bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth‐induced changes in the proteome of Helicobacter pylori
Authors: Uwins Christina , Deitrich Christian , Argo Evelyn , Stewart Elizabeth , Davidson Ian , Cash Phil
Published: March 2006

Helicobacter pylori is a major human pathogen that is responsible for a number of gastrointestinal infections. We have used 2‐DE to characterise protein synthesis in bacteria grown either on solid agar‐based media or in each of two broth culture media (Brucella and brain heart infusion (BHI) broth). Significant differences were observed in the proteomes of bacteria grown either on agar‐based or in broth media. Major changes in protein abundance were identified using principal component analysis (PCA), which delineated the profiles derived for the three key growth conditions (i.e. agar plates, Brucella and BHI broth). Proteins detected across the gel series were identified by peptide mass mapping and Edman sequencing. A number of proteins associated with protein synthesis in general as well as specific amino acid synthesis were depressed in broth‐grown bacteria compared to plate‐grown bacteria. A similar reduction was also observed in the abundance of proteins involved in detoxification. Two of the most abundant spots, identified as UreB and GroEL, in plate‐grown bacteria showed a >140‐fold drop in abundance in bacteria grown in Brucella broth compared to bacteria grown on agar plates. Two protein spots induced in bacteria grown in broth culture were both identified as glyceraldehyde 3‐phosphate dehydrogenase based on their N‐terminal amino acid sequences derived by Edman degradation. The underlying causes of the changes in the proteins abundance were not clear, but it was likely that a significant proportion of the changes were due to the alkaline pH of the broth culture media.
Enantiomeric separation of some demethylated analogues of clofibric acid by capillary zone electrophoresis and nano‐liquid chromatography
Authors: Fantacuzzi Marialuigia , Bettoni Giancarlo , D'Orazio Giovanni , Fanali Salvatore
Published: March 2006

The enantiomeric separation of some demethylated analogues of clofibric acid, namely 2‐(6‐chloro‐benzothiazol‐2‐ylsulfanyl)‐, 2‐(6‐methoxy‐benzothiazol‐2‐ylsulfanyl)‐, 2‐(quinolin‐2‐yloxy)‐, 2‐(6‐chloro‐quinolin‐2‐yloxy)‐, 2‐(7‐chloro‐quinolin‐4‐yloxy)‐propionic acid (compounds A–E, respectively), has been studied by CZE and nano‐LC using for the first technique two β‐CD derivatives and vancomycin added to the BGE and vancomycin‐modified silica particles for the second one, with the aim to find the optimum experimental conditions for the baseline resolution. The type and the concentration of the chiral selector added to the BGE, the buffer pH, the type of organic modifier and its concentration, the capillary temperature and the applied voltage played a very important role in the enantioresolution of the analysed compounds. The use of 6‐monodeoxy‐6‐monoamino‐β‐CD allowed to achieve baseline resolution of four of five clofibric acid derivatives in less than 10 min while heptakis‐(2,3,6‐tri‐O‐methyl)‐β‐CD partially resolved the same compounds in their enantiomers. Employing vancomycin as the chiral selector in CZE, the counter‐current partial filling method was chosen achieving baseline resolution of four analytes. All the studied compounds were enantioresolved employing a capillary column packed with vancomycin stationary phase by nano‐LC, and the resolution was strongly influenced by the concentration of the organic modifier and by the pH of the mobile phase. The best results were achieved at pH 4.5 in presence of 60% of methanol (MeOH). However, longer analysis times were observed in the experiments carried out by nano‐LC.
Development of a capillary electrophoretic method for the analysis of low‐molecular‐weight amines from metal working fluid aerosols and ambient air
Authors: Fekete Agnes , Frommberger Moritz , Ping Guichen , Lahaniatis Majlinda R. , Lintelman Jutta , Fekete Jeno , Gebefugi Istvan , Malik Ashok Kumar , Kettrup Antonius , Schmitt‐Kopplin Philippe
Published: March 2006

A method for the determination of low‐molecular‐weight amines from indoor and ambient air was developed using a concentration device followed by CE coupled with indirect spectrophotometric and mass spectrometric detection that enables a reliable, rapid‐response and easy‐to‐operate method. In indirect detection method, the selected amines were separated from interfering metal ions and amino alcohols present in the samples with an imidazole‐based buffer with ethanol and EDTA as modifier. By replacing imidazole with ammonium, the final buffer was applicable for MS detection for the analytes with m/z higher than 50. A novel monolithic polymer material based on poly(methacrylate‐acrylate) copolymer was developed for sampling short‐chain amines from the gaseous phase. The selected analysis conditions were applied to quantify the selected short‐chain amines with detection limits for the whole procedure determined between 1 and 2 µg/filter when 40 L air was sampled with 1 L/min velocity. Improved linearity and precision were obtained when the raw, time‐scaled electropherogram data were transformed into mobility‐scale applied for the determination of the performance characteristics of the methods. The applicability of the process of data transformation into the mobility scale was demonstrated by studying the matrix effect of water‐miscible metal working fluid (stable water–oil emulsion) and of ambient air as real samples. CE‐indirect UV and CE‐MS, combined with the possibility of rapid air sampling, can be useful for the estimation of short‐term exposure of the selected biogenic amines.
Influence of detergent additives on mobility of native and subviral rhinovirus particles in capillary electrophoresis
Authors: Kremser Leopold , Petsch Martina , Blaas Dieter , Kenndler Ernst
Published: March 2006

The electrophoretic properties of two human rhinovirus (HRV) serotypes, HRV2 and HRV14, their subviral particles, and their capsid proteins were investigated by CE using borate buffer, pH 8.3, as BGE and three different detergents as additives. In addition, the influence of modification of the capsid with an amine reactive fluorescent dye, Cy3.5, on migration in the electric field was assessed. We found that the reproducibility of the electrophoretic results was decisively dependent on the presence of the detergents above their respective CMC. As compared to the strong ionic detergent SDS, the nonionic, mild detergent dodecylpoly(ethyleneglycol ether) (D‐PEG) efficiently and reproducibly resolved both, native viruses as well as subviral particles. Most of the analytes behaved as expected except native HRV2; this serotype showed a dramatically higher anionic mobility in SDS than in D‐PEG. Additionally, its mobility decreased when each positive charge contributed from a lysine at the capsid surface was substituted by four negative charges upon derivatization with Cy3.5. We discuss the possibility that this effect is caused by differences in number and in arrangement of exposed lysines in the two serotypes leading to differences in the amount of bound SDS micelles.
Simul 5 – Free dynamic simulator of electrophoresis
Authors: Hruška Vlastimil , Jaroš Michal , Gaš Bohuslav
Published: March 2006

We introduce the mathematical model of electromigration of electrolytes in free solution together with free software Simul, version 5, designed for simulation of electrophoresis. The mathematical model is based on principles of mass conservation, acid–base equilibria, and electroneutrality. It accounts for any number of multivalent electrolytes or ampholytes and yields a complete picture about dynamics of electromigration and diffusion in the separation channel. Additionally, the model accounts for the influence of ionic strength on ionic mobilities and electrolyte activities. The typical use of Simul is: inspection of system peaks (zones), stacking and preconcentrating analytes, resonance phenomena, and optimization of separation conditions, in either CZE, ITP, or IEF.
An experimental study on bubble growth within a narrow space under an interference‐piece
Authors: Yasukawa Yukio
Published: February 2006

An experimental study was performed on the nucleate boiling of water in the narrow space formed between an interference piece and the heating surface of heat‐proof glass. A past study analyzed the bubble growth due to the conduction of heat through the thinner liquid layer under a growing bubble; however, the thickness of the liquid layer was not shown experimentally. This study investigates the thickness of this thinner layer by the interference method. Almost no change was found on the thickness of the liquid layer under both the interference‐piece and on the heating surface. The experimental results indicate the great contribution of superheating of the test liquid to bubble growth. © 2006 Wiley Periodicals, Inc. Heat Trans Asian Res, 35(2): 165–177, 2006; Published online in Wiley InterScience (www.interscience. wiley.com). DOI 10.1002/htj.20100
Thermophysical properties of molten tungsten measured with an electrostatic levitator
Authors: Paradis Paul‐François , Ishikawa Takehiko , Fujii Ryuichi , Yoda Shinichi
Published: February 2006

Thermophysical properties of liquid and supercooled tungsten were measured using non‐contact techniques in combination with an electrostatic levitator. Over the 3125–3707 K temperature range, the density measurements can be expressed as ρ (T) = 16.7(± 0.33)× 103 − 1.08(± 0.08) (kgċ m−3) with Tm = 3695 K, leading to a volume expansion coefficient of 6.6×10−5 K−1. In addition, over the 3398–3695 K temperature range, the surface tension (γ) and viscosity (η) data can be expressed respectively as γ (T) = 2.48× 103(± 75) − 0.31(± 0.08) (T − Tm) (10−3 Nċ m−1) and η (T) = 0.11(± 0.02) exp[12.8(± 4.1) × 104/(RT)] (10−3 Paċ s). © 2006 Wiley Periodicals, Inc. Heat Trans Asian Res, 35(2): 152–164, 2006; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/htj.20101
Research on the thermal radiation characteristics of ionic crystals
Authors: Yong‐hong Wu , De‐hong Xia
Published: February 2006

Based on the microscopic mechanism of thermal radiation of ionic crystals, a model of damping oscillators is established to study and calculate the thermal radiation characteristics of ionic crystals, i.e., absorptivity, reflectivity, and transmissivity. Then the model can be used to calculate the infrared spectra of ionic crystals as well. Comparison of the numerical results with experimental results indicates that the results calculated by the model are in good agreement with the infrared spectra measured. © 2006 Wiley Periodicals, Inc. Heat Trans Asian Res, 35(2): 101–109, 2006; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/htj.20106
Acute effects of caffeine in volunteers with different patterns of regular consumption
Authors: Hewlett Paul , Smith Andrew
Published: March 2006

Rationale

The effects of caffeine on mood and performance are well established. One explanation of these effects is that caffeine removes negative effects induced by prior caffeine withdrawal. This was tested here by comparing effects of caffeine in withdrawn consumers and non‐consumers (who by definition were not withdrawn).

Objectives

The present study aimed to determine whether caffeine withdrawal influenced mood and performance by comparing regular consumers who had been withdrawn from caffeine overnight with non‐consumers. Following this the effects of acute caffeine challenges were compared in withdrawn consumers and non‐consumers. In addition, comparisons were made between those with higher and lower caffeine consumption.

Methods

One hundred seventy‐six volunteers participated in the study. Regular caffeine consumption was assessed by questionnaire and this showed that 56 of the sample did not regularly consume caffeinated beverages. Volunteers were instructed to abstain from caffeine overnight and then completed a baseline session measuring mood and a range of cognitive functions at 08.00 the next day. Following this approximately half of the volunteers were given 1 mg/kg caffeine in a milkshake or water (in the ‘no caffeine’ condition they were given just the milkshake or water) and the test battery repeated one hour later. A second test battery was carried out at 12.00 and a second caffeine challenge at 13.00. A final test session was carried out at 15.00.

Results

The baseline data revealed little evidence of effects of caffeine withdrawal on performance and mood. In contrast to this, caffeine produced a number of significant improvements in performance. There were some differences in the effects of caffeine on regular and non‐consumers, with caffeine tending to reduce reaction time in regular consumers while the opposite was true for non‐consumers.

Conclusions

The present results show little evidence of effects of caffeine withdrawal on performance. In contrast, caffeine challenge produced improvements in aspects of performance and these were often not modified by regular caffeine consumption patterns. The differences in effects of caffeine that were observed between non‐consumers and regular consumers were in functions that were unaffected by caffeine withdrawal. These findings show that the observed beneficial effects of caffeine cannot be interpreted in terms of a reversal of caffeine withdrawal. Copyright © 2006 John Wiley & Sons, Ltd.
Intertemporal pricing and price discrimination: a semiparametric hedonic analysis of the personal computer market
Authors: Stengos T. , Zacharias E.
Published: March 2006

We apply a smooth coefficient semiparametric model to a unique high‐frequency data set to examine the intertemporal pricing of personal computers. Furthermore, we test whether firms charge differential component prices for their top performance personal computers and whether premium firms charge both a premium for all their computers and a premium for their top performance ones. We find nonlinear effects in the pricing of personal computer components. We also find that firms in general do not charge differential prices for the components of their top performance computers. In addition, high‐quality firms charge higher premia only for their most advanced products. Copyright © 2006 John Wiley & Sons, Ltd.
Magazine prices revisited
Authors: Willis Jonathan L.
Published: March 2006

This paper examines price adjustment behaviour in the magazine industry. In a frequently cited study, Cecchetti (1986) constructs a reduced‐form (S, s) model for firms. Cecchetti assumes that a firm's pricing rules are fixed for non‐overlapping three‐year intervals and estimates the model using a conditional logit specification from Chamberlain (1980). The estimates are inconsistent, however, due to the duration‐dependent specification of the model. Two alternative specifications are used to obtain consistent estimation. The consistent estimates continue to provide strong evidence in favour of state‐dependent pricing models, but only weak evidence on the behaviour of price adjustment costs. Copyright © 2006 John Wiley & Sons, Ltd.
Brain, mind, and dyadic change processes
Authors: Neborsky Robert J.
Published: March 2006

For many individuals attachment trauma is at the core of psychoneurosis and personality disorder. Combining theoretical aspects of psychodynamic therapy, developmental neuroscience, and attachment styles provides a useful framework for intensifying emotion and accelerating the course of treatment. A bihemispheric model is considered. The model addresses the challenge in treating the implicit trauma, which resides in the right hemisphere. This is achieved without resorting to interpretation, which is largely a left hemispheric process. The article presents a patient who benefits from a brief emotionally based psychotherapy that was completed after a course of a 20‐year psychoanalysis. © 2006 Wiley Periodicals, Inc. J Clin Psychol: In Session 62: 523–538, 2006.
Emotional processing in the treatment of psychosomatic disorders
Authors: Coughlin Della Selva Patricia
Published: March 2006

Research has supported a hypothesis that was clinically suggested decades ago: that those who have functional disorders exhibit significant deficits in emotional processing. Studies have systematically documented that patients who habitually repress their emotions simultaneously suppress their immune response, rendering them vulnerable to physical illness and early death. Conversely, those who are encouraged to experience and express their feelings demonstrate improvement in immune function, physical condition, and psychological well‐being. This article describes a psychotherapy designed to interrupt defensive processes and facilitate the direct experience of previously disavowed feelings in the treatment of patients who have physical ailments. Verbatim transcripts from the treatment of a woman who had a life‐threatening illness illustrate the therapeutic techniques designed to facilitate emotional processing and restore health. Data from 14 years of follow‐up interviews reveal a sustained remission in her disease. © 2006 Wiley Periodicals, Inc. J Clin Psychol: In Session 62: 539–550, 2006.
Quantum transformation in trauma and treatment: Traversing the crisis of healing change
Authors: Fosha Diana
Published: March 2006

Three ideas are discussed from the vantage point of accelerated experiential dynamic psychotherapy (AEDP): Quantum changes operate not only in trauma, but also in healing; attachment plays a major role in whether fear or excitement is the response to novelty; experientially exploring the experience of transformation can itself lead to quantum change. Vignettes from an AEDP session show a patient's grappling with the startling novelty of experience that the transformational process, particularly when occurring in quantum leaps, evokes, and illustrate the phenomenology of cascading transformations. They illustrate how the therapist's emotional engagement and attachment orientation, experiential techniques, and metatherapeutic processing activate state transformations and entrain adaptive healing processes. © 2006 Wiley Periodicals, Inc. J Clin Psychol: In Session 62: 569–583, 2006.
Adhesion and proliferation of fibroblasts on RF plasma‐deposited nanostructured fluorocarbon coatings: Evidence of FAK activation
Authors: Rosso Francesco , Marino Gerardo , Muscariello Livio , Cafiero Gennaro , Favia Pietro , D'Aloia Erica , d'Agostino Riccardo , Barbarisi Alfonso
Published: February 2006

We used combined plasma‐deposition process to deposit smooth and nanostructured fluorocarbon coatings on polyethylenethereftalate (PET) substrates, to obtain surfaces with identical chemical composition and different roughness, and investigate the effect of surface nanostructures on adhesion and proliferation of 3T3 Swiss Albino Mouse fibroblasts. Untreated PET and polystyrene (PS) were used as controls for cell culture. We have found that the statistically significant increase of cell proliferation rate and FAK (a nonreceptor tyrosine kinase) activation detected on ROUGH fluorocarbon surfaces is due to the presence of nanostructures. Changes in cytoskeletal organization and phospho FAK (tyr 397) localization were evident after 60 min on cells adhering to ROUGH surfaces. This change was characterized by the formation of actin stress fibers along lamellar membrane protrusion instead of usual focal contacts. Also the morphology of the adhering fibroblasts (60 min) adhering on ROUGH surfaces was found quite different compared to cells adhering on smooth ones. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
cDNA‐array profiling of melanomas and paired melanocyte cultures
Authors: Mischiati Carlo , Natali Pier Giorgio , Sereni Alessia , Sibilio Leonardo , Giorda Ezio , Cappellacci Sandra , Nicotra Maria Rita , Mariani Giustino , Di Filippo Franco , Catricalà Caterina , Gambari Roberto , Grammatico Paola , Giacomini Patrizio
Published: March 2006

Three paired (from the same donor) sets of melanoma cells and normal melanocytes, established as early‐passage cultures from metastatic lesions and the uninvolved skin of three patients, were comparatively cDNA profiled by macroarrays (approximately 1,200 genes) and reverse transcription (RT)‐PCR. While 145 gene products were significantly (at least twofold) upregulated or downregulated in at least 1 pair, and 23 were in at least 2 pairs, only 3 (the signal transducer and activator of transcription STAT2, collagen type VI, and CD9) were concordantly modulated (downregulation) in all 3 pairs. Array results were validated by RT‐PCR on a small panel of surgically removed nevocellular nevi and metastatic melanoma lesions, and by immunohistochemistry on a large panel of benign and malignant lesions of the nevomelanocytic lineage. The three gene products were downregulated at different stages of melanoma progression. STAT2 was detectable in nevi (5/5) and most primary melanomas (11/12), but was lost in 10/15 metastatic lesions. Collagen type VI was expressed in nevi (5/5) and primary melanomas below a Breslow thickness of 1 mm (3/3), but was lost in 24/24 primary melanomas above this threshold, and in metastatic melanomas (10/10). The tetraspanin CD9 molecule was expressed in 18/18 nevi, but was lost in 20/28 primary melanomas (including thin lesions), and in 24/52 metastatic lesions. These data provide the proof of principle that cDNA profiling of paired melanocyte/melanoma cultures sorts out novel, early signatures of melanocyte transformation that could contribute to the clinical management of patients at high risk of metastatic disease. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Expression and regulation of CReMM, a chromodomain helicase‐DNA‐binding (CHD), in marrow stroma derived osteoprogenitors
Authors: Marom R. , Shur I. , Hager G.L. , Benayahu D.
Published: March 2006

This study follows the expression of CReMM, a new CHD family member, in osteoprogenitors. CReMM expression was analyzed in primary cultured mesnchymal cells from rat and human. Analysis in ex vivo cultured marrow stromal cells (MSC) from rats revealed higher level of CReMM in cells from young (3 months), when compared to cells from old (15 months) rats. CReMM level was higher in human MSC then in mature trabecular bone cells (TBC). Within the MSC population, osteogenic clones showed higher levels of CReMM then non‐osteogenic ones. We used bone marrow derived osteogenic cell line (MBA‐15) to elaborate on the regulation of CReMM expression in correlation with cell proliferation and co‐expression with alkaline phosphatase (ALK). CReMM is highly expressed in proliferating cells and is inversely related to expression of ALK. MBA‐15 cells were challenged with dexamethasone (Dex) or 17β‐estradiol and quantification of CReMM at the protein (ELISA) and mRNA (RT‐PCR) levels had shown that Dex upregulated CReMM levels. Since CReMM is regulated by Dex, we analyzed the interaction of CReMM with the glucocorticoid receptor (GR), which mediates Dex action. Co‐immunopercipitation (Co‐IP) demonstrated an association between CReMM and GR. In summary, CReMM is a CHD protein expressed by osteoprogenitors, and we suggest it plays a role in mediating transcriptional response to hormones that coordinate osteoblast function. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Downregulation of Akt activity contributes to the growth arrest induced by FGF in chondrocytes
Authors: Priore Riccardo , Dailey Lisa , Basilico Claudio
Published: March 2006

Unregulated FGF signaling produced by activating FGFR3 mutations causes several forms of dwarfism‐associated chondrodysplasias in humans and mice. FGF signaling inhibits chondrocyte proliferation by activating multiple signal transduction pathways that all contribute to chondrocyte growth arrest and induction of some aspects of differentiation. Previous studies had identified the Stat1 pathway, dephosphorylation of the Rb family proteins p107 and p130, induction of p21 expression and sustained activation of MAP kinases as playing a role in the FGF response of chondrocytes. We have examined the role of Akt (PKB) in the response of chondrocytes to FGF signaling. Differently from what is observed in many other cell types, FGF does not activate Akt in chondrocytes, and Akt phosphorylation is actually downregulated after FGF treatment. By expressing a constitutively activated, myristylated form of Akt (myr‐Akt) in the RCS chondrosarcoma cell line, we show that Akt activation partially counteracts the inhibitory effect of FGF signaling. The response of myr‐Akt expressing cells to FGF is identical to parental RCS in the first few hours after treatment, but then diverges as myr‐Akt cells show decreased p130 phosphorylation, increased cyclin E/cdk2 activity and continue to proliferate at a slow rate. Constitutive Akt activation does not affect p21 expression but appears to influence directly cdk/cyclin activity. On the other hand, the induction of differentiation‐related genes is unchanged in myr‐Akt cells. These results identify Akt downregulation as an important aspect of the response of chondrocytes to FGF that, however, only affects chondrocyte proliferation and not the ability of FGF to induce differentiation genes. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Functions of human replication protein A (RPA): From DNA replication to DNA damage and stress responses
Authors: Zou Yue , Liu Yiyong , Wu Xiaoming , Shell Steven M.
Published: March 2006

Human replication protein A (RPA), a heterotrimeric protein complex, was originally defined as a eukaryotic single‐stranded DNA binding (SSB) protein essential for the in vitro replication of simian virus 40 (SV40) DNA. Since then RPA has been found to be an indispensable player in almost all DNA metabolic pathways such as, but not limited to, DNA replication, DNA repair, recombination, cell cycle, and DNA damage checkpoints. Defects in these cellular reactions may lead to genome instability and, thus, the diseases with a high potential to evolve into cancer. This extensive involvement of RPA in various cellular activities implies a potential modulatory role for RPA in cellular responses to genotoxic insults. In support, RPA is hyperphosphorylated upon DNA damage or replication stress by checkpoint kinases including ataxia telangiectasia mutated (ATM), ATR (ATM and Rad3‐related), and DNA‐dependent protein kinase (DNA‐PK). The hyperphosphorylation may change the functions of RPA and, thus, the activities of individual pathways in which it is involved. Indeed, there is growing evidence that hyperphosphorylation alters RPA–DNA and RPA–protein interactions. In addition, recent advances in understanding the molecular basis of the stress‐induced modulation of RPA functions demonstrate that RPA undergoes a subtle structural change upon hyperphosphorylation, revealing a structure‐based modulatory mechanism. Furthermore, given the crucial roles of RPA in a broad range of cellular processes, targeting RPA to inhibit its specific functions, particularly in DNA replication and repair, may serve a valuable strategy for drug development towards better cancer treatment. J. Cell. Physiol. 208: 267–273, 2006. © 2006 Wiley‐Liss, Inc.
Proteasome inhibitors shorten replicative life span and induce a senescent‐like phenotype of human fibroblasts
Authors: Torres Claudio , Lewis Lindsey , Cristofalo Vincent J.
Published: March 2006

The proteasome constitutes the main non‐lysosomal cellular protease activity, and plays a crucial role not only in the disposal of unwanted material, but also in the regulation of numerous cellular processes. Previously, we have reported that during the replicative senescence of WI‐38 fibroblasts there is a significant impairment in proteasome activity, which probably has important implications in the control of MAPK signaling and cellular proliferation. In this study, we report the potential role of the proteasome in the generation of the senescent phenotype in WI‐38 fibroblasts. Our results indicate that inhibition of proteasome activity leads to an impairment in cell proliferation, and a shortening of the life span. The results also indicate that inhibition of the proteasome in young cells induces a premature senescent‐like phenotype, as indicated by the increase in senescence‐associated β‐galactosidase (SA β‐gal) activity and the abundance of both p21 and collagenase mRNAs, as well as a decreased level of EPC‐1 mRNA known markers of cellular senescence, not previously shown to depend on proteasome activity. Together, our results suggest a molecular mechanism for the lack of responsiveness of human cells to growth factors, and point towards a role for the proteasome in the control of the life span of both cells and organisms. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Consequences of point mutations in trout anion exchanger 1 (tAE1) transmembrane domains: Evidence that tAE1 can behave as a chloride channel
Authors: Martial Sonia , Guizouarn Hélène , Gabillat Nicole , Pellissier Bernard , Borgese Franck
Published: February 2006

In this study, we have shown that, when expressed in Xenopus oocytes, trout anion exchanger 1 (tAE1) was able to act as a bifunctional protein, either an anion exchanger or a chloride conductance. Point mutations of tAE1 were carried out and their effect on Cl− conductance and Cl− unidirectional flux were studied. We have shown that mutations made in transmembrane domain 7 had dramatic effects on tAE1 function. Indeed, when these residues were mutated, either individually or together (mutants E632K, D633G, and ED/KG), Cl− conductance was reduced to 28–44% that of wild‐type tAE1. Moreover, ion substitution experiments showed that anion selectivity was altered. However, the exchanger function was unchanged, as evidenced by the fact that Cl− influx and Km were identical for each of these mutants and similar to the wild‐type protein parameters. By contrast, mutations made in the C‐terminal domains of the protein (R819M, Q829K) affected both transport functions. Cl− conductance was increased by ∼200% with respect to tAE1 and anion selectivity was impaired. Likewise, Cl− influx was increased by ∼260% and was no longer saturable. These and other mutations carried out in transmembrane domains 7, 8, 12–14 of tAE1 allow us to demonstrate without doubt that, in addition to its anion exchanger activity, tAE1 can also function as a chloride channel. Above all, this work led us to identify amino acids involved in this double function organization. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Activation of smooth muscle and myenteric plexus cells of jejunum via toll‐like receptor 4
Authors: Rumio Cristiano , Besusso Dario , Arnaboldi Francesca , Palazzo Marco , Selleri Silvia , Gariboldi Silvia , Akira Shizuo , Uematsu Satoshi , Bignami Paola , Ceriani Valerio , Ménard Sylvie , Balsari Andrea
Published: March 2006

The cell types of the gut expressing Toll‐like receptor 4, which recognizes specifically bacterial lipopolysaccharides, as well as the functionality of this receptor, have remained controversial. We aimed to clarify these issues. Mouse and human intestinal specimens were stained immunohistochemically to detect Toll‐like receptor 4 expression. Smooth muscle and myenteric plexus cells but not enterocytes revealed receptor expression. Murine intestinal smooth muscle and myenteric plexus cells but not enterocytes showed nuclear translocation of nuclear factor‐kappaB after in vivo stimulation with lipopolysaccharide. Moreover, lipopolysaccharide added to human jejunum biopsies free of epithelial cells induced release of interleukin‐8 (IL‐8). We can conclude that Toll‐like receptor 4 is not expressed in epithelial layer, but rather on smooth muscle and myenteric plexus cells and that expression is functional. The expression of Toll‐like receptor 4 on smooth muscle and myenteric plexus cells is consistent with the possibility that these cells are involved in intestinal immune defense; the low or absent expression of Toll‐like receptor 4 on enterocytes might explain the intestinal epithelium hyporesponsiveness to the abundance of LPS in the intestinal lumen. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Differential activity of the FGF‐4 enhancer in F9 and P19 embryonal carcinoma cells
Authors: Boer Brian , Bernadt Cory T. , Desler Michelle , Wilder Phillip J. , Kopp Janel L. , Rizzino Angie
Published: March 2006

Transcription factors Oct‐3/4 and Sox2 behave as global regulators during mammalian embryogenesis. They work together by binding co‐operatively to closely spaced HMG and POU motifs (HMG/POU cassettes). Recently, it was suggested that a critical Sox2:Oct‐3/4 target gene, FGF‐4, is expressed at lower levels in P19 than in F9 embryonal carcinoma (EC) cells, due to lower levels of Sox2 in P19 than in F9 cells. We tested this possibility to better understand how FGF‐4 expression is modulated during development. Although we found that P19 EC cells express ∼10‐fold less FGF‐4 mRNA than F9 EC cells, we determined that Sox2 levels do not differ markedly in F9 and P19 EC cells. We also determined that Sox2 and Oct‐3/4 work together equally well in both EC cell lines. Moreover, in contrast to an earlier prediction based on in vitro binding studies, we demonstrate that the function of the HMG/POU cassettes of the FGF‐4 and UTF1 genes does not differ significantly in these EC cell lines when tested in the context of a natural enhancer. Importantly, we determined that the FGF‐4 promoter is highly responsive to a heterologous enhancer in both EC cell lines; whereas, the FGF‐4 enhancer is 7‐ to 10‐fold less active in P19 than in F9 EC cells. Because F9 and P19 EC cells are likely to represent cells at different stages of mammalian development, we suggest that this difference in FGF‐4 enhancer activity may reflect a mechanism used to decrease, but not abolish, FGF‐4 expression as the early embryo develops. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.
Impact of transgene expression on drug metabolism following systemic adenoviral vector administration
Authors: Callahan Shellie M. , Boquet Michael P. , Ming Xin , Brunner Lane J. , Croyle Maria A.
Published: February 2006

Background

Systemic administration of a first‐generation adenovirus expressing E. coli beta‐galactosidase (AdlacZ) alters expression and function of two hepatic drug‐metabolizing enzymes, cytochrome P450 (CYP) 3A2 and 2C11, for 14 days. The objective of these studies was to determine how the transgene cassette influences CYP expression and function.

Methods

Sprague‐Dawley rats were given 5.7 × 1012 viral particles (vp)/kg of either: AdlacZ, Ad expressing murine erythropoietin (Epo), Ad without a transgene (Null), or phosphate‐buffered saline (Vehicle). Hepatic CYP protein expression, activity, mRNA and alanine aminotransferase (ALT) levels were analyzed 0.25, 1, 4, and 14 days following a single intravenous injection.

Results

Administration of Epo did not alter CYP3A2 activity, but induced RNA levels by a factor of 2 at 4 and 14 days (P⩽0.01). This vector suppressed CYP2C11 activity levels by 45% at 1 day (P⩽0.05) and RNA levels throughout the study period (P⩽0.05). The Null vector suppressed CYP3A2 activity by 36, 63, 34, and 45% at 0.25, 1, 4 and 14 days, respectively (P⩽0.05). CYP2C11 activity was suppressed 1 day after administration (41%) and RNA levels were suppressed at 6 h (53%) and 1 day (36%, P⩽0.05). In contrast, AdlacZ suppressed both CYP3A2 and 2C11 at all time points.

Conclusions

The immunogenic and biological nature of the transgene cassette can influence changes in CYP3A2, but not the 2C11 isoform. The shift in transcription and translation of protein for maintenance of physiologic homeostasis to production of viral proteins and transgene product and their associated toxicity during viral infection may explain our observations. Copyright © 2006 John Wiley & Sons, Ltd.
α‐Melanocyte‐stimulating hormone gene therapy reverses carbon tetrachloride induced liver fibrosis in mice
Authors: Lee Tsung‐Hsing , Jawan Bruno , Chou Wen‐Ying , Lu Cheng‐Nan , Wu Chia‐Ling , Kuo Hsiao‐Mei , Concejero Allan M. , Wang Cheng‐Haung
Published: February 2006

Background

Hepatic fibrosis represents a process of healing and scarring in response to chronic liver injury. Effective therapies are lacking. We have previously demonstrated that α‐melanocyte‐stimulating hormone (α‐MSH) gene therapy protects against thioacetamide‐induced acute liver failure in mice. Recent reports showed that collagen metabolism is a novel target of α‐MSH. Therefore, the aim of this study is to investigate whether α‐MSH gene therapy possesses anti‐hepatic fibrogenic effect in mice.

Methods

Liver fibrosis was induced in mice by administering carbon tetrachloride (CCl4) continuously for 10 weeks. α‐MSH expression plasmid was delivered via electroporation after liver fibrosis had been established. Histopathology, reverse‐transcription polymerase chain reaction (RT‐PCR), immunoblotting, and gelatin zymography were used to investigate its possible mechanisms of action.

Results

α‐MSH gene therapy reversed established liver fibrosis in CCl4‐treated mice. RT‐PCR revealed that α‐MSH gene therapy attenuated the liver TGF‐β1, collagen α1, and cell adhesion molecule mRNA upregulation. Following gene transfer, both the activation of α‐smooth muscle actin (α‐SMA) and cyclooxygenase‐2 (COX‐2) was significantly attenuated. Further, α‐MSH significantly increased matrix metalloproteinase (MMP) activity with tissue inhibitors of matrix metalloproteinase (TIMP) inactivation.

Conclusions

We have demonstrated that α‐MSH gene therapy reversed established liver fibrosis in mice. It also prevented the upregulated fibrogenic and proinflammatory gene response after CCl4administration. Its collagenolytic effect may be attributed to MMP and TIMP modulation. In summary, α‐MSH gene therapy may be an effective therapeutic modality against liver fibrosis with potential clinical use. Copyright © 2006 John Wiley & Sons, Ltd.
Synthesis of fluorine‐18‐labelled 5‐ and 6‐fluoro‐2‐pyridinamine
Authors: Abrahim Aiman , Angelberger Peter , Kletter Kurt , Müller Markus , Joukhadar Christian , Erker Thomas , Langer Oliver
Published: March 2006

A one‐pot radiosynthesis method to prepare the new fluorine‐18‐labelled fluoropyridine derivatives 5‐[18F]fluoro‐2‐pyridinamine and 6‐[18F]fluoro‐2‐pyridinamine in two to three reaction steps was developed. The first step consisted of no‐carrier‐added nucleophilic aromatic substitution of commercially available halogen‐substituted 2‐pyridinecarboxamide or 2‐pyridinecarbonitrile derivatives with K[18F]F‐K222 in DMSO at 150–180°C. The [18F]fluoride incorporation yields ranged from 67 to 98% for all studied precursor molecules. It is remarkable that 5‐bromo‐2‐pyridinecarbonitrile gave almost quantitative [18F]fluoride incorporation at the meta‐position (5‐position) of the pyridine ring after only 5 min of heating at 150°C. After base‐catalysed hydrolysis of the [18F]fluorinated pyridinecarbonitriles into their corresponding carboxamides, the latter were transformed in a Hofmann‐type rearrangement reaction into the respective amines by treatment of crude reaction mixtures with bromine and aqueous base (20–30% conversion yield). Reaction mixtures were purified by reversed‐phase semipreparative HPLC followed by strong cation exchange solid‐phase extraction to afford 5‐[18F]fluoro‐2‐pyridinamine and 6‐[18F]fluoro‐2‐pyridinamine in non‐decay‐corrected radiochemical yields of 6–10% in a total synthesis time of 83–112 min. The preparation of 5‐[18F]fluoro‐2‐pyridinamine is one of very few examples demonstrating the feasibility of nucleophilic meta‐[18F]fluorination of a pyridine derivative. Both 5‐[18F]fluoro‐2‐pyridinamine and 6‐[18F]fluoro‐2‐pyridinamine are new potentially useful radiolabelled synthons for radiopharmaceutical chemistry. Copyright © 2006 John Wiley & Sons, Ltd.
Local staging of rectal cancer using combined pelvic phased‐array and endorectal coil MRI
Authors: Tatli Servet , Mortele Koenraad J. , Breen Elizabeth L. , Bleday Ronald , Silverman Stuart G.
Published: March 2006

Purpose

To assess the accuracy of MRI, using a pelvic phased‐array coil and an endorectal coil, for preoperative local staging of rectal cancer.

Materials and Methods

Fifty‐one patients (26 males and 25 females) with adenocarcinoma of the rectum underwent preoperative MRI and surgical resection of their tumors. Surgical pathology staging was compared to MRI staging (using the TNM classification) obtained both retrospectively by a reader blinded to surgical findings and prospectively (radiological reports). In addition, patients were stratified according to surgical treatment groups (stage I = T1‐2/N0, stage II = T3/N0, stage III = Tx/N1‐2).

Results

At pathology, 36 of 51 (68%) tumors were classified as T0‐T2, and 15 (32%) were classified as T3. Overall, the sensitivity and specificity of MRI readings for T3 staging were 93% and 86%, respectively (positive predictive value (PPV) = 74%, negative predictive value (NPV) = 97%, accuracy = 88%). MRI correctly predicted lymph node metastases in 11 of 13 patients with a sensitivity of 85% and specificity of 69% (PPV = 58%, NPV = 90%, accuracy = 74%). MRI correctly predicted surgical treatment groups in 33 of 39 (85%) patients. Interobserver agreement between the retrospective and prospective readings was excellent (κ = 0.85) for prediction of T3 tumor and good (κ = 0.80) for prediction of nodal involvement.

Conclusion

Combined endorectal and pelvic phased‐array coil MRI can be used reliably to select which patients should receive preoperative chemoradiotherapy. It is highly predictive in terms of excluding T3 tumors, but still has limitations in predicting lymph node metastasis. J. Magn. Reson. Imaging 2006. © 2006 Wiley‐Liss, Inc.
T1ρ relaxation mapping in human osteoarthritis (OA) cartilage: Comparison of T1ρ with T2
Authors: Regatte Ravinder R. , Akella Sarma V.S. , Lonner J.H. , Kneeland J.B. , Reddy Ravinder
Published: March 2006

Purpose

To quantify the spin‐lattice relaxation time in the rotating frame (T1ρ) in various clinical grades of human osteoarthritis (OA) cartilage specimens obtained from total knee replacement surgery, and to correlate the T1ρ with OA disease progression and compare it with the transverse relaxation time (T2).

Materials and Methods

Human cartilage specimens were obtained from consenting patients (N = 8) who underwent total replacement of the knee joint at the Pennsylvania Hospital, Philadelphia, PA, USA. T2‐ and T1ρ‐weighted images were obtained on a 4.0 Tesla whole‐body GE Signa scanner (GEMS, Milwaukee, WI, USA). A 7‐cm diameter transmit/receive quadrature birdcage coil tuned to 170 MHz was employed.

Results

All of the surgical knee replacement OA cartilage specimens showed elevated relaxation times (T2 and T1ρ) compared to healthy cartilage tissue. In various grades of OA specimens, the T1ρ relaxation times varied from 62 ± 5 msec to 100 ± 8 msec (mean ± SEM) depending on the degree of cartilage degeneration. However, T2 relaxation times varied only from 32 ± 2 msec to 45 ± 4 msec (mean ± SEM) on the same cartilage specimens. The increase in T2 and T1ρ in various clinical grades of OA specimens were ∼5–50% and 30–120%, respectively, compared to healthy specimens. The degenerative status of the cartilage specimens was also confirmed by histological evaluation.

Conclusion

Preliminary results from a limited number of knee specimens (N = 8) suggest that T1ρ relaxation mapping is a sensitive noninvasive marker for quantitatively predicting and monitoring the status of macromolecules in early OA. Furthermore, T1ρ has a higher dynamic range (>100%) for detecting early pathology compared to T2. This higher dynamic range can be exploited to measure even small macromolecular changes with greater accuracy compared to T2. Because of these advantages, T1ρ relaxation mapping may be useful for evaluating early OA therapy. J. Magn. Reson. Imaging 2006. © 2006 Wiley‐Liss, Inc.
Interactive magnetic resonance cholangiography (MRC) with adaptive averaging
Authors: Graves Martin J. , Kershaw Lucy E. , Black Richard T. , Sala Evis , Skinner Jane , Lomas David J.
Published: March 2006

Purpose

To implement and evaluate a technique for adaptively averaging 2D magnetic resonance cholangiography (MRC) images obtained using an interactive imaging system with a view to improving image quality at reduced fields of view (FOVs).

Materials and Methods

Images were obtained using an interactive implementation of a single‐shot half‐Fourier rapid acquisition with relaxation enhancement (RARE) technique. Software was developed for adaptively averaging images, and an evaluation was performed in a phantom and a cohort of 10 patients referred for standard MRC. Adaptively averaged and standard single‐shot MRC images were evaluated with respect to their ability to demonstrate the common bile duct and main left and right intrahepatic duct branches.

Results

In all patient studies there was no difference in the ability of either the adaptive technique or the standard single‐shot method to demonstrate the common bile duct and the main left and right intrahepatic duct branches. However, in seven of the 10 patient studies the adaptive technique provided better visualization of the peripheral bile duct system (P = 0.035; sign test). There was no difference in the diagnostic confidence of the two techniques (P = 0.32, Wilcoxon signed‐rank test).

Conclusion

Adaptive averaging of MRC images obtained using an interactive imaging paradigm significantly improves visualization of peripheral intrahepatic ducts. J. Magn. Reson. Imaging 2006. © 2006 Wiley‐Liss, Inc.
Hemodynamic characterization of focal hepatic lesions: Role of ferucarbotran‐enhanced dynamic MR imaging using T2‐weighted multishot spin‐echo echo‐planar sequence
Authors: Hori Masatoshi , Murakami Takamichi , Kim Tonsok , Iannaccone Riccardo , Abe Hisashi , Onishi Hiromitsu , Tomoda Kaname , Catalano Carlo , Passariello Roberto , Nakamura Hironobu
Published: March 2006

Purpose

To investigate the role of ferucarbotran‐enhanced dynamic MR imaging using multishot spin‐echo echo‐planar sequence in the evaluation of hemodynamics of focal hepatic lesions.

Materials and Methods

Sixty‐three focal hepatic lesions (24 benign and 39 malignant) from 53 consecutive patients who underwent both ferucarbotran‐enhanced MR imaging and dynamic computed tomography (CT) were included in this study. MR imaging was performed with a 1.5‐T scanner with a phased‐array coil. T2‐weighted multishot spin‐echo echo‐planar sequences (TR/TE = 1714–2813/80 msec) were obtained during a single breathhold before and 15, 60, 120, 180, and 600 seconds after intravenous injection of ferucarbotran. The enhancement patterns of lesions were classified into three categories by a study coordinator on the basis of dynamic CT images as hypervascular, hypovascular, and hemangioma type. The study coordinator created mean contrast‐to‐noise ratio of lesions vs. time curves for each enhancement pattern for quantitative analyses. Moreover, three radiologists separately and blindly reviewed MR images, and then assigned three confidence scores for the three enhancement patterns to each lesion. Sensitivity, specificity, and receiver operating characteristic analyses were performed.

Results

Quantitative analyses showed characteristic enhancement curves for each enhancement pattern. Mean sensitivities/specificities were 0.816/0.882, 0.897/0.863, and 0.800/0.989 for hypervascular, hypovascular, and hemangioma types, respectively. Mean areas under the receiver operating characteristic curve were 0.886 for hypervascular type and 0.913 for hypovascular type.

Conclusion

Ferucarbotran‐enhanced dynamic MR imaging can be used to successfully characterize the hemodynamics of focal hepatic lesions. J. Magn. Reson. Imaging 2006. © 2006 Wiley‐Liss, Inc.