bluuebunny/crossref_metadata_2025 huggingface dataset:
MOCVD Growth of Transparent Conducting Cd2SnO4 Thin Films
Authors: Metz A.W. , Lane M.A. , Kannewurt C.R. , Poeppelmeier K.R. , Marks T.J.
Published: December 2004

The first preparation of transparent conducting Cd2SnO4 thin films by a simple MOCVD process is described. As‐deposited films using Cd(hfa)2(TMEDA) (Figure), at 365  °C are found to be highly crystalline with a relatively wide range of grain size of 100–300  nm. XRD indicates a cubic spinel Cd2SnO4 crystal structure and the possible presence of a small amount of CdO. The films exhibit conductivities of 2170 S/cm and a bandgap of 3.3 eV, rivaling those of commercial tin‐doped indium oxide.
MOCVD of KNbO3 Ferroelectric Films and their Characterization
Authors: Romanov M.V. , Korsakov I.E. , Kaul A.R. , Stefanovich S.Yu. , Bolshakov I.A. , Wahl G.
Published: December 2004

KNbO3 in the form of films is a highly acclaimed material due to its potential application in surface acoustic wave (SAW), and nonlinear optic devices. Single‐source powder flash evaporation MOCVD of epitaxial KNbO3 films was accomplished, for the first time, with potassium tert‐butoxide and niobium heteroligand complex, Nb(OiPr)4(thd) used as volatile metal–organic precursors. The microstructure of the films was found to be dependent on the substrate used (MgO or SrTiO3) and deposition temperature. A new approach to reach cation stoichiometry of deposited films deficient in potassium, consisting of a post‐deposition annealing with a KNbO3/K3NbO4 powder mixture, was proposed. The device quality of the films was verified by high second harmonic generation (SHG) output. The effect of the oxygen non‐stoichiometry of films on the phase transition temperature was proven.
UHV Surface Chemistry of bis(ethylcyclopentadienyl)ruthenium, (C2H5C5H4)2Ru on an Oxide Substrate
Authors: Luo B. , Wang Q. , White J.M.
Published: December 2004

The interactions of an organometallic precursor to Ru thin films, bis(ethylcyclopentadienyl)ruthenium, (C2H5C5H4)2Ru, with an amorphous Al2O3 substrate were probed using tools of ultra‐high vacuum surface science. The precursor adsorbs and desorbs with no decomposition on Al2O3 that is bare, or covered with patches of O‐saturated Ru. In the absence of pre‐existing Ru sites, there was no reaction up to 950 K with or without co‐dosed O2 at pressures up to 10–6 torr. Film‐forming reactions do occur under UHV dosing conditions when Ru sites and chemisorbed oxygen are both present. The distribution of chemical states of surface oxygen changes with temperature and influences reaction kinetics and film growth.
Leading the way to double‐digit growth
Authors: Gandossy Robert P. , Greenslade Shelli
Published: December 2004

The five strategies leaders in fast‐growth companies pursue.
On‐line waiting: The role of download time and other important predictors on attitude toward e‐retailers
Authors: Rose Gregory M. , Meuter Matthew L. , Curran James M.
Published: December 2004

Across both marketing and management‐information‐system disciplines, download time has been identified as an important factor for on‐line business success. This research is intended to clarify counterintuitive findings generated from previous research and Web‐based experimentation. Here the roles of several factors, including download time, on the overall evaluation of an e‐retailer are explored. With an experimental design, the article explores the impact of 5‐, 30‐, and 45‐second download delays on attitude toward an e‐retailer. Consistent with prior works, findings from both a main study and replication study indicate that objective download delay is not a critical determinant of attitude toward an e‐retailer. However, findings show there are other direct and indirect delay effects, including those resulting from perceived retailer control of delay, that impact Web‐system success. © 2005 Wiley Periodicals, Inc.
Structural relaxation and evolution of yield stress in epoxy glass aged under shear strain
Authors: Kawakami Hiroshi , Otsuki Ryohei , Nanzai Yukuo
Published: December 2004

Hollow cylindrical specimens of annealed epoxy glass were twisted and then aged for various periods of time under shear strain. At the end of the aging process, we twisted the specimens again to determine the stress–strain relations. For specimens aged under a shear strain of 0.005 or 0.01, the stress relaxation behavior was almost independent of the amount of strain imposed, and the value of stress at the upper yield point, regardless of aging time, was almost the same as that of the annealed specimen. On the other hand, for specimens aged under a strain of 0.02 or 0.04, the stress relaxation behavior depended on the value of the strain applied, and the value of stress at the upper yield point first decreased and subsequently increased with increasing aging time. These results led us to the following conclusions: If epoxy glass is strained largely, the originally stable structure becomes unstable. Also, when epoxy glass is aged under strain, the stability of the structure continues to decrease for a short period of time after deformation ceases, and then increases with increasing aging time. POLYM. ENG. SCI. 45:20–24, 2005. © 2004 Society of Plastics Engineers.
Experimental study on the flow and deposition of powder particles in rotational molding
Authors: Olinek J. , Anand C. , Bellehumeur C.T.
Published: December 2004

The nature of powder flow and its effect on particle deposition in rotationally molded parts were considered in this work. Experiments were carried out to observe the effects of various parameters, such as particle characteristics and operating conditions, on the deposition patterns of polyethylene powders and micropellets. The results indicate that the polymeric powders were cohesive enough to prevent size segregation at ambient temperature; however, segregation occurred when particles that had a smooth surface and regular shape were used. During processing, however, a new phenomenon of reverse cohesive segregation was observed. The results showed that the final deposition patterns are controlled primarily by the initial segregation patterns, as well as by the heating rate and rotation speed, which affect the evolution of adhesive forces between particles during heating and melt deposition process. An order‐of‐magnitude analysis was conducted to evaluate the development of cohesive forces between particles, and to estimate their effects on the movement of particles. This study provides a better understanding of the flow characteristics of polymer particles during the rotational molding process, which is very important in the development of techniques for fabricating composites and multilayered products. Polym. Eng. Sci. 45:62–73, 2005. © 2004 Society of Plastics Engineers.
Preparation and characterization of poly(styrene‐co‐butadiene) and polybutadiene rubber/clay nanocomposites
Authors: Song M , Wong CW , Jin J , Ansarifar A , Zhang ZY , Richardson M
Published: December 2004

Poly(styrene‐co‐butadiene) rubber (SBR) and polybutadiene rubber (BR)/clay nanocomposites have been prepared. The effects of the incorporation of inorganically and organically modified clays on the vulcanization reactions of SBR and BR were analysed by rheometry and differential scanning calorimetry. A reduction in scorch time (ts1) and optimum time (t95) was observed for both the rubbers when organoclay was added and this was attributed to the amine groups of the organic modifier. However, ts1 and t95 were further increased as the clay content was increased. A reduction in torque value was obtained for the organoclay nanocomposites, indicating a lower number of crosslinks formed. The organoclays favoured the vulcanization process although the vulcanizing effect was reduced with increasing clay content. The tensile strength and elongation of SBR were improved significantly with organoclay. The improvement of the tensile properties of BR with organoclay was less noticeable than inorganic‐modified clay. Nevertheless, these mechanical properties were enhanced with addition of clay. The mechanical properties of the nanocomposites were dependent on filler size and dispersion, and also compatibility between fillers and the rubber matrix. Copyright © 2004 Society of Chemical Industry
Relaxational behavior and swelling‐pH master curves of poly[(diethylaminoethyl methacrylate)‐graft‐(ethylene glycol)] hydrogels
Authors: Podual Kairali , Peppas Nicholas A.
Published: December 2004

Poly[(diethylaminoethyl methacrylate)‐graft‐(ethylene glycol)] hydrogels were prepared with a molar ratio of 10:1 of diethylaminoethyl methacrylate to poly(ethylene glycol) of number‐average molecular weights (Mn) 200, 400 and 1000 g mol−1 using tetra(ethylene glycol) dimethacrylate to give a crosslinking ratio between 0.5 and 4.0 %. Glucose oxidase and catalase were immobilized in the matrix during polymerization. The maximum enzyme loading used was 6.6 × 10−4 g of glucose oxidase per g of polymer. The equilibrium and dynamic swelling properties of these hydrogels were investigated. The pH‐dependent equilibrium swelling characteristics showed a sharp transition between the swollen and the collapsed state at pH 7.0. The dynamic response of the hydrogel discs to pH was analyzed in pulsatile pH conditions. The effects of particle size, crosslinking and molecular weight of poly(ethylene glycol) (PEG) on the dynamic swelling response were investigated. The pulsatile nature of the response was analyzed using Boltzmann superposition. Swelling–pH master curves were obtained. Copyright © 2004 Society of Chemical Industry
Peptide mass fingerprinting by matrix‐assisted laser desorption ionization mass spectrometry of proteins detected by immunostaining on nitrocellulose
Authors: Dufresne‐Martin Geneviève , Lemay Jean‐François , Lavigne Pierre , Klarskov Klaus
Published: December 2004

We have developed an approach that allows peptide mass mapping by matrix‐assisted laser desorption ionization‐mass spectrometry of proteins visualized on a nitrocellulose membrane by immunochemical detection. Proteins are separated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE), electroblotted onto a nitrocellulose membrane and after blocking with a nonprotein‐containing polymer such as polyvinylpyrrolidone 40 (PVP‐40) or Tween 20, the proteins are stained with fount India ink. After incubation with primary and, if required, secondary peroxidase‐coupled antibodies, immunochemically reactive proteins can be visualized using conventional enhanced chemiluminescence detection and assigned to the India ink‐stained membrane by simple superposition. The proteins of interest are excised, submitted to “on‐membrane” cleavage and the peptides are analyzed by mass spectrometry. Protein‐based blocking reagents normally used in standard immunodetection protocols, such as skimmed milk, can be employed. We have obtained high‐quality mass spectra of bovine serum albumin (BSA) detected on an immunoblot with an estimated amount of 100 fmol applied onto the gel, indicating the sensitivity of the present method. In addition, the approach is demonstrated with two other commercially available proteins, a serum protein, the successful identification of a tyrosine phosphorylated protein from total rat liver homogenate and serine phosphorylated proteins from an EcR 293 nuclear extract separated by two‐dimensional (2‐D) SDS‐PAGE.
Mass spectrometry and database search in the analysis of proteins from the fungus Pleurotus ostreatus
Authors: Matis Maja , Žakelj‐Mavrič Marija , Peter‐Katalinić Jasna
Published: December 2004

Tandem mass spectrometry is a method of choice for rapid analysis in proteomics. Identification and characterization of proteins from organisms with sequenced genomes is today a routine procedure as will be identification of proteins from organisms with unsequenced genomes with new developing tools. Here, we report the use of isotopic labeling with electrospray ionisation (ESI)‐tandem mass spectrometry for de novo sequencing in combination with database search taking advantage of different programs for identification of fungal proteins. Using this approach we could identify the proteins of interest. Nevertheless, the identification of a novel protein responsible for the conversion of testosterone into androstenedione was still a difficult task, mostly due to the low homology of steroid transforming enzymes, especially those from microorganisms. Protein p27 was identified as the vanillate O‐demethylase oxidoreductase, p33 and p36 as two isoenzymes of malate dehydrogenase, and p45 as citrate synthase. By rechecking the sequences using additional programs it could be shown that the protein p36 has a higher local homology to the steroid‐transforming enzyme than to the malate dehydrogenase. Therefore, we assume that p36 is a pluripotent enzyme most probably responsible for the 17β‐hydroxysteroid dehydrogenase activity.
Kinetic and thermodynamic study of the reaction of pyridoxal 5′‐phosphate with L‐tryptophan
Authors: Echevarría Gerardo R. , Santos José G. , Basagoitia Andrea , Blanco Francisco García
Published: December 2004

The apparent rate constants for formation (k1) and hydrolysis (k2) of the Schiff bases formed by reaction of pyridoxal 5′‐phosphate with L‐tryptophan were determined at various pH values, at different temperatures and at constant ionic strength (0.1 M). Also obtained were the elementary rate constants for formation and hydrolysis of the Schiff bases corresponding to the different chemical species present in the media, and the pK values of the Schiff's bases. The activation and thermodynamic parameters for the formation and hydrolysis of the Schiff's bases also were determined. Some of the ΔH0 and ΔS0 values for the individual processes were found to be positive. In basic media the enthalpic factor is unfavorable but the entropic contribution leads to a negative ΔG0. Copyright © 2004 John Wiley & Sons, Ltd.
Gloucester Fishery: insights from a group modeling intervention
Authors: Otto Peter , Struben Jeroen
Published: December 2004

System dynamics provides a powerful support mechanism for resolving problems in highly complex and dynamic contexts. Effectively building a system dynamics model in a client‐group environment is a challenging task, particularly with a heterogeneous clientele and a variety of stakeholders inﬂuencing the boundaries of the model. In such environments, understanding and formulating central concerns is as important as it is challenging. This article describes a group model building initiative designed to study the implications of various policies aimed at revitalizing the ﬁshing industry in Gloucester, Massachusetts. While the iterative and interactive methods in this group modeling intervention helped ﬁshermen and community members to communicate effectively with one another, deﬁne problems and improve their understanding of the critical interactions in the system, the resulting model was used to help the Gloucester Community Development Cooperation to communicate potential impacts of a prospective ﬁsh factory to a wider audience. Copyright © 2004 John Wiley & Sons, Ltd.
Common ground for institutional economics and system dynamics modeling
Authors: Atkinson Glen
Published: December 2004

Institutional economics traces its roots to the works, beginning in the late nineteenth century, of Thorstein Veblen, John R. Commons and Wesley C. Mitchell. They believed that orthodox economic theory, based on deduction from axioms, was not a proper foundation to study the economy. They attempted to establish relations between economic actors as deﬁned by important economic institutions. Classical theory, on the other hand, is based on natural law rather than human organization. Natural law provided a ﬁxed structure, and this reduced uncertainty in economic theory. Change was within the structure but the structure was always stable. Institutional economists examine institutions that provide economic order, and they study the endogenous forces that cause these institutions to evolve. The author suggests that these are some of the same elements that describe system dynamics. Copyright © 2004 John Wiley & Sons, Ltd.
Upregulation of DNA repair genes in active cirrhosis associated with hepatocellular carcinoma
Authors: Zindy Pierre , Andrieux Lise , Bonnier Dominique , Musso Orlando , Langouët Sophie , Campion Jean Pierre , Turlin Bruno , Clément Bruno , Théret Nathalie
Published: December 2004

Phenotypic changes in injured livers involve complex network of genes whose interplays may lead to fibrosis and cirrhosis, a major risk of hepatocellular carcinoma. Gene expression profiles in fibrotic livers were analyzed by using cDNA microarray, hierarchical clustering and gene ontology. Analyses of a major cluster of upregulated genes in cirrhosis identified a new set of genes involved in DNA repair and damage. The upregulation of DNA repair genes was confirmed by real‐time quantitative polymerase chain reaction and associated with necroinflammatory activity (P
< 0.001). Increased DNA repair activity in cirrhosis with inflammatory activity may reflect increased DNA damages as a consequence of chronic liver injury.
NSPc1, a mainly nuclear localized protein of novel PcG family members, has a transcription repression activity related to its PKC phosphorylation site at S183
Authors: Gong Yanhua , Wang Xu , Liu Jin , Shi Lei , Yin Bin , Peng Xiaozhong , Qiang Boqin , Yuan Jiangang
Published: December 2004

Nervous system polycomb 1 (NSPc1) shares high homology with vertebrate PcG proteins Mel‐18 and Bmi‐1. The mRNA of NSPc1 is highly expressed in the developmental nervous system [Mech. Dev. 102 (2001) 219–222]. However, the functional characterization of NSPc1 protein is not clear. In the present study, using Western blotting technique, we aimed to describe the distributions of NSPc1 protein in rat tissues and cell lines. The subcellular localization of NSPc1 was examined in HeLa and SH‐SY5Ycell lines, and its transcriptional repression activity was examined in COS‐7 cell line. We found that the NSPc1 protein was localized mainly in the nucleus. NSPc1 remarkably repressed the transcription. Most interestingly, both the C‐terminal of NSPc1 and two phosphorylation sites in the C‐terminal, especially the PKC phosphorylation site at S183, were important in mediating transcription repression. Taken together, results from our study suggest that NSPc1, as a typical PcG family member, has powerful transcriptional repression ability, which may be related to the PKC signaling pathway.
Activation of tyrosine kinase of EGFR induces Gβγ‐dependent GRK–EGFR complex formation
Authors: Gao Jingxia , Li Jiali , Chen Yuejun , Ma Lan
Published: December 2004

This study demonstrated that activation of tyrosine kinase of epidermal growth factor receptor (EGFR) induces its association with G protein‐coupled receptor kinase 2 (GRK2). Immunoprecipitation experiments showed that EGF stimulation increased GRK2 binding to EGFR complex in HEK293 cells coexpressing EGFR and GRK2. The EGF‐induced GRK2–EGFR complex formation was greatly reduced by perturbation of EGFR and Src tyrosine kinase activity. Furthermore, studies with GRK2 mutants showed that neither catalytic activity nor the N‐terminal domain of GRK2 was required for EGF‐induced GRK2–EGFR complex formation. However, overexpression of Gβγ scavengers blocked EGF‐induced formation of GRK2–EGFR complex.
Modelling fishing location choice within mixed fisheries: English North Sea beam trawlers in 2000 and 2001
Authors: Hutton Trevor , Mardle Simon , Pascoe Sean , Clark Robin A.
Published: December 2004

Numerous studies have proposed methodologies to model fisher behaviour with the aim of predicting the outcomes of decision-making on board a fishing vessel. Both short- and long-term processes (e.g. investment) impact fleet dynamics. The proposed structure of the models has tended to depend upon the nature of the fishery and the control variables (technical restrictions, quotas, effort control, and/or closed areas). For example, within the context of multi-stock, multi-fleet fisheries (mixed fisheries), a skipper will allocate effort (as input to the production process) to harvest a range of species. Spatial complexity is normally excluded in models of behaviour. In this paper, two spatial analyses are presented for modelling location choice: an analysis based on a random utility model (RUM), and a simplified simulation model of individual vessels that depends on the results of the RUM. These models are applied to the English beam-trawl fleet operating in the North Sea in 2000. The results from the RUM indicate that the number of trips, the average trip length, and the average effort in each ICES rectangle are significant variables affecting location choice, in addition to catch rate for the previous year (1999), weighted by value. The last result is used as an assumption in a simulation model of fishing effort, i.e. fishers make decisions on spatial location of operation on the basis of past catch rates. The simulation model is used to predict the distribution of the same fleet for one month during the temporary closure in the North Sea in 2001. The predicted values for effort relate well to the fishing patterns observed.
Management of vesico‐vaginal fistulas in women
Authors: Ahmad S. , Nishtar A. , Hafeez G.A. , Khan Z.
Published: December 2004

Objective

Urinary fistulas resulting from obstetric trauma remains a major problem in developing countries. Surgical success rates, however, range from 61% to 95%. This study reviews the main causes of fistulas, and the surgical techniques and success rates of surgical closure.

Method

A total of 1086 cases of urinary fistulas resulting from obstetric trauma over 25 years were reviewed. All women received surgical intervention. Various types of fistulas as well as specific surgical techniques are described, and patient age, parity, and probable cause of fistula were compared with findings from studies in Africa.

Results

The highest number of cases (47.6%) occurred in women aged between 26 and 35 years; moreover, 54.43% of the women were para 6 and higher. Obstructed labor contributed to 72.78%, cesarean hysterectomy to 7.60%, cesarean section to 7.30%, and other obstetrical traumatic procedures to 12.28% of the cases. Surgical success was achieved in 84.5% of the cases. The major residual problem of stress incontinence was noted in 5% of the cases.

Conclusion

Besides prevention, the basic principles of fistula repair are the most important factors in achieving surgical success. Better preoperative preparation and rigorous measures for preventing postoperative infection can help increase the success rate of fistula repair.
Dysmenorrhea in Japanese women
Authors: Osuga Y. , Hayashi K. , Kobayashi Y. , Tokyokawa S. , Momoeda M. , Koga K. , Yoshino O. , Tsutsumi O. , Hoshiai H. , Terakawa N. , Taketani Y.
Published: December 2004

A third of Japanese women suffer from dysmenorrhea that requires analgesics, and the prevalence of dysmenorrhea generally shows inverse relationships with parity and age.
l‐Arginine treatment for asymmetric fetal growth restriction
Authors: Xiao X.M. , Li L.P.
Published: December 2004

Objectivities

To investigate the effects of l‐Arginine in treating asymmetric fetal growth restriction (FGR).

Methods

A total of 66 pregnant women whose fetuses were diagnosed with asymmetric fetal growth restriction were divided into two groups. Group 1 consisted of 36 women who were given routine therapy alone; group 2 consisted of 30 women who were given l‐Arginine and routine therapy; and the control group consisted of 30 more women with a normal pregnancy.

Results

Before treatment, mean maternal serum levels of NO2−/NO3− were significantly lower in groups 1 and 2 than in the control group (P<0.01). After treatment, maternal serum levels of NO2−/NO3− were considerably higher in group 2 than in group 1 (P<0.01). Mean birth weight was significantly higher in group 2 than in group 1 (P<0.05), but still lower in group 2 than in the control group (P<0.01).

Conclusions

A deficiency in nitric oxide may play an important role in the development of asymmetric fetal growth restriction. l‐Arginine can be used to increase maternal NO2−/NO3− levels and newborn birth weight.
On characterizing the 4 C's in supply chain management
Authors: Lejeune Miguel A. , Yakova Nevena
Published: December 2004

Supply chain management is a field at the confluence of many other disciplines; it has been studied under a number of perspectives, which has played a role for the crossbreeding of the discipline. In this paper, we propose a typology of supply chain configurations that contributes to tie together terms that have been used disjointedly for describing various forms of supply chain. It also enables researchers to develop theories related to the identified typology, rather than continue expanding efforts on separate, unconnected studies. The typology is configurational, consistent, and based on the correspondence (Sheppard and Sherman, 1998) between Fiske's theory of relational forms (1990) and the concept of interdependence, analyzed in the context of supply chain management. Our framework encompasses four main configurations – the communicative, coordinated, collaborative, and co‐opetitive – and we refer to these as the 4 C's in supply chain management. We discuss the benefits of the proposed typology from managerial and research perspectives, and suggest some research extensions.
The impact of alignment between virtual enterprise and information technology on business performance in an agile manufacturing environment
Authors: Cao Qing , Dowlatshahi Shad
Published: December 2004

Manufacturing companies are facing rapid and unanticipated changes in their business environment. Agile manufacturing (AM) is a manufacturing paradigm that focuses on smaller scale, modular production facilities, and agile operations capable of dealing with turbulent and changing environments. From several enablers of AM, virtual enterprise (VE) and information technology (IT) were chosen. This empirical study explored the impact of the alignment between VE and IT on business performance in an AM setting. Extensive data collection strategy and several tests are used to establish the reliability and validity of the data collected. Extensive analyses of the data using structural equation molding were performed for five hypotheses. The results indicate that both VE and IT had positive influences on business performance. It was also established that the alignment between VE and IT had a positive impact on business performance. Further, it was shown that the impact of the alignment between VE and IT on business performance was more significant than the impact of VE and IT on business performance individually. In conclusion, the assessment of the results along with future research directions is provided.
Experimental studies on the effects of recombinant human matrix metalloproteinases on herniated disc tissues—how to facilitate the natural resorption process of herniated discs
Authors: Haro Hirotaka , Komori Hiromichi , Kato Tsuyoshi , Hara Yasushi , Tagawa Masahiro , Shinomiya Kenichi , Spengler Dan M.
Published: December 2004

Purpose. Recently, MMP‐7 and MMP‐3 have been found to play a crucial role in the natural resorption process of herniated discs. We therefore examined the role of these recombinant human matrix metalloproteinases (rh MMPs) in the treatment of herniated discs.

Methods. (a) Surgical samples of herniated disc were cultured in the presence or absence of rh MMPs, and wet weight was measured 24 h later. (b) The rh MMPs were administered into normal rabbit intervertebral discs, and after 1 week spine samples were stained with Safranin O. (c) The rh MMPs were administered into canine herniated discs in vivo. Myelography and MRI were performed prior to and 1 week after administration. Spine samples were examined histologically. Whole disc tissue was collected, total protein was extracted, and Western blot analysis was performed.

Results. (a) Proteoglycan degradation was found in MMP‐7, MMP‐3, and chymopapain‐treated samples. MMP‐7 and chymopapain‐treated samples displayed a significant loss in wet weight (p < 0.01). (b) Normal disc tissues after administration of rh MMP‐7, MMP‐3, and chymopapain showed an extensive loss of Safranin O staining. (c) The rh MMP‐7‐treated discs had a marked decrease in protruded herniation by MRI. Herniated discs after administration of MMP‐7 and chymopapain showed a significant decrease in protruded mass 7 days after administration compared with saline‐treated discs when evaluated by myelography (p <0.01). The rh MMP‐7‐treated discs displayed a clear loss of Safranin O staining in the nucleus pulposus. Proteoglycan expression was barely detectable in disc tissues after MMP‐7 administration, whereas obvious expression was obtained in saline‐treated or untreated disc tissues.

Conclusions. Exposure to rh MMP‐7 resulted in promising proteoglycan loss in human surgical samples, normal rabbit intervertebral discs, and natural canine herniated discs. Administration of rh MMP‐7 may facilitate the resorption process of herniated discs. © 2004 Orthopaedic Research Society. Published by Elsevier Ltd. All rights reserved.
A sensitivity analysis of the fed‐batch animal‐cell bioreactor with respect to some control parameters
Authors: Lavric Vasile , Ofiţeru Irina Dana , Woinaroschy Alexandru
Published: December 2004

Animal cell culture is widely used in the manufacture of valuable products, and this process is nowadays seeing a rapid expansion. The growth of animal cells is a complex process, because the cells are very sensitive to environmental changes (in, for example, nutrients, pH, temperature, oxygen and osmolarity) during this phase and to the toxic compounds produced by the cell itself. Ammonia and lactate are the two major waste materials of cell culture. They can have inhibitory effects on cell growth and product (monoclonal antibodies among others) formation. In order to model the behaviour of a fed‐batch animal cell bioreactor producing monoclonal antibodies, it is necessary to use a complex kinetic model with optimal operating patterns ensuring high productivities. Good knowledge of such domains of operating parameters, together with the understanding of the response of this rather complex system to small modifications in the working conditions, are essential for on‐line control to improve the quality of product and the yield of an animal cell culture. The present study focuses on the sensitivity analysis of a fed‐batch animal cell bioreactor with respect to some candidate control parameters (substrate set‐point concentrations, feeding time‐step patterns and concentration of feeding solutions), emphasizing the influence of these on the overall performance of the system.
Effect of oxidative stress on the production of recombinant human interferon‐γ in Escherichia coli
Authors: Mohammadian‐Mosaabadi Jafar , Naderi‐Manesh Hossein , Maghsoudi Nader , Khalilzadeh Rasoul , Shojaosadati Seyd Abbas , Ebrahimi Marzieh
Published: December 2004

To reduce the degradation pathway of rhIFN‐γ (recombinant human interferon‐γ), the susceptibility against oxidative stress during fermentation in Escherichia coli cells and in purification process were investigated. Fermentations of recombinant E. coli were performed at 5, 30 and 60% DO (dissolved oxygen) concentrations. The expressed rhIFN‐γ is purified in three‐step chromatography processes with 99.9% purity under reducing and non‐reducing conditions. Recombinant IFN‐γ production, carbonyl and dimeric contents and antiviral‐specific activity of purified protein were monitored. The increase in carbonyl content was taken to be an indicator of protein oxidation. DO concentration did not show any noticeable effect on rhIFN‐γ production. During fermentation, the carbonyl‐group content showed no significant increase at 5 and 30% DO but a 10‐fold increase at 60% DO concentration was observed. The antiviral‐specific activity of purified rhIFN‐γ was decreased 35 and 69% at 30 and 60% DO concentrations respectively. After the purification process, under non‐reducing conditions, the activity of purified protein decreased to 30% of its original value. The degree of dimeric forms of protein was found to depend on the O2 concentration during fermentation and purification.
Regulation of all members of the antizyme family by antizyme inhibitor
Authors: MANGOLD Ursula , LEBERER Ekkehard
Published: December 2004

ODC (ornithine decarboxylase) is the rate-limiting enzyme in polyamine biosynthesis. Polyamines are essential for cellular growth and differentiation but enhanced ODC activity is associated with cell transformation. Post-translationally, ODC is negatively regulated through members of the antizyme family. Antizymes inhibit ODC activity, promote ODC degradation through the 26 S proteasome and regulate polyamine transport. Besides the ubiquitously expressed antizymes 1 and 2, there is the tissue-specific antizyme 3 and an yet uncharacterized antizyme 4. Antizyme 1 has been shown to be negatively regulated through the AZI (antizyme inhibitor) that binds antizyme 1 with higher affinity compared with ODC. In the present study, we show by yeast two- and three-hybrid protein–protein interaction studies that AZI interacts with all members of the antizyme family and is capable of disrupting the interaction between each antizyme and ODC. In a yeast-based ODC complementation assay, we show that human ODC is able to complement fully the function of the yeast homologue of ODC. Co-expression of antizymes resulted in ODC inhibition and cessation of yeast growth. The antizyme-induced growth inhibition could be reversed by addition of putrescine or by the co-expression of AZI. The protein interactions could be confirmed by immunoprecipitation of the human ODC–antizyme 2–AZI complexes. In summary, we conclude that human AZI is capable of acting as a general inhibitor for all members of the antizyme family and that the previously not yet characterized antizyme 4 is capable of binding ODC and inhibiting its enzymic activity similar to the other members of the antizyme family.
Identification of a novel rat hepatic gene induced early by insulin, independently of glucose
Authors: COFFY Sandrine , DECAUX Jean-François , GIRARD Jean , de KEYZER Yves , ASFARI Maryam
Published: December 2004

We used mRNA differential display to identify new genes induced early after exposure to insulin. Our screening strategy was based on the comparison of gene expression during the time course of insulin induction in the liver of 12-day-old suckling rats both in vivo and in vitro. A novel, early induced transcript, EIIH, was identified that encodes a 353-amino-acid protein with several features suggesting that it may be secreted or bound to membranes. EIIH is also distantly related to a variety of LRR (leucine-rich repeat) proteins. Insulin treatment increased EIIH mRNA levels in the hepatocytes of suckling, fasted adult and STZ (streptozotocin)-treated diabetic rats, where insulin was required to maintain the basal level of EIIH expression. EIIH expression was induced during the suckling/weaning transition, and remained detectable thereafter. Tissue distribution analysis in adult rats revealed a pattern of expression mainly in the liver, intestine and islets of Langerhans, closely following that of the Glut2 (glucose transporter 2), suggesting that it may play a role in carbohydrate metabolism. EIIH may be a primary target of the transcriptional regulation by insulin, and may therefore constitute a new model to study the mechanisms by which insulin acts on gene transcription.
Molecular basis of pollen-related food allergy: identification of a second cross-reactive IgE epitope on Pru av 1, the major cherry (Prunus avium) allergen
Authors: WICHE Regina , GUBESCH Michaela , KÖNIG Herbert , FÖTISCH Kay , HOFFMANN Andreas , WANGORSCH Andrea , SCHEURER Stephan , VIETHS Stefan
Published: December 2004

Birch (Betula verrucosa) pollen-associated food allergy is a well-characterized syndrome, which is due to the cross-reactivity of IgE antibodies to homologous allergens in various foods. One crossreacting area on the major birch pollen allergen Bet v 1 and its homologue in cherry (Prunus avium) Pru av 1 has already been identified. This is the so-called ‘P-loop’ region, which encompasses amino acid residues around position 45 and is found on the two virtually identical tertiary protein structures. We tried to determine an additional IgE cross-reacting patch on Pru av 1 and Bet v 1. The putative IgE-binding region on Pru av 1 was localized with a mAb (monoclonal antibody) that was generated against Bet v 1, and cross-reacts with several Bet v 1 homologues in food and inhibits the binding of patients' IgE to Pru av 1. mAb reactivity pattern was analysed and amino acid positions 28 and 108 of Pru av 1 were selected and mutated by site-directed mutagenesis. The Pru av 1 mutants were produced as recombinant proteins and characterized for their folding, mAb- and IgE-binding capacity and allergenic potency with a cellular assay using the humanized rat basophilic leukaemia cell line RBL-25/30. Amino acid position 28 is involved in a second major IgE-binding region on Pru av 1 and probably on Bet v 1. The identification of this second major IgE-binding region is an essential prerequisite to understand the phenomenon of cross-reactivity and its clinical consequences, and to produce hypoallergenic proteins for an improved immunotherapy of type I allergy.
Differential induction of quinone reductase by phytoestrogens and protection against oestrogen-induced DNA damage
Authors: BIANCO Nicole R. , CHAPLIN Laura J. , MONTANO Monica M.
Published: December 2004

Quinone reductase (QR) is a phase II detoxification enzyme that plays an important role in detoxifying quinones and may help maintain the antioxidant function of the cell. We have previously observed that QR is up-regulated by anti-oestrogens, but not oestrogen, in breast cancer cells via ERβ (oestrogen receptor β) transactivation. Such QR induction appears to protect breast cells against oestrogen-induced oxidative DNA damage, most likely by reducing reactive oestrogen metabolites termed catecholestrogen-quinones back to the hydroxy-catecholestrogens which may be conjugated. We now report that the phytoestrogens biochanin A, genistein and resveratrol also up-regulate QR expression in breast cancer cells. We observe that regulation can occur at the transcriptional level, preferentially through ERβ transactivation at the electrophile response element of the QR gene promoter. By chromatin immunoprecipitation analysis, we show binding of ERα and ERβ to the QR promoter, with increased ERβ binding in the presence of resveratrol. Functional studies show that biochanin A and resveratrol, but not genistein, can significantly protect against oestrogen-induced oxidative DNA damage in breast cancer cells. Antisense technology was used to determine whether such protection was dependent on ERβ or QR. Our results with resveratrol are consistent with our hypothesis that the protective ability of resveratrol is partially dependent on the presence of ERβ and QR. In conclusion, we postulate that phytoestrogen-mediated induction of QR may represent an additional mechanism for breast cancer protection, although the effects may be specific for a given phytoestrogen.
Human cyclo-oxygenase-1 and an alternative splice variant: contrasts in expression of mRNA, protein and catalytic activities
Authors: SCHNEIDER Claus , BOEGLIN William E. , BRASH Alan R.
Published: December 2004

The two COX (cyclo-oxygenase) isoenzymes COX-1 and -2 catalyse the initial step in the conversion of arachidonic acid into PG (prostaglandin) hormones. The identification of an mRNA transcript encoding a splice variant of human COX-1 was reported more than a decade ago [Diaz, Reginato and Jimenez (1992) J. Biol. Chem. 267, 10816–10822], yet catalytic activity and tissue expression of the corresponding spliced protein remained uncharacterized. The splice variant lacks amino acids 396–432, corresponding to the last 37 amino acids of exon 9 of the gene encoding COX-1. These amino acids form a loop at one side of the peroxidase active site of the protein. We expressed the full-length and spliced COX-1 cDNAs in COS-7 and Sf9 insect cells, and determined the PG-forming activity using incubations with radiolabelled arachidonic acid and HPLC analyses. When expressed in either system, abundant PG formation was observed with the full-length COX-1, whereas the spliced protein did not form any detectable product. Peroxidase activity was readily detected in microsomes prepared from COS-7 cells transfected with COX-1 but not with the splice variant. In reverse transcriptase–PCR experiments, we detected the mRNA for the alternatively spliced and full-length COX-1 in human brain, tonsil and colon tissue, yet we were unable to detect expression of the spliced protein in the same tissues using immunoprecipitation and Western-blot analyses. We conclude that, whereas the mRNA transcript for the spliced COX-1 is present in various human tissues, the corresponding protein is either not formed or subject to rapid proteolytic degradation.
Apolipoprotein CI overexpression is not a relevant strategy to block cholesteryl ester transfer protein (CETP) activity in CETP transgenic mice
Authors: GAUTIER Thomas , MASSON David , JONG Miek C. , PAIS DE BARROS Jean-Paul , DUVERNEUIL Linda , LE GUERN Naig , DECKERT Valérie , DUMONT Laure , BATAILLE Amandine , ZAK Zoulika , JIANG Xian-Cheng , HAVEKES Louis M. , LAGROST Laurent
Published: December 2004

ApoCI (apolipoprotein CI) is a potent inhibitor of plasma CETP [CE (cholesteryl ester) transfer protein]. The relevance of apoCI overexpression as a method for CETP blockade in vivo was addressed in the present study in CETPTg/apoCITg mice (mice expressing both human CETP and apoCI). Despite a significant reduction in specific CETP activity in CETPTg/apoCITg mice compared with CETPTg mice [transgenic mouse to human CETP; 46.8±11.1 versus 101.8±25.7 pmol·h−1·(μg of plasma CETP)−1 respectively; P<0.05], apoCI overexpression increased both the CETP mass concentration (3-fold increase; P<0.05) and the hepatic CETP mRNA level (4-fold increase, P<0.005), leading to an increase in total plasma CE transfer activity (by 39%, P<0.05). The ratio of apoB-containing lipoprotein to HDL (high-density lipoprotein) CE was 10-fold higher in CETPTg/apoCITg mice than in apoCITg mice (P<0.0005). It is proposed that the increased CETP expression in CETPTg/apoCITg mice is a direct consequence of liver X receptor activation in response to the accumulation of cholesterol-rich apoB-containing lipoproteins. In support of the latter view, hepatic mRNA levels of other liver X receptor-responsive genes [ABCG5 (ATP-binding cassette transporter GS) and SREBP-1c (sterol-regulatory-binding protein-1c)] were higher in CETPTg/apoCITg mice compared with CETPTg mice. In conclusion, overexpression of apoCI, while producing a significant inhibitory effect on specific CETP activity, does not represent a suitable method for decreasing total CE transfer activity in CETPTg/apoCITg mice, owing to an hyperlipidaemia-mediated effect on CETP gene expression.
Enhancement of endotoxin neutralization by coupling of a C12-alkyl chain to a lactoferricin-derived peptide
Authors: ANDRÄ Jörg , LOHNER Karl , BLONDELLE Sylvie E. , JERALA Roman , MORIYON Ignacio , KOCH Michel H. J. , GARIDEL Patrick , BRANDENBURG Klaus
Published: December 2004

Antibacterial peptide acylation, which mimics the structure of the natural lipopeptide polymyxin B, increases antimicrobial and endotoxin-neutralizing activities. The interaction of the lactoferricin-derived peptide LF11 and its N-terminally acylated analogue, lauryl-LF11, with different chemotypes of bacterial lipopolysaccharide (LPS Re, Ra and smooth S form) was investigated by biophysical means and was related to the peptides' biological activities. Both peptides exhibit high antibacterial activity against the three strains of Salmonella enterica differing in the LPS chemotype. Lauryl-LF11 has one order of magnitude higher activity against Re-type, but activity against Ra- and S-type bacteria is comparable with that of LF11. The alkyl derivative peptide lauryl-LF11 shows a much stronger inhibition of the LPS-induced cytokine induction in human mononuclear cells than LF11. Although peptide–LPS interaction is essentially of electrostatic nature, the lauryl-modified peptide displays a strong hydrophobic component. Such a feature might then explain the fact that saturation of the peptide binding takes place at a much lower peptide/LPS ratio for LF11 than for lauryl-LF11, and that an overcompensation of the negative LPS backbone charges is observed for lauryl-LF11. The influence of LF11 on the gel-to-liquid-crystalline phase-transition of LPS is negligible for LPS Re, but clearly fluidizing for LPS Ra. In contrast, lauryl-LF11 causes a cholesterol-like effect in the two chemotypes, fluidizing in the gel and rigidifying of the hydrocarbon chains in the liquid-crystalline phase. Both peptides convert the mixed unilamellar/non-lamellar aggregate structure of lipid A, the ‘endotoxic principle’ of LPS, into a multilamellar one. These data contribute to the understanding of the mechanisms of the peptide-mediated neutralization of endotoxin and effect of lipid modification of peptides.
IAP proteins: sticking it to Smac
Authors: DUCKETT Colin S.
Published: December 2004

Dogma has it that suppression of the programmed cell death pathway by the IAP (inhibitor of apoptosis) proteins is achieved by their direct enzymic inhibition of the chief executioners of the apoptotic process, the caspases. In turn, the IAPs themselves can be neutralized by Smac/DIABLO (second mitochondrial activator of caspases/direct IAP binding protein with low pI), a protein which in healthy cells is thought to be sequestered in the mitochondria, but which, in response to apoptotic stimuli, is released from the mitochondria into the cytosol where it can bind to IAPs, displacing caspases and thus perpetuating the apoptotic signal. While this is an elegant and attractive model, recent studies have suggested that IAPs can also suppress apoptotic cell death independently of their ability to inhibit caspases, and two reports in this issue of the Biochemical Journal reach the interesting conclusion that the cytoprotective IAPs, ML-IAP (melanoma IAP) and ILP-2 (IAP-like protein 2), exert their effects not through direct caspase inhibition, but through the neutralization of Smac/DIABLO. The predicted outcome of these studies is a delicately controlled equilibrium between the activities of IAPs and Smac/DIABLO, leading to a dynamic regulation of the apoptotic threshold.