bluuebunny/crossref_metadata_2025 huggingface dataset:
Information and Communication Technology Policy in European Cities: A Comparative Approach
Authors: Cohen Galit , Nijkamp Peter
Published: October 2002

Information and communication technology (ICT) is widely accepted as a potentially favourable set of instruments, which may improve the welfare and competitiveness of nations and cities. Nowadays, both public and private actors aim to exploit the expected benefits of ICT developments. The authors seek to investigate the potential of ICT use at an urban level and, in particular, to shed more light on various factors that influence urban ICT policies in the public domain. First, a conceptual framework, designed to improve understanding of the driving forces of urban ICT policies, is outlined. It focuses on the way decisionmakers perceive their city, and shape their opinions about ICT; it addresses in particular the way these decisionmakers evaluate the importance of ICT for their city. Next, interviews with urban decisionmakers in different European cities in three countries (Austria, Spain, and the Netherlands) are used to analyse the complex relationship between perceived urban characteristics (for example, nature of problems and urban image), personal attitudes towards ICT, administrative features of the cities concerned, and perceptions of the relevance of ICT to the cities. The authors' main focus is on the identification of a possible systematic relationship between the aforementioned explanatory factors and urban decisionmakers' attitudes towards ICT policies. Understanding the decisionmakers' perceptions is an important step towards grasping the nature and substance of the policy itself, and may explain some of the variance among different cities. Because the ‘urban ICT’ discourse is still relatively new, an open-interview method is used to capture a variety of different views and perceptions on ICT and on the information age in the city. With the aid of qualitative content analysis, the interview results are transformed into a more systematic and comparable form. The results suggest that even interviewees from the same city may have a different understanding of their urban reality whereas, on the other hand, cities with different characteristics may appear to suffer from similar problems. Moreover, the authors found a wide range of attitudes toward ICT and its expected social impacts, although most of the interviewees appeared to be more sceptical than had been expected. The authors identified a clear need for a more thorough investigation of background factors and, therefore an approach originating from the field of artificial intelligence—rough-set analysis—was deployed to offer a more rigorous analysis. This approach helped in the characterisation and understanding of perceptions and attitudes regarding urban policies, problems, and images.
The Usefulness of the GIS—Fuzzy Set Approach in Evaluating the Urban Residential Environment
Authors: Oh Kyushik , Jeong Yeunwoo
Published: October 2002

The authors' focus was to determine the usefulness of the fuzzy set approach in evaluating the urban residential environment, compared with the crisp (or Boolean) approach. Particular emphasis was placed upon the comparison of evaluation results produced by the two methods within a geographic information system (GIS). This comparison highlighted the advantages of the GIS-fuzzy set approach as follows. First, it was revealed that the fuzzy set approach could reduce excessive abstraction or exaggeration in environmental phenomena. Hence, without the loss of valuable information, more accurate decisionmaking can be rendered. Second, by integrating membership functions into GIS, greater efficiency of the entire evaluation process was achieved.
Contribution of Color to Face Recognition
Authors: Yip Andrew W , Sinha Pawan
Published: October 2002

One of the key challenges in face perception lies in determining how different facial attributes contribute to judgments of identity. In this study, we focus on the role of color cues. Although color appears to be a salient attribute of faces, past research has suggested that it confers little recognition advantage for identifying people. Here we report experimental results suggesting that color cues do play a role in face recognition and their contribution becomes evident when shape cues are degraded. Under such conditions, recognition performance with color images is significantly better than that with gray-scale images. Our experimental results also indicate that the contribution of color may lie not so much in providing diagnostic cues to identity as in aiding low-level image-analysis processes such as segmentation.
The Boogie-Woogie Illusion
Authors: Cavanagh Patrick , Anstis Stuart
Published: October 2002

A grid of vertical and horizontal lines, each composed of light and dark squares, is moved rigidly at 45° to the vertical on a gray surround. When the luminance of the background is set midway between the luminances of the light and dark squares, the squares appear to race along the lines even though they are actually ‘painted’ on the lines. The effect arises from the unequal apparent speeds of the lines and their textures. The light and dark squares along the lines define a first-order pattern whose apparent speed, parallel or along the line, is close to veridical. The lines themselves have no overall luminance difference from the background so that they are defined by a second-order difference. As reported elsewhere, apparent speed is reduced for second-order motion so that the motion perpendicular to the line is perceived as slower than the motion along the line even though they are physically equal. The imbalance creates the impression that the small squares are moving along the lines rather than moving rigidly with them.
Gab1 and SHP-2 promote Ras/MAPK regulation of epidermal growth and differentiation
Authors: Cai Ti , Nishida Keigo , Hirano Toshio , Khavari Paul A.
Published: October 2002

În epidermis, Ras can influence proliferation and differentiation; however, regulators of epidermal Ras function are not fully characterized, and Ras effects on growth and differentiation are controversial. EGF induced Ras activation in epidermal cells along with phosphorylation of the multisubstrate docking protein Gab1 and its binding to SHP-2. Expression of mutant Gab1Y627F deficient in SHP-2 binding or dominant-negative SHP-2C459S reduced basal levels of active Ras and downstream MAPK proteins and initiated differentiation. Differentiation triggered by both Gab1Y627F and SHP-2C459S could be blocked by coexpression of active Ras, consistent with Gab1 and SHP-2 action upstream of Ras in this process. To study the role of Gab1 and SHP-2 in tissue, we generated human epidermis overexpressing active Gab1 and SHP-2. Both proteins stimulated proliferation. In contrast, Gab1Y627F and SHP-2C459S inhibited epidermal proliferation and enhanced differentiation. Consistent with a role for Gab1 and SHP-2 in sustaining epidermal Ras/MAPK activity, Gab1−/− murine epidermis displayed lower levels of active Ras and MAPK with postnatal Gab1−/− epidermis, demonstrating the hypoplasia and enhanced differentiation seen previously with transgenic epidermal Ras blockade. These data provide support for a Ras role in promoting epidermal proliferation and opposing differentiation and indicate that Gab1 and SHP-2 promote the undifferentiated epidermal cell state by facilitating Ras/MAPK signaling.
Both midzone and astral microtubules are involved in the delivery of cytokinesis signals
Authors: Murata-Hori Maki , Wang Yu-li
Published: October 2002

To address the mechanism that coordinates cytokinesis with mitosis, we have studied the dynamics of aurora B, a chromosomal passenger protein involved in signaling cytokinesis. Photobleaching analyses indicated dynamic exchange of aurora B between a centromeric and a cytoplasmic pool before anaphase onset, and stable associations with microtubules after anaphase onset. Bleaching near centromeres upon anaphase onset affected the subsequent appearance of fluorescence along midzone microtubules, but not that near the lateral equatorial cortex, suggesting that there were centromeric-dependent and -independent pathways that transported aurora B to the equator. The former delivered centromeric aurora B along midzone microtubules, whereas the latter delivered cytoplasmic aurora B along astral microtubules. We suggest that cultured cells use midzone microtubules as the primary signaling pathway for cytokinesis, whereas embryos, with their stockpile of cytoplasmic proteins and large sizes, rely primarily on astral microtubules.
Pricing on the Internet
Authors: Kung Mui , Monroe Kent B. , Cox Jennifer L.
Published: October 2002

Conventional theories suggest that the Internet will drive down prices and lead to perfectly competitive prices. However, there is contradictory evidence indicating that online prices are not absolutely lower than offline stores. Regardless, the Internet gives rise to many opportunities for leveraging pricing strategies, in research and testing capabilities, customer segmentation, dynamic pricing, product differentiation, developing brand loyalty, including shipping and handling in the profitability analysis, offering multiple versions, and creating or participating in electronic marketplaces. The trading platform of eBay, Priceline’s reverse auction, and price comparison Web sites are examples of novel Internet pricing models that are helping create a new pricing paradigm.
Kinetic and dynamic studies of liposomal bupivacaine and bupivacaine solution after subcutaneous injection in rats
Authors: Yu Hsiu-Ying , Li Shyh-Dar , Sun Pin
Published: October 2002

The pharmacodynamics and pharmacokinetics of bupivacaine in solution and in liposome preparations following subcutaneous administration were studied in rats. Multilamellar vesicles entrapping bupivacaine solution were prepared. The local anaesthetic effect was estimated by the tail-flick test in Wistar rats treated with 1 mg bupivacaine in 0.2-ml. preparations. Plasma concentrations of bupivacaine were determined by high-performance liquid chromatography. The results showed that both bupivacaine solution and bupivacaine liposomes revealed local anaesthetic effects in the initial tail-flick test (15 min after injection). With bupivacaine liposomes, the duration of action was 5-fold (447 ± 28.9 vs 87 ± 6.7 min), the maximum possible effect was 2-fold (100 ± 0 vs 47.6 ± 13%), and the peak plasma concentration (Cmax) was less than one-fifth (0.12 ± 0.04 vs 0.65 ± 0.04 μg mL−1) that with bupivacaine solution. The sensory block effect of bupivacaine solution completely resolved at 90 min, while the plasma concentration of bupivacaine was still more than half the Cmax. Bupivacaine liposomes resulted in a low and relatively constant plasma level (approx. 0.1 μg mL−1) and a pronounced local anaesthetic effect throughout the experimental period (> 7 h). In conclusion, bupivacaine liposomes elevated the intensity and prolonged the duration of the local anaesthetic effect of bupivacaine, and suppressed the systemic absorption rate of encapsulated bupivacaine.
Rapid determination of vitamin C by NIR, MIR and FT-Raman techniques
Authors: Yang Hong , Irudayaraj Joseph
Published: October 2002

Rapid spectroscopic determination of vitamin C in food and pharmaceutical products using infrared and Raman techniques was proposed. In this study, near-infrared (NIR), Fourier transform near-infrared (FT-NIR), Fourier transform infrared-attenuated total reflectance (FTIR-ATR), diffuse reflectance (DRIFTS), Fourier transform infrared-photoacoustic (FTIR-PAS) and FT-Raman spectroscopy were used in conjunction with partial least squares (PLS) regression to quantify vitamin C in powdered mixtures and solutions. Results indicate that the methods adopted have high prediction correlation. R2 values were 0.999 for FTIR-ATR, 0.976 for DRIFTS, 0.966 for FTIR-PAS, 0.988 for NIR, 0.992 for FT-NIR and 0.95 for FT-Raman, with an overall prediction error of 0.2–3.0%. The time required to complete an experiment ranged from 5 s (NIR) to 3 min (FT-Raman). The FTIR and FT-Raman techniques can be complementary tools for qualitative and quantitative characterization of the samples. Infrared and Raman techniques can be used to quantify vitamin C in foods and pharmaceutical products.
Uptake of cyclic dipeptide by PEPT1 in Caco-2 cells: phenolic hydroxyl group of substrate enhances affinity for PEPT1
Authors: Mizuma Takashi , Narasaka Takuo , Awazu Shoji
Published: October 2002

Uptake of cyclic dipeptides by H+/oligopeptide cotransporter (PEPT1) was studied in monolayers of the human intestinal cell line, Caco-2. The cyclic dipeptides studied were cyclic glycylphenylalanine (cyclo(Gly-Phe)), cyclic phenylalanylserine (cyclo(Phe-Ser)), cyclic seryltyrosine (cyclo(Ser-Tyr)) and cyclic glycyltyrosine (cyclo(Gly-Tyr)). These molecules have both peptide bonds and aromatic rings, and are similar in structure to cephalexin and cephadroxil, which are transported by PEPT1. Cellular uptake of these cyclic dipeptides was pH dependent, and was inhibited by the addition of PEPT1 substrates such as glycylsarcosine, indicating PEPT1-mediated transport. Michaelis constants (Km) for these cyclic dipeptides were cyclo(Ser-Tyr) < cyclo(Phe-Ser), and cyclo(Gly-Tyr) < cyclo(Gly-Phe), indicating that tyrosine possessing phenol moiety has higher affinity for PEPT1 than phenylalanine possessing benzen moiety. The Km for cephadroxil possessing phenol moiety was reportedly lower than that for cephalexin possessing benzen moiety. Therefore, it was concluded that the phenolic hydroxyl group of the substrate may enhance affinity for PEPT1.
PC cluster parallel finite element analysis of sloshing problem by earthquake using different network environments
Authors: Kashiyama Kazuo , Tanaka Seizo , Sakuraba Masaaki
Published: October 2002

This paper presents a parallel finite element method for the analysis of the sloshing problem caused by earthquakes. The incompressible Navier–Stokes equation based on Arbitrary Lagrangian–Eulerian description is used as the governing equation. The SUPG/PSPG formulation is employed to improve the numerical stability and the accuracy. Parallel implementation of the unstructured grid based formulation was carried out on a PC cluster. The present method was applied to analyse the sloshing problem of a rectangular tank and an actual reservoir. The effect of parallelization on the efficiency of the computations was examined using a number of different network environments. Copyright © 2002 John Wiley & Sons, Ltd.
A priori pivoting in solving the Navier–Stokes equations
Authors: Wille S. Ø. , Loula A. F. D.
Published: October 2002

Mixed finite element formulations of incompressible Navier–Stokes Equations leads to non‐positive definite algebraic systems inappropriate for iterative solution techniques. However, introducing a suitable preconditioner, the mixed finite element equation system becomes positive definite and solvable by iterative techniques. The present work suggests a priori pivoting sequences for parallel and serial implementations of incomplete Gaussian factorization. Tests are performed for the driven cavity problem in two and three dimensions. Copyright © 2002 John Wiley & Sons, Ltd.
Self designing structures: a new evolutionary rule for thickness distribution in 2D problems
Authors: Marckmann G. , Bettess P. , Peseux B.
Published: October 2002

This paper deals with an evolutionary rule for material distribution in topology optimization problems. This rule is determined in order to satisfy the design constraints that can appear in an industrial design problem. Some other topology optimization techniques are presented before describing our scheme. The classical MBB problem is used to illustrate the capabilities of our approach. The influences of a design parameter and the mesh refinement are discussed. Copyright © 2002 John Wiley & Sons, Ltd.
Studies on biodegradation of phenol using response surface methodology
Authors: Sheeja R Y , Murugesan T
Published: October 2002

Biodegradation of phenol was studied using Pseudomonas pictorum (NCIM 2077) immobilized on alginate and activated carbon – alginate beads. Control experiments were also performed using free cells and non‐inoculated activated carbon – alginate beads. The entrapped alginate and activated carbon – alginate beads suffer from a concentration gradient for oxygen in the interior of the beads and hence free cells showed better degradation at lower concentrations of phenol. The results on entrapped alginate beads were modeled using response surface methodology which determines the dependency of the maximum percentage of phenol degraded as a function of the independent variables, namely initial phenol concentration, initial pH, temperature, and diameter of the immobilized beads. The predicted values are in close agreement with the experimental values with the coefficient of correlation equal to 0.9999 and 0.9993 for both P pictorum – alginate beads and activated carbon – P pictorum – alginate beads respectively.

© 2002 Society of Chemical Industry
Selective recovery of silver ions from aqueous solutions using modified silica beads with Adogen 364
Authors: Lee Wonyoung , Kim Changmook , Yi Jongheop
Published: October 2002

In this work, a method to prepare new adsorbents for the recovery of silver ions was studied. Cheap silica beads for industrial use were obtained and silanized with dichlorodimethylsilane after pretreatment with hot water to increase the silanol contents, followed by immobilization of Adogen 364 (a mixture of trialkyl tertiary amines, Witco Co), a commercial solvent extractant that has a good affinity with silver ions. This sample, designated as K2WDA, was proven to be effective for the adsorption of silver ions from aqueous solutions with fast adsorption rates where sodium nitrate was added as a complexation agent to increase the adsorption capacity of the basic extractant. The adsorption capacity of K2WDA at pH 5 was 0.125 mmol g−1 and corresponded to the proposed mechanism that silver ions adsorb on the functional group with a 1:1 molecular ratio. Selectivity for silver ions in synthetic plating wastewater containing Cu2+ and Ni2+ ions was excellent due to thermodynamic stability. In addition, 80% of the silver ions adsorbed could be recovered with sodium thiosulfate solution rather than acids and the solution was concentrated more than 10 times compared with the original solution. For practical application, a semi‐batch reactor was considered and numerical modeling was performed.

© 2002 Society of Chemical Industry
A first trimester trisomy 13/trisomy 18 risk algorithm combining fetal nuchal translucency thickness, maternal serum free β‐hCG and PAPP‐A
Authors: Spencer Kevin , Nicolaides Kypros H.
Published: October 2002

This study examines 45 cases of trisomy 13 and 59 cases of trisomy 18 and reports an algorithm to identify pregnancies with a fetus affected by trisomy 13 or 18 by a combination of maternal age fetal nuchal translucency (NT) thickness, and maternal serum free β‐hCG and PAPP‐A at 11–14 weeks of gestation. In this mixed trisomy group the median MoM NT was increased at 2.819, whilst the median MoMs for free β‐hCG and PAPP‐A were reduced at 0.375 and 0.201 respectively. We predict that with the use of the combined trisomy 13 and 18 algorithm and a risk cut‐off of 1 in 150 will for a 0.3% false positive rate allow 95% of these chromosomal defects to be identified at 11–14 weeks. Such algorithms will enhance existing first trimester screening algorithms for trisomy 21. Copyright © 2002 John Wiley & Sons, Ltd.
Prenatal diagnosis of trisomy 21 by i(21q): a rare case of fetoplacental chromosomal discrepancy
Authors: Gilardi J. L. , Perrotin F. , Paillet C. , Blesson S. , Cave H. , Briault S. , Moraine C.
Published: October 2002

Objective

A study was conducted to explain the mechanism of an unusual discrepancy between short‐ and long‐term culture examination methods of chorionic villus sampling (CVS).

Method

In a 29‐year‐old Caucasian woman, transabdominal CVS was carried out at 12 weeks of gestation. Non‐mosaic karyotype 46,XX,i(21q) was found on long‐term CVS culture but number and morphology of chromosomes were normal on short‐term culture, amniocyte culture, hygroma colli fluid and fetal fibroblast.

Results

Chromosomal aberration probably appeared after the trophoblast cell line differentiation, four days after fertilization, by means of a 21 centromere misdivision and formation of a i(21q) with secondary positive selection of the 46,XX,i(21q) cell line and loss of the 46,XX in the fetus.

Conclusion

The restricted number of cases with this type of discrepancy limits the possibility of drawing generalised conclusions. In case of discrepancy, we recommend confirmation by amniocentesis or by fetal blood combined with sonographic examination to provide a more definitive diagnosis. Copyright © 2002 John Wiley & Sons, Ltd.
Prenatal diagnosis of Crigler–Najjar syndrome type I by single‐strand conformation polymorphism (SSCP)
Authors: Francoual Jeanne , Trioche Pascale , Mokrani Chahnez , Seboui Hassen , Khrouf Naïma , Chalas Jacqueline , Clement Marina , Capel Liliane , Tachdjian Gérard , Labrune Philippe
Published: October 2002

Crigler–Najjar syndrome type I (CN‐I) is a rare and severe inherited disorder of bilirubin metabolism, caused by the total deficiency of bilirubin‐UDP‐glucuronosyltransferase (UGT) activity. Enzymatic diagnosis cannot be performed in chorionic villi or amniocytes as UGT is not active in these tissues. The cloning of the UGT1 gene and the identification of disease‐causing mutations have led to the possibility of performing DNA‐based diagnosis. Here we report DNA‐based prenatal diagnosis of CN‐I in two Tunisian families in whom CN‐I patients were diagnosed. As we had previously shown that CN‐I was, in Tunisia, associated with homozygosity for the Q357R mutation within the UGT1 gene, we were able to detect this mutation in both families and to show that it was easily recognized by single‐strand conformation polymorphism (SSCP) analysis. In both cases, SSCP analysis of fetal DNA showed that the fetus was heterozygous for the Q357R mutation. In one family, the pregnancy was carried to term and a healthy baby was born, whereas, in the other family, the pregnancy is still continuing. Thus the prenatal diagnosis of CN‐I is possible, provided disease‐causing mutations have been identified. SSCP analysis of DNA prepared either from amniocytes or from chorionic villi is a simple, reliable and fast method for prenatal diagnosis. Copyright © 2002 John Wiley & Sons, Ltd.
Using a qualitative research method for building a software maintenance methodology
Authors: Polo Macario , Piattini Mario , Ruiz Francisco
Published: October 2002

This article explains our experience of using Action Research to develop a software maintenance methodology involving two organizations: a group of university researchers and a software services organization. The concept of ‘methodology’ comprises a wide set of elements whose identification, definition and integration is not a trivial task, due to the magnitude of the project and to the different nature of the organizations. The use of Action Research was a key factor in the progress of the research and has been essential in the adoption of the methodology within the software services organization. Copyright © 2002 John Wiley & Sons, Ltd.
CD++: a toolkit to develop DEVS models
Authors: Wainer Gabriel
Published: October 2002

The features of a toolkit for modeling and simulation based on the DEVS formalism are presented. The tool is built as a set of independent software pieces running on different platforms. Not only are the main characteristics of the environment presented, a focus on its use is also considered by inclusion of application examples for a variety of problems. Many models can be defined in an automated fashion, simplifying the construction of new models and easing their verification. The use of this formal approach has allowed the development of safe and cost‐effective simulations, significantly reducing development time. Copyright © 2002 John Wiley & Sons, Ltd.
Primary Hyperparathyroidism Revisited in Menopausal Women with Serum Calcium in the Upper Normal Range at Population‐based Screening 8 Years Ago
Authors: Lundgren Ewa , Hagström Emil G. , Lundin Jonas , Winnerbäck Kajsa , Roos Johanna , Ljunghall Sverker , Rastad Jonas
Published: October 2002

Population‐based screening showed 2.1% prevalence of primary hyperparathyroidism (pHPT) in postmenopausal women. Individuals with total serum (s)‐calcium levels of 2.55 mmol/L or more at screening were diagnosed with pHPT when subsequent analysis supported inappropriately elevated intact parathormone (PTH) levels in relation to even normal s‐calcium levels. The arbitrary diagnostic criteria were validated by parathyroidectomy. Herein we reinvestigated biochemical signs of pHPT in women not diagnosed with pHPT due to s‐calcium 2.50 to 2.54 mmol/L (group A, n = 160) at screening or due to appropriate PTH levels on two occasions after screening (group B, n = 70). Altogether, 99 women in group A and 47 in group B underwent reinvestigation 8.8 years after screening when they were 65 to 84 years old. The s‐calcium levels averaged 2.56 mmol/L and had increased in group A (mean 0.04 mmol/L) and decreased in group B (mean 0.05 mmol/L). A total of 48 and 18 females (48%, 38%), respectively, met the previously validated criteria of pHPT. Altogether 21% of them were hypercalcemic (range 2.60–3.12 mmol/L). Subgroup analysis showed that PTH had not increased with time (n = 47) and that atherogenic blood lipids, but not glucose levels, were similar in pHPT patients and matched controls (n = 37). Assuming the existence of pHPT already at screening, the prevalence of pHPT could be adjusted to 3.4%. Even the most liberal diagnostic criteria utilized at pHPT screening seemed to underdiagnose the disease by inefficient cutoff limits for s‐calcium and PTH. Because one‐fifth of the women with pHPT progressed to hypercalcemia, long‐term follow‐up is advocated for those with s‐calcium in the upper normal range.
Effect of Surgery on the Outcome of Midgut Carcinoid Disease with Lymph Node and Liver Metastases
Authors: Hellman Per , Lundström Tobias , Öhrvall Ulf , Eriksson Barbro , Skogseid Britt , Öberg Kjell , Tiensuu Janson Eva , Åkerström Göran
Published: October 2002

We have evaluated survival and tumor‐related symptoms in the presence of mesenteric lymph node and liver metastases in relation to surgical procedures in 314 patients
(148 women, mean age at diagnosis 61 years; 249 with liver metastases)
treated for midgut carcinoid tumors. Of the operated patients, 46%
presented with severe abdominal pain and intestinal obstruction and were operated on before the diagnosis. Medical treatment (somatostatin analogs, interferon‐a) was initiated in 67% and 86%, respectively.
Surgical attempts included small intestine or ileocecal/right‐sided colon resection with excision of mesenteric lymph node metastases. Most of the patients (n = 286) had mesenteric lymph node metastases; 33% of them had unresectable mesenteric lymph node metastases and underwent surgery without mesenteric dissection.
Patients who underwent resection for the primary tumor had a longer survival than those with no resection (median survival 7.4 vs. 4.0
years; p <0.01). Patients who underwent successful excision of mesenteric metastases had a significantly longer survival than those with remaining lymph node metastases. Patients operated on for a primary tumor but with remaining lymph nodes but no liver metastases and who subsequently received interferon and somatostatin analog treatment had a median survival of 7.4 years. Resection of the primary tumor and the mesenteric lymph node metastases led to a significant reduction in tumor‐related symptoms. Surgery to remove the primary intestinal tumor including mesenteric lymph node metastases is supported by the present results, even in the presence of liver metastases. Liver metastases and significant preoperative weight loss are identified as major negative prognostic factors for survival.
Prolactin activation of IRF‐1 transcription involves changes in histone acetylation
Authors: Book McAlexander Melissa , Yu-Lee Li-yuan
Published: October 2002

In response to prolactin (PRL) signaling, transcription of the interferon regulatory factor‐1 gene (IRF‐1) is rapidly induced during early G1, declines in mid G1, and rises again over the G1/S transition phase of the cell cycle in Nb2 T cells. Using chromatin immunoprecipitation assays, we show that histone H4 acetylation increases over a 1.7 kb region of the IRF‐1 promoter in early G1 and again at the G1/S transition in response to PRL stimulation. These results demonstrate a correlation between histone H4 hyperacetylation at the IRF‐1 promoter and biphasic transcription of IRF‐1 in response to PRL signaling in vivo.
Overexpression of glutamine:fructose‐6‐phosphate‐amidotransferase induces transforming growth factor‐β1 synthesis in NIH‐3T3 fibroblasts
Authors: Weigert Cora , Brodbeck Katrin , Lehmann Rainer , Häring Hans U. , Schleicher Erwin D.
Published: October 2002

Increased flux through the hexosamine biosynthetic pathway with glutamine:fructose‐6‐phosphate‐amidotransferase (GFAT) as rate‐limiting enzyme has been linked to the enhanced bioactivity of the prosclerotic cytokine transforming growth factor β1 (TGF‐β1) in fibrotic complications, particularly in diabetic kidney disease. Here, we investigate in a stable transfection system the effect of overexpression of GFAT on TGF‐β1 synthesis in NIH‐3T3 fibroblasts. We demonstrate a 1.8‐fold stimulation of TGF‐β1 mRNA and a 1.9‐fold increased protein expression, whereas TGF‐β2 production was not upregulated. The 1.5‐fold enhanced TGF‐β1 promoter activity suggests a transcriptional regulation. The elevated TGF‐β1 protein is biologically active since GFAT‐overexpressing cells exhibit a 2‐fold fibronectin production. The results indicate a GFAT‐dependent induction of TGF‐β1 synthesis.
Reconstitution of hepatitis C virus protease activities in yeast
Authors: Mak Paul , Palant Olga , Labonté Patrick , Plotch Steve
Published: October 2002

The hepatitis C virus (HCV) protease genes (NS2/3 and NS3) were expressed in yeast with their natural substrates fused to a ligand‐dependent transcriptional activator, the retinoic acid receptor (RARβ). RARβ can activate transcription in yeast cells in response to retinoic acids. We hypothesized that cis‐cleavage at the NS2–3 or NS3–4A junctions by the appropriate HCV proteases would release RARβ, thereby activating transcription of a reporter gene. Our results from Western blot analyses and reporter gene activation indicate that the wild‐type NS2/3 and NS3 enzymes are catalytically active in yeast cells, whereas mutations in the catalytic domain of NS2(C993V) and NS3(S1165A) lead to inactive enzymes. We conclude that HCV NS2/3 and NS3 protease activities can be reconstituted in yeast.
Structural elements of the osteopontin SVVYGLR motif important for the interaction with α4 integrins
Authors: Green Philip M. , Ludbrook Steven B. , Miller David D. , Horgan Carmel M.T. , Barry Simon T.
Published: October 2002

The osteopontin SVVYGLR motif binds the integrins α4β1 and α9β1. We show that α4β7 also interacts with this motif and that an SVVYGLR‐OH peptide antagonises the α4β7 MAdCAM interaction. The important elements of this motif required to bind α4β1 and α4β7 were probed using a series of mutated peptides based around SVVYGLR. Leu167 is important for the interaction with α4 integrins, as is the C‐terminal carboxylic acid of Arg168 exposed by thrombin cleavage. The importance of the acidic group means that SVVYGLR has structural elements in common with other α4 integrin‐binding motifs and suggests why thrombin cleavage activates this motif.
A study on the human mitochondrial RNA polymerase activity points to existence of a transcription factor B‐like protein
Authors: Prieto-Martı́n Ascensión , Montoya Julio , Martı́nez-Azorı́n Francisco
Published: October 2002

In the present work, the RNA polymerase activity of the human mitochondrial RNA polymerase mature protein (h‐mtRPOLm) is shown, and its molecular activity calculated (2.1±0.9 min−1). An activity analysis of h‐mtRPOLm and deleted versions of it has demonstrated that the entire recombinant protein is required for this activity. In addition, h‐mtRPOLm alone or in presence of the known mitochondrial transcription factors (human mitochondrial transcription factor A and/or human mitochondrial transcription termination factor) is not able to initiate transcription from the specific human mitochondrial promoters pointing to the existence of a human mitochondrial transcription factor B‐like protein.
Evidence from time resolved studies of the P700+/A1− radical pair for photosynthetic electron transfer on both the PsaA and PsaB branches of the photosystem I reaction centre
Authors: Muhiuddin I.P. , Heathcote P. , Carter S. , Purton S. , Rigby S.E.J. , Evans M.C.W.
Published: October 2002

Kinetic analysis using pulsed electron paramagnetic resonance (EPR) of photosynthetic electron transfer in the photosystem I reaction centres of Synechocystis 6803, in wild‐type Chlamydomonas reinhardtii, and in site directed mutants of the phylloquinone binding sites in C. reinhardtii, indicates that electron transfer from the reaction centre primary electron donor, P700, to the iron–sulphur centres, Fe–SX/A/B, can occur through either the PsaA or PsaB side phylloquinone. At low temperature reaction centres are frozen in states which allow electron transfer on one side of the reaction centre only. A fraction always donates electrons to the PsaA side quinone, the remainder to the PsaB side.
Activation of c‐Jun amino‐terminal kinase is required for retinoic acid‐induced neural differentiation of P19 embryonal carcinoma cells
Authors: Wang Hong , Ikeda Shoko , Kanno Shin-ichiro , Guang Li Ming , Ohnishi Motoko , Sasaki Masato , Kobayashi Takayasu , Tamura Shinri
Published: October 2002

P19 embryonal carcinoma cells are known to differentiate into neurons and glia when treated with relatively high concentrations (>100 nM) of retinoic acid (RA). Concomitant with this RA‐induced neural differentiation, we observed an activation of the c‐Jun amino‐terminal kinase (JNK). JNK was required for the RA‐induced neural differentiation, because dominant‐negative JNK blocked the differentiation. Studies using protein phosphatase inhibitors and protein kinase inhibitors suggested that both okadaic acid‐sensitive protein phosphatase(s) and protein kinase C participate in the RA‐induced activation of JNK.
Nitrotyrosine mimics phosphotyrosine binding to the SH2 domain of the src family tyrosine kinase lyn
Authors: Mallozzi Cinzia , Di Stasi Anna Maria Michela , Minetti Maurizio
Published: October 2002

The nitration of tyrosine residues in protein occurs through the action of reactive oxygen and nitrogen species and is considered a marker of oxidative stress under pathological conditions. The most active nitrating species so far identified is peroxynitrite, the product of the reaction between nitric oxide and superoxide anion. Previously, we have reported that in erythrocytes peroxynitrite irreversibly upregulates lyn, a tyrosine kinase of the src family. In this study we investigated the possible role of tyrosine nitration in the mechanism of lyn activation. We found that tyrosine containing peptides modelled either on the C‐terminal tail of src kinases or corresponding to the first 15 amino acids of human erythrocyte band 3 were able to activate lyn when the tyrosine was substituted with 3‐nitrotyrosine. The activity of nitrated peptides was shared with phosphorylated but not with unphosphorylated, chlorinated or scrambled peptides. Recombinant lyn src homology 2 (SH2) domain blocked the capacity of the band 3‐derived nitrotyrosine peptide to activate lyn and we demonstrated that this peptide specifically binds the SH2 domain of lyn. We propose that nitropeptides may activate src kinases through the displacement of the phosphotyrosine in the tail from its binding site in the SH2 domain. These observations suggest a new mechanism of peroxynitrite‐mediated signalling that may be correlated with the upregulation of tyrosine phosphorylation observed in several pathological conditions.
Solution NMR studies of the integral membrane proteins OmpX and OmpA from Escherichia coli
Authors: Fernández César , Hilty Christian , Bonjour Sophie , Adeishvili Koba , Pervushin Konstantin , Wüthrich Kurt
Published: October 2002

Membrane proteins are usually solubilized in polar solvents by incorporation into micelles. Even for small membrane proteins these mixed micelles have rather large molecular masses, typically beyond 50 000 Da. The NMR technique TROSY (transverse relaxation‐optimized spectroscopy) has been developed for studies of structures of this size in solution. In this paper, strategies for the use of TROSY‐based NMR experiments with membrane proteins are discussed and illustrated with results obtained with the Escherichia coli integral membrane proteins OmpX and OmpA in mixed micelles with the detergent dihexanoylphosphatidylcholine (DHPC). For OmpX, complete sequence‐specific NMR assignments have been obtained for the polypeptide backbone. The 13C chemical shifts and nuclear Overhauser effect data then resulted in the identification of the regular secondary structure elements of OmpX/DHPC in solution, and in the collection of an input of conformational constraints for the computation of the global fold of the protein. For OmpA, the NMR assignments are so far limited to about 80% of the polypeptide chain, indicating different dynamic properties of the reconstituted OmpA β‐barrel from those of OmpX. Overall, the present data demonstrate that relaxation‐optimized NMR techniques open novel avenues for studies of structure, function and dynamics of integral membrane proteins.
A refined structure of human aquaporin‐1
Authors: de Groot Bert L. , Engel Andreas , Grubmüller Helmut
Published: October 2002

A refined structure of the human water channel aquaporin‐1 is presented. The model rests on the high resolution X‐ray structure of the homologous bacterial glycerol transporter GlpF, electron crystallographic data at 3.8 Å resolution and a multiple sequence alignment of the aquaporin superfamily. The crystallographic R and free R values (36.7% and 37.8%) for the refined structure are significantly lower than for previous models. Improved geometry and enhanced stability in molecular dynamics simulations demonstrate a significant improvement of the aquaporin‐1 structure. Comparison with previous aquaporin‐1 models shows significant differences, not only in the loop regions, but also in the core of the water channel.
Insights from the structure of the yeast cytochrome bc1 complex: crystallization of membrane proteins with antibody fragments
Authors: Hunte Carola
Published: October 2002

The ubiquinol:cytochrome c oxidoreductase (EC 1.20.2.2, QCR or cytochrome bc
1 complex) is a component of respiratory and photosynthetic electron transfer chains in mitochondria and bacteria. The complex transfers electrons from quinol to cytochrome c. Electron transfer is coupled to proton translocation across the lipid bilayer, thereby generating an electrochemical proton gradient, which conserves the free energy of the redox reaction. The yeast complex was crystallized with antibody Fv fragments, a promising technique to obtain well‐ordered crystals from membrane proteins. The high‐resolution structure of the yeast protein reveals details of the catalytic sites of the complex, which are important for electron and proton transfer.
Diva/Boo is a negative regulator of cell death in human glioma cells
Authors: Naumann Ulrike , Weit Simone , Wischhusen Jörg , Weller Michael
Published: October 2002

Diva is a novel proapoptotic member of the Bcl‐2 protein family which binds apoptosis activating factor‐1 (APAF‐1). Diva is identical with Boo which was identified as a novel antiapoptotic Bcl‐2 family protein. Here, we report that Diva promotes the cell cycle exit of human glioma cells in response to serum deprivation and inhibits apoptosis of these cells induced by CD95 ligand or chemotherapeutic drugs. In glioma cells, Diva interferes with apoptotic signaling downstream of cytochrome c release, but upstream of caspase activation, consistent with an inhibitory effect on the mitochondrial amplification step involving the apoptosome and APAF‐1.
Differences in proton leak kinetics, but not in UCP3 protein content, in subsarcolemmal and intermyofibrillar skeletal muscle mitochondria from fed and fasted rats
Authors: Iossa Susanna , Lionetti Lillà , Mollica Maria Pia , Crescenzo Raffaella , Botta Monica , Samec Sonia , Dulloo Abdul G. , Liverini Giovanna
Published: October 2002

We have investigated the effect of 24‐h fasting on basal proton leak and uncoupling protein (UCP) 3 expression at the protein level in subsarcolemmal and intermyofibrillar skeletal muscle mitochondria. In fed rats, the two mitochondrial populations displayed different proton leak, but the same protein content of UCP3. In addition, 24‐h fasting, both at 24 and 29°C, induced an increase in proton leak only in subsarcolemmal mitochondria, while UCP3 content increased in both the populations. From the present data, it appears that UCP3 does not control the basal proton leak of skeletal muscle mitochondria.
GD3 glycosphingolipid contributes to Fas‐mediated apoptosis via association with ezrin cytoskeletal protein
Authors: Giammarioli Anna Maria , Garofalo Tina , Sorice Maurizio , Misasi Roberta , Gambardella Lucrezia , Gradini Roberto , Fais Stefano , Pavan Antonio , Malorni Walter
Published: October 2002

Efficiency of Fas‐mediated apoptosis of lymphoid cells is regulated, among other means, by a mechanism involving its association with ezrin, a cytoskeletal protein belonging to the 4.1 family of proteins. In the present work, we provide evidence for a further molecule that associates to ezrin in Fas‐triggered apoptosis, the disialoganglioside GD3. In fact, as an early event, GD3 redistributed in membrane‐associated domains in uropods and co‐localized with ezrin. Co‐immunoprecipitation analyses confirmed this result, indicating a GD3–ezrin association. Altogether, these results are suggestive for a role of GD3 in Fas/ezrin‐mediated apoptosis, supporting the view that uropods contain a multimolecular signaling complex involved in Fas‐mediated apoptosis.
Defining the repeating elements in the cysteine‐rich region (CRR) of the CD18 integrin β2 subunit
Authors: Tan Suet-Mien , Walters Susannah E. , Mathew Elizabeth C. , Robinson Martyn K. , Drbal Karel , Shaw Jacqueline M. , Law S.K.Alex
Published: October 2002

The cysteine‐rich region (CRR) of the integrin β subunits is organised into four repeating elements. By expression of a panel of truncated β2 subunits, and CRR segments fused to the C‐terminal end of a CD4 soluble fragment, the segment required for the expression of two monoclonal antibody conformational epitopes was determined. This segment, E482–Q574, contains 16 cysteines representing two repeating units. We have thus defined the CRR unit motif of ‘xC‐‐‐C‐‐‐C‐‐‐CxCxxCxC‐‐‐Cx’, where ‘x’ represents a single residue, and ‘‐‐‐’ represents a stretch of four to 14 residues.
Homodimerization of presenilin N‐terminal fragments is affected by mutations linked to Alzheimer's disease
Authors: Cervantes Sara , Gonzàlez-Duarte Roser , Marfany Gemma
Published: October 2002

Mutations on human presenilins 1 and 2 cause dominant early‐onset familial Alzheimer's disease (FAD). Presenilins are polytopic transmembrane proteins endoproteolytically processed in vivo to N‐ and C‐terminal fragments (NTFs and CTFs). The functional presenilin unit consists of a high molecular weight complex that contains both fragments. Here we show NTF:NTF, CTF:CTF and NTF:CTF interactions by yeast two‐hybrid and in vivo endoplasmic reticulum split‐ubiquitin assays. Our results also highlight the involvement of HL1 – the hydrophilic loop between TMI and TMII – in the NTF:NTF binding site. Besides, nine FAD‐linked presenilin mutations substantially affected HL1:HL1 binding. From the evidence of NTF and CTF homodimerization, we propose the contribution of two NTFs and two CTFs, instead of a single NTF:CTF heterodimer, to the functional presenilin–γ‐secretase complex and that FAD mutations affect the assembly or stability of this complex.
Inhibition of the membrane fusion machinery prevents exit from the TGN and proteolytic processing by furin
Authors: Band Arja M. , Määttä Juha , Kääriäinen Leevi , Kuismanen Esa
Published: October 2002

The Semliki Forest virus (SFV) glycoprotein precursor p62 is processed to the E2 and E3 during the transport from the trans‐Golgi network (TGN) to the cell surface. We have studied the regulation of the membrane fusion machinery (Rab/N‐ethylmaleimide (NEM)‐sensitive fusion protein (NSF)/soluble NSF attachment protein (SNAP)–SNAP receptor) in this processing. Activation of the disassembly of this complex with recombinant NSF stimulated the cleavage of p62 in permeabilized cells. Inactivation of NSF with a mutant α‐SNAP(L294A) or NEM treatment inhibited processing of p62. Rab GDP dissociation inhibitor inhibited the cleavage. Inactivation of NSF blocks the transport of SFV glycoproteins and vesicular stomatitis virus G‐glycoprotein from the TGN membranes to the cell surface. The results support the conclusion that inhibition of membrane fusion arrests p62 in the TGN and prevents its processing by furin.
Apaf‐1 overexpression partially overcomes apoptotic resistance in a cisplatin‐selected HeLa cell line
Authors: Kamarajan Pachiyappan , Sun Nian-Kang , Sun Chun-Ling , Chao Chuck C.-K
Published: October 2002

Inhibition of caspase‐3‐mediated apoptosis has been hypothesized to be associated with chemoresistance. Investigations of apoptosis revealed that cytosolic cytochrome c is associated with a complex of apoptotic protease activating factor‐1 (Apaf‐1), an adapter molecule, and caspase‐9 to activate caspase‐3. However, whether these apoptotic molecules are involved in acquired cisplatin resistance is not understood. The present work shows reduced activation of caspase‐3 and apoptosis in a cisplatin‐selected HeLa cell line. Ac‐DEVD‐CHO, a caspase‐3 inhibitor, inhibited cisplatin‐induced apoptosis about 60–70% in both cell lines. Ac‐LEHD‐CHO, a caspase‐9 inhibitor or Ac‐IETD‐CHO, a caspase‐8 inhibitor, inhibited cisplatin‐induced caspase‐3 activation and apoptosis similarly in both cell lines. In addition, cisplatin induced the activation of caspase‐9, the upstream activator of caspase‐3, in a dose‐dependent manner, and the activation of caspase‐9 was less induced in resistant cells. The accumulation of cytosolic cytochrome c, an activator of caspase‐9, and the induction of the mitochondrial membrane‐associated voltage‐dependent anion channel were also reduced in cisplatin‐resistant cells. However, the concentration of Bcl‐2 family proteins in cisplatin‐resistant cells was normal. The concentration of Apaf‐1 was unaltered in both cell lines. Increasing the cellular concentration of Apaf‐1 through the transient expression of the gene increased the induction of apoptosis in resistant cells, associated with enhanced activation of caspase‐9, caspase‐3 and DNA fragmentation factor. Regression analysis reveals that the modification factor, the ratio of the slope in the linear range of the dose–response curve with Apaf‐1 to the slope without Apaf‐1, is 1.5 and 4.75 in the HeLa and cisplatin‐resistant HeLa cells, respectively. These results indicate that apoptosis and caspases are less induced in cisplatin‐selected HeLa cells. They also suggest that ectopic overexpression of Apaf‐1 may partially reverse the acquired cisplatin resistance.