology and proliferation were assessed on H&E and Ki‐67 stained sections, respectively. The re‐expression of a disc cell phenotype was assessed using toluidine blue staining and immunohistochemistry (with antibodies to collagen types I, II, IIA, VI and X and to the glycosaminoglycans, chondroitin‐4‐ and ‐6‐sulfate and keratan sulfate. RT‐PCR was performed using oligonucleotide primers to collagen types I, II and X, aggrecan, link protein and small leucine‐rich PGs.

Results  H&E staining of 10‐µm thick cryosections revealed an even distribution of loaded cells throughout the scaffold at day 1 being maintained through to day 28. Toluidine blue staining showed the presence of GAGs, increasing with time. Ki‐67 staining indicated that approximately 5% of cells were proliferating at all time points. Immunohistochemistry demonstrated the production of collagen types I, II, IIA, VI and X, and the glycosaminoglycans, chondroitin‐4‐, ‐6‐sulfates and keratan sulfate. RT‐PCR results showed mRNA expression of fibromodulin throughout the experiment, lumican at days 14, 21 and 28. Types II and X collagen were present at days 21 and 28.

Discussion  Combining 0.8% alginate with PVA retained 100% of the seeded cells and allowed an even distribution of cells throughout the scaffold. The immunohistochemistry and RT‐PCR demonstrated that the system allowed the bovine nucleus cells to express phenotypic markers expressed by disc cells in vivo. These preliminary results indicate that the PVA/alginate system could act as a suitable delivery device for cells during autologous repair of the intervertebral disc or other connective tissues such as meniscus.
An interaction between opticin and heparan sulfate may provide the molecular basis for vitreoretinal adhesion
Authors: John Hindson V. , Gallagher John T. , Bishop Paul N.
Published: August 2004

Introduction  Opticin is a member of the extracellular matrix small leucine‐rich repeat (SLRP) proteoglycan/protein family, which was originally identified in the eye associated with the collagen fibrils of the vitreous humour. A putative heparin/heparan sulfate (HS) binding motif (RKERKRR) was identified at the N‐terminus of human opticin, but this is absent in the bovine form. Furthermore, the strength of attachment between the vitreous and the retina was observed to be species‐dependent and related to the presence or absence of this motif. We hypothesized that opticin cross‐links the collagen fibrils of the vitreous to HS proteoglycans in the inner limiting lamina (a basement membrane on the inner surface of the retina), contributing towards vitreoretinal adhesion.

Materials and methods  Recombinant human and bovine opticin were expressed in 293‐EBNA cells and purified to apparent homogeneity. Solid phase assays and surface plasmon resonance studies were used to characterize interactions between immobilized heparin/HS and opticin.

Results  Solid phase and BIAcore data revealed that human opticin binds heparin/HS and binds to heparin with a dissociation constant of approximately 20 nm. By contrast bovine opticin, which lacks the basic cluster, bound severalfold less tightly. Competition studies with heparin oligosaccharides indicated that the heparin/HS binding site is greater than 6 monosaccharides in length. Heparin, HS, chondroitin sulfate A (CS‐A), dermatan sulfate and hyaluronan all competed with heparin for binding to human opticin but CS‐C did not.

Discussion  Work to date suggests that the N‐terminal sequence RKERKRR contributes significantly to the binding of opticin to heparin/HS. Vitreoretinal adhesion plays a key role in a number of eye diseases and inhibitors of the opticin–HS interaction could be of therapeutic value.
ADAMTS‐4 activity on a proteoglycan vs. a polypeptide is controlled by the ancillary domains
Authors: Jones Gavin C. , Vankemmelbeke Mireille N. , Verheijen Jan H. , DeDood Claudia , Day Anthony J. , Buttle David J.
Published: August 2004

Introduction  In combination with the catalytic domain, the ancillary domains of the ADAMTS' are proposed to regulate activity via interactions with sulfated GAGs, the extracellular matrix (ECM) and cell surface. Interactions with both GAGs and the ECM have been attributed to the thrombospondin (TSP) type 1 motifs and spacer region (Kuno and Matsushima 1998; Flannery et al. 2002). ADAMTS‐1, ‐4 and ‐5, all undergo cleavage within their respective spacer regions (Flannery et al. 2002; Rodriguez‐Manzaneque et al. 2000; Georgiadis et al. 2002), an event which has been reported to increase activity towards the interglobular domain of aggrecan and decrease the heparin affinity of ADAMTS‐4 (Flannery et al. 2002; Gao et al. 2002).

Materials and methods  V5‐ and His‐tagged recombinant human ADAMTS‐4 constructs terminating after the catalytic (▵DTS), disintegrin‐like (▵TS), TSP (▵S) or spacer region (Full) were expressed in High‐Five cells. Proteoglycanase activities of the resultant proteins were assayed with solution digests of aggrecan and a polyacrylamide‐entrapped aggrecan particle assay. Proteolytic activity was measured using a novel, nonglycosylated, reporter substrate assay.

Results  All forms of ADAMTS‐4 were active to varying degrees in the reporter substrate assay. Digestion of aggrecan in solution digests was apparent in all proteins with the exception of the catalytic domain in isolation (▵DTS). Activity towards aggrecan decreased with increasing truncation of the protein.

Discussion  Removal of the cysteine‐rich‐spacer domain and further C‐terminal truncations decrease the activity of ADAMTS‐4 towards aggrecan, whilst the proteolytic activity remains intact. Cleavages releasing the ancillary domains of ADAMTS' may therefore alter the catalytic activity of these enzymes against proteoglycans and also nonglycosylated polypeptides. More information is required about potential substrates for the processed forms of ADAMTS‐4.
Conformation of glycosaminoglycans by ion mobility mass spectrometry
Authors: Jin Lan , Barran Perdita E. , Lyon Malcolm , Uhrín Dušan
Published: August 2004

Introduction  Glycosaminogycans (GAGs) are polysaccharides found on most animal cell surfaces and extracellular matrices. They constitute an important class of biologically active macromolecules that are implicated in the biological activity of morphogens, growth factors, cytokines, chemokines and enzymes. Intervention in signalling pathways they are involved in, represents a valuable therapeutical opportunity for cancer treatment, wound healing and specific organ regeneration. GAGs are linear polysaccharides with alternating uronic acid and hexosamine residues. Their polysaccharide precursors are extensively modified (epimerization, N‐deacetylation N‐, O‐sulfation) creating a wide heterogeneity to their structures. The binding specificity of GAGs is encoded in their primary structures but ultimately depends on how this basis set of functional groups is presented to a protein in three‐dimensional space.

Methods  As a part of our efforts to characterize the conformation of GAGs, we present here our preliminary results of conformational studies of heparin‐derived oligosaccharides by ion mobility mass spectrometry. This MS technique is designed to study conformation of mass‐selected gas‐phase ions. When ions drift through an inert gas under the influence of a weak electric field, their arrival time depends on their collisional cross sections. From this data, the experimental cross sectional areas are easily derived. These are then compared with those obtained from candidate geometries generated from molecular modelling, and hence conformation(s) of studied compounds can be determined.

Results and discussion  We have measured the cross sectional areas of four heparin oligosaccharides: one disaccharide and three tetrasaccharides. Amongst these are two fully sulfated species and also two tetrasaccharides each with one sulfate group missing. Our samples were prepared by enzymatic digestion of heparin and contain a nonreducing terminal unsaturated glycosyluronic acid ring. We employ the GLYCAM 2000 force field (Woods) and the AMBER (Case et al. 2002) suite of programs to model the gas‐phase conformations of the oligosaccharides. We have generated AMBER parameters for sulfate groups using ab initio calculations and are in the process of generating structures for which we calculate the cross sectional areas using a projection approximation (Wyttenbach et al. 1997). We present a comparison of experimental cross sections with those of calculated models. We also compare them with cross sections calculated using oligosaccharide fragments extracted from X‐ray and NMR structures of heparin oligosaccharides – protein complexes.
New insights into the alternating sequences of heparan sulfate
Authors: Murphy Kevin J. , Lyon Malcolm , Merry Catherine L. R. , Thompson James E. , Roberts Ian S. , Gallagher John T.
Published: August 2004

Introduction  The sulfated domains (S‐domains) within HS are thought to be the primary binding site for protein ligands (Gallagher and Lyon, 2000). Studies on the binding of multimeric chemokines to HS have suggested that N‐acetylated sequences (NAc‐domains) act to space the S‐domains correctly so binding can occur at more than one HS binding site within a protein (Lortat‐Jacob et al. 1995; Stringer et al. 2002). So far, little functional significance has been ascribed to either alternating N‐sulfate/NAc or lone N‐sulfate types of sequence, although alternating sequences are thought to occur at the borders between S‐domains and NAc‐domains, forming transition zones between the two. Existing scission techniques used to excise oligosacchrides from the intact chain for structural and functional analysis disrupt the transition zones (heparinase III), or the S‐domains (heparinase I) or both (low pH nitrous acid).

Materials and methods  Radiolabelled HS was isolated from cultured 3T3 fibroblasts HS. Its patterns of degradation using bacteriophage K5 lyase as well as conventional heparinases I/III and nitrous acid were compared. K5 lyase‐resistant fragments were characterized by size exclusion chromatography and strong‐anion exchange HPLC.

Results  The work presented demonstrates how the enzyme K5‐lyase specifically cleaves only within NAc‐domains requiring sequences of at least five consecutive N‐acetylated disaccharides within the HS chain. The enzyme leaves intact S‐domains together with their flanking transition zones in situ.

Discussion  The characterization of oligosaccharides generated by K5 lyase cleavage of the intact HS chain provides new insights into the domain structure of HS.
Site‐specific N‐linked glycosylation analysis on the human salivary mucin MUC5B using Precursor Ion Discovery on the CAPLC Q‐TOF system
Authors: Ritchie Mark , Knight David , Kirkham Sara , Thornton David
Published: August 2004

Introduction  Mucins are large highly O‐glycosylated oligomeric glycoproteins that form the polymer matrix of mucus, which forms part of the innate immune system. MUC5B, the predominant gel‐forming mucin in saliva, is around 600 kDa in mass before glycosylation. O‐linked glycosylation is abundant in defined repeated domains and may account for 80% of the observed mass of the secreted. Not much is known about the N‐linked glycosylation, although there are at least 34 potential sites. Despite little being known about the structure, N‐glycosylation has been shown to be important but not essential for mucin synthesis (Perez‐Vilar & Hill 1999). Here, we aim to study the N‐linked glycosylation site specifically using mass spectrometry

Materials and methods  A sample of purified human MUC5B mucin was digested with trypsin and separated by size exclusion chromatography (Thornton et al. 1999). The previously uncharacterized fractions were analysed by online LC Q‐Tof MS in order to characterize any N‐glycosylated peptides present. Glycopeptide discovery was steered by the Precursor Ion Discovery function, which detects potential glycopeptides by looking for sugar oxonium ions during intermittent high‐collision energy surveys. Candidate ions are then selected for MSMS, which reveals both the carbohydrate content and the glycosylation site location.

Results  Glycopeptides were located and site specifically analysed. Data suggests N‐linked occupancy of numerous sites, and microheterogeneity was observed.

Discussion  This is the first study to investigate the N‐glycans on oligomeric gel‐forming mucins. The N‐glycans are localized in the N‐ and C‐terminal portions of the MUC5B polypeptide in areas which have sparse O‐glycosylation. These regions of the molecule are known to be involved in the disulfide bond‐mediated oligomerization of the mucin monomers. Furthermore, they may be potential sites for interaction with other components of saliva that are important for biological function. It may be that they have a role in stabilizing these important regions of the molecule, or they may be sites of interaction with other salivary components.
Investigating the role of heparin sulfate proteoglycans in hereditary multiple exostoses (HME) tumourigenesis
Authors: Scholefield Z.M. , Hassell J.r , West L. , Govindraj P. , Hecht J.T. , Rushton G. , Oldershaw R. , Hardingham T. , Merry C.L.R. , Gallagher J.T.
Published: August 2004

Introduction  Heparin sulfate (HS) has long been implicated in the bone deformity hereditary multiple exostoses (HME), and it is now clear that HME is associated with mutations in the HS biosynthetic genes EXT1 and EXT2. Interestingly, HME is also associated with an increased risk of chondro‐ and osteo‐sarcomas.

Methods and results  Preliminary analysis of GAG samples purified from fibroblasts of both HME and isolated non‐HME exostoses patients reveal a dramatic shift in the ratio of CS : HS, with the HME and isolated cases having a much higher proportion of CS relative to normal controls. This is true in the case of both shed and cell surface material but is far more extreme in the latter, with the HS reducing from approximately 45% in the controls to less than 10% in HME patients. Initial analysis also reveals shortened chain length within these samples; indeed they often have two populations of chains present. Simple analysis of the total disaccharide composition of these samples demonstrates no significant differences against controls. However, detailed analysis of the subpopulations of chains (as determined by chain length) within these samples as well as cartilaginous samples from exostoses patients may provide further insight into the changes that occur within the biosynthetic pathway following disrupted EXT function.

We are also carrying out immunocytochemistry with a variety of HS‐specific antibodies with the aim to further investigate normal HS structure and localization. This is being carried out on human primary chondrocytes isolated from normal patients and also adult mesenchymal stem cells as they undergo differentiation into chondrocytes. HS has been identified in both these cell types, and it is hoped that the manipulation of these cells through RNAi of different enzymes of the HS biosynthetic pathway will provide a suitable model for studying what changes may occur in cellular HS structures over the initial differentiation process in the growth plate.

Discussion  Together, these investigations should provide a good model to allow us to determine the role of HS in chondrocyte differentiation and maturation in both normal and diseased states.
Epitope mapping of proteins for heparin binding sites
Authors: Vivès Romain R. , Crublet Elodie , Andrieu Jean‐Pierre , Gagnon Jean , Lortat‐Jacob Hugues
Published: August 2004

Introduction  Heparan sulfate (HS) is a cell surface and basement membrane, sulfated polysaccharide involved in a huge array of biological processes, such as cell proliferation, chemo‐attraction, inflammation, matrix assembly, embryo development or viral attachment. These multiple functions stem from HS ability to bind and modulate the activity of a large number of proteins. The interaction of HS with its ligands is primarily, although not exclusively, of an electrostatic nature and involves the recognition of basic domains on the protein by specific motifs of the polysaccharide. Importantly, subtle differences in HS structure have been shown to dramatically affect the polysaccharide activity, as observed for HS regulation of bFGF signalling. Therefore, the identification of both protein and saccharide structural determinants implicated in HS/protein binding is critical for understanding the biological role of the interaction. In this study, we have developed a simple technique enabling the identification of HS‐binding sites on proteins.

Materials and methods  Briefly, proteins were immobilized on heparin beads, using an adapted protocol of a zero‐length two‐step cross‐linking method and submitted to proteolytic digestion. Peptides retained on beads, i.e. containing amino acids involved in heparin binding, were then identified by solid phase N‐terminus protein sequencing performed directly on the heparin beads.

Results  This technique was used to analyse a number of HS protein ligands, including the envelope protein gC of a herpes virus (pseudorabies) and the chemokine RANTES. Results obtained pinpointed residues known to be critical for HS binding and thus validate the method.

Conclusion  This technique constitutes a novel tool that should greatly facilitate HS binding site cartography of the ever‐expanding family of heparin‐binding proteins and provides an attractive alternative to more complex approaches such as site‐directed mutagenesis.
Are There Investor Clienteles in Rental Housing?
Authors: Wood Gavin A. , Tu Yong
Published: August 2004

The possible existence of investor clientele groups has received little attention in the real estate finance literature. In this paper we develop a clientele model, which in equilibrium produces a clustering of investors by tax characteristics. Low‐tax‐bracket investors are concentrated in low‐value rental housing that attracts rents which are high in relation to property values. On the other hand, only high‐tax‐bracket investors will be observed in high‐value rental housing, and they charge rents that are low in relation to property values. An empirical model is specified and estimated using a cross section of investors in Australian private rental housing markets. Investor clienteles are detected among property investors, though there is a weak diversification effect indicating that clientele effects may be stronger among single property investors.
Reaction of House Prices to a New Rapid Transit Line: Chicago's Midway Line, 1983–1999
Authors: McMillen Daniel P. , McDonald John
Published: August 2004

This study examines the effect of the new rapid transit line from downtown Chicago to Midway Airport on single‐family house prices before and after the opening of the line. The results show that the housing market anticipated the opening of the line. House prices were being affected by proximity to the stations in the late 1980s and early 1990s—after the plans for the line were well known. The difference between the increase in the value of homes within the sample area as compared with properties farther away from the new transit stations was approximately $216 million between 1986 and 1999.
Homeownership Determinants for Chinese Americans: Assimilation, Ethnic Concentration and Nativity
Authors: Painter Gary , Yang Lihong , Yu Zhou
Published: August 2004

Chinese homeownership rates in the Los Angeles Consolidated Metropolitan Statistical Area adjusted by socioeconomic and housing market characteristics are, on average, 18 percentage points higher than those of native white households. This finding runs contrary to most immigration literature, which suggests that immigrants usually lag behind the host society in measures of economic well‐being. This study focuses on two additional factors, which most economic studies of homeownership choice ignore, that may play a role in helping Chinese households achieve high homeownership in ways that other immigrant groups do not. The results of this analysis find that the high homeownership rates cannot be explained by the English skills of households. The cultural influence of homeowning peers may have partially contributed to the higher homeownership of Chinese households. While living in ethnic Chinese communities lowers homeownership rates, in general, it helps improve the likelihood that Chinese immigrants will own a home. Finally, we find that there is great diversity among Chinese subgroups with respect to their likelihood of owning a home, but very little diversity with respect to the education and income level of Chinese households across subgroups.
Alveolar and plasma concentrations of interleukin‐8 and vascular endothelial growth factor following oesophagectomy
Authors: Cree R. T. J. , Warnell I. , Staunton M. , Shaw I. , Bullock R. , Griffin S. M. , Baudouin S. V.
Published: August 2004

The acute respiratory distress syndrome occurs in approximately 10% of all patients undergoing elective oesophagectomy. Local increases in lung pro‐inflammatory cytokines have been previously detected in high‐risk patients before the development of the acute respiratory distress syndrome. We hypothesised that similar changes would occur following oesophagectomy. Two groups of patients were studied. In the collapsed lung group (n = 11), interelukin‐8 and vascular endothelial growth factor were measured in bronchoalveolar lavage samples obtained from the intra‐operative collapsed lung after operation. In the ventilated lung group (n = 10), bronchoalveolar lavage was performed after operation from the ventilated lung and cytokines measured. Cytokines were also measured in peripheral blood samples before and after operation. Bronchoalveolar lavage cytokine levels in both lungs were of an order of magnitude greater than in peripheral blood. Pulmonary pro‐inflammatory cytokine release occurs following oesphageal surgery and may indicate subclinical lung injury.
Interobserver reliability between a nurse and anaesthetist of tests used for predicting difficult tracheal intubation
Authors: Hilditch W. G. , Kopka A. , Crawford J. M. , Asbury A. J.
Published: August 2004

We examined the interobserver reliability, between a nurse and anaesthetist, of five tests used to predict difficult tracheal intubation: mouth opening; thyromental distance; head and neck movement; mandibular luxation; and assessment of oropharyngeal view. For each test, an anaesthetic nurse and a specialist registrar anaesthetist were trained to use a standard method of examination. Most of the tests had either good or very good reliability. Assessment of mouth opening demonstrated only moderate reliability and assessment of oropharyngeal view demonstrated poor reliability. The interobserver reliability estimates between a nurse and an anaesthetist are similar to those previously demonstrated between two anaesthetists.