bluuebunny/crossref_metadata_2025 huggingface dataset:
Genomic and physiological studies of early cryptochrome 1 action demonstrate roles for auxin and gibberellin in the control of hypocotyl growth by blue light
Authors: Folta Kevin M. , Pontin Mariela A. , Karlin‐Neumann George , Bottini Rubén , Spalding Edgar P.
Published: October 2003

Blue light inhibits elongation of etiolated Arabidopsis thaliana hypocotyls during the first 30 min of irradiation by a mechanism that depends on the phototropin 1 (phot1) photoreceptor. The cryptochrome 1 (cry1) photoreceptor begins to exert control after 30 min. To identify genes responsible for the cry1 phase of growth inhibition, mRNA expression profiles of cry1 and wild‐type seedlings were compared using DNA microarrays. Of the roughly 420 genes found to be differentially expressed at the point of cry1 response incipience, approximately half were expressed higher and half lower in cry1 relative to the wild type. Many of the cry1‐dependent genes encoded kinases, transcription factors, cell cycle regulators, cell wall metabolism enzymes, gibberellic acid (GA) biosynthesis enzymes, and auxin response factors. High‐resolution growth studies supported the hypothesis that genes in the last two categories were indeed relevant to cry1‐mediated growth control. Inhibiting GA4 biosynthesis with a 3β‐hydroxylase inhibitor (Ca‐prohexadione) restored wild‐type response kinetics in cry1 and completely suppressed its long‐hypocotyl phenotype in blue light. Co‐treatment of cry1 seedlings with Ca‐prohexadione plus GA4 completely reversed the effects of the inhibitor, restoring the long‐hypocotyl phenotype typical of the mutant. Treatment of wild‐type seedlings with GA4 was not sufficient to phenocopy cry1 seedlings, but co‐treatment with IAA plus GA4 produced cry1‐like growth kinetics for a period of approximately 5 h. The genomic and physiological data together indicate that blue light acting through cry1 quickly affects the expression of many genes, a subset of which suppresses stem growth by repressing GA and auxin levels and/or sensitivity.
Vacuolar cation/H+ exchange, ion homeostasis, and leaf development are altered in a T‐DNA insertional mutant of AtNHX1, the Arabidopsis vacuolar Na+/H+ antiporter
Authors: Apse Maris P. , Sottosanto Jordan B. , Blumwald Eduardo
Published: October 2003

The function of vacuolar Na+/H+ antiporter(s) in plants has been studied primarily in the context of salinity tolerance. By facilitating the accumulation of Na+ away from the cytosol, plant cells can avert ion toxicity and also utilize vacuolar Na+ as osmoticum to maintain turgor. As many genes encoding these antiporters have been cloned from salt‐sensitive plants, it is likely that they function in some capacity other than salinity tolerance. The wide expression pattern of Arabidopsis thaliana sodium proton exchanger 1 (AtNHX1) in this study supports this hypothesis. Here, we report the isolation of a T‐DNA insertional mutant of AtNHX1, a vacuolar Na+/H+ antiporter in Arabidopsis. Vacuoles isolated from leaves of the nhx1 plants had a much lower Na+/H+ and K+/H+ exchange activity. nhx1 plants also showed an altered leaf development, with reduction in the frequency of large epidermal cells and a reduction in overall leaf area compared to wild‐type plants. The overexpression of AtNHX1 in the nhx1 background complemented these phenotypes. In the presence of NaCl, nhx1 seedling establishment was impaired. These results place AtNHX1 as the dominant K+ and Na+/H+ antiporter in leaf vacuoles in Arabidopsis and also suggest that its contribution to ion homeostasis is important for not only salinity tolerance but development as well.
Stress memory in plants: a negative regulation of stomatal response and transient induction of rd22 gene to light in abscisic acid‐entrained Arabidopsis plants
Authors: Goh Chang‐Hyo , Nam Hong Gil , Park Yu Shin
Published: October 2003

All organisms, including plants, perceive environmental stress, and they use this information to modify their behavior or development. Here, we demonstrate that Arabidopsis plants have memory functions related to repeated exposure to stressful concentrations of the phytohormone abscisic acid (ABA), which acts as a chemical signal. Repeated exposure of plants to ABA (40 µm for 2 h) impaired light‐induced stomatal opening or inhibited the response to a light stimulus after ABA‐entrainment under both dark/light cycle and continuous light. Moreover, there were transient expressions of the rd22 gene during the same periods under both the growing conditions. Such acquired information in ABA‐entrained plants produced a long‐term sensitization. When the time of light application was changed, a transient induction of the rd22 gene in plants after ABA‐entrainment indicated that these were light‐associated responses. These transient effects were also observed in kin1, rab18, and rd29B. The transient expression of AtNCED3, causing the accumulation of endogenous ABA, indicated a possible regulation by ABA‐dependent pathways in ABA‐entrained plants. An ABA immunoassay supported this hypothesis: ABA‐entrained plants showed a transient increase in endogenous ABA level from 220 to 250 pmol g−1 fresh mass at 1–2 h of the training period, whereas ABA‐deficient (aba2) mutants did not. Taking into account these results, we propose that plants have the ability to memorize stressful environmental experiences, and discuss the molecular events in ABA‐entrained plants.
Enhanced formation of flowers in salt‐stressed Arabidopsis after genetic engineering of the synthesis of glycine betaine
Authors: Sulpice Ronan , Tsukaya Hirokazu , Nonaka Hideko , Mustardy Laszlo , Chen Tony H. H. , Murata Norio
Published: October 2003

Previously, we showed that transformation with the codA gene for choline oxidase allows plants to synthesize glycine betaine (GB) and enhances their ability to tolerate various kinds of stress during germination and vegetative growth. In this study, we examined the tolerance of transformed plants to salt stress at the reproductive stage, which is the stage at which plants are most sensitive to environmental stress. Salt‐shock treatment of wild‐type plants for 3 days resulted in the abortion of flower buds and decreased the number of seeds per silique. These deleterious effects were clearly visible 6 days after the termination of salt‐shock treatment. Microscopic examination of floral structures revealed that salt stress inhibited the development of anthers, pistils, and petals. In particular, the production of pollen grains and ovules was dramatically inhibited. These effects of salt stress were significantly reduced by transformation with the codA gene, and our observations suggested that the enhanced tolerance of the transgenic plants was a result of the accumulation of GB in the reproductive organs. Indeed, levels of GB in flowers, siliques, and inflorescence apices were about five times higher than in leaves.
AtAGP30, an arabinogalactan‐protein in the cell walls of the primary root, plays a role in root regeneration and seed germination
Authors: Van Hengel Arjon J. , Roberts Keith
Published: October 2003

Arabinogalactan‐proteins (AGPs) are extracellular proteoglycans that are implicated in many plant growth and developmental processes, but in no case has a biological function been assigned to a particular AGP. AtAGP30 is a non‐classical AGP core protein from Arabidopsis that is expressed only in roots. Analysis of the corresponding mutant, agp30, has revealed that the wild‐type gene product is required in vitro for root regeneration and in planta for the timing of seed germination. The mutant shows a suppression of the abscisic acid (ABA)‐induced delay in germination and altered expression of some ABA‐regulated genes. This suggests that AtAGP30 functions in the ABA response. By analogy to proteoglycan‐mediated regulation of growth‐factor‐signalling pathways in animals, our data indicate that phytohormone activity in plants can be modulated by AGPs.
Reconstruction of the central carbon metabolism of Aspergillus niger
Authors: David Helga , Åkesson Mats , Nielsen Jens
Published: October 2003

The topology of central carbon metabolism of Aspergillus niger was identified and the metabolic network reconstructed, by integrating genomic, biochemical and physiological information available for this microorganism and other related fungi. The reconstructed network may serve as a valuable database for annotation of genes identified in future genome sequencing projects on aspergilli. Based on the metabolic reconstruction, a stoichiometric model was set up that includes 284 metabolites and 335 reactions, of which 268 represent biochemical conversions and 67 represent transport processes between the different intracellular compartments and between the cell and the extracellular medium. The stoichiometry of the metabolic reactions was used in combination with biosynthetic requirements for growth and pseudo‐steady state mass balances over intracellular metabolites for the quantification of metabolic fluxes using metabolite balancing. This framework was employed to perform an in silico characterisation of the phenotypic behaviour of A. niger grown on different carbon sources. The effects on growth of single reaction deletions were assessed and essential biochemical reactions were identified for different carbon sources. Furthermore, application of the stoichiometric model for assessing the metabolic capabilities of A. niger to produce metabolites was evaluated by using succinate production as a case study.
Deflavination and reconstitution of flavoproteins
Authors: Hefti Marco H. , Vervoort Jacques , van Berkel Willem J. H.
Published: October 2003

Flavoproteins are ubiquitous redox proteins that are involved in many biological processes. In the majority of flavoproteins, the flavin cofactor is tightly but noncovalently bound. Reversible dissociation of flavoproteins into apoprotein and flavin prosthetic group yields valuable insights in flavoprotein folding, function and mechanism. Replacement of the natural cofactor with artificial flavins has proved to be especially useful for the determination of the solvent accessibility, polarity, reaction stereochemistry and dynamic behaviour of flavoprotein active sites. In this review we summarize the advances made in the field of flavoprotein deflavination and reconstitution. Several sophisticated chromatographic procedures to either deflavinate or reconstitute the flavoprotein on a large scale are discussed. In a subset of flavoproteins, the flavin cofactor is covalently attached to the polypeptide chain. Studies from riboflavin‐deficient expression systems and site‐directed mutagenesis suggest that the flavinylation reaction is a post‐translational, rather than a cotranslational, process. These genetic approaches have also provided insight into the mechanism of covalent flavinylation and the rationale for this atypical protein modification.
Identification and characterization of eukaryotic initiation factor 5A‐2
Authors: Clement Paul M. J. , Henderson C. Allen , Jenkins Zandra A. , Smit‐McBride Zeljka , Wolff Edith C , Hershey John W. B. , Park Myung Hee , Johansson Hans E
Published: October 2003

The phylogenetically conserved eukaryotic translation initiation factor 5A (eIF5A) is the only known cellular protein to contain the post‐translationally derived amino acid hypusine [Nε‐(4‐amino‐2‐hydroxybutyl)lysine]. Both eIF5A and its hypusine modification are essential for sustained cell proliferation. Normally only one eIF5A protein is expressed in human cells. Recently, we identified a second human EIF5A gene that would encode an isoform (eIF5A‐2) of 84% sequence identity. Overexpression of eIF5A‐2 mRNA in certain human cancer cells, in contrast to weak normal expression limited to human testis and brain, suggests EIF5A2 as a potential oncogene. However, eIF5A‐2 protein has not been described in human or mammalian cells heretofore. Here, we describe the identification of eIF5A‐2 protein in human colorectal and ovarian cancer lines, SW‐480 and UACC‐1598, that overexpress eIF5A‐2 mRNAs. Functional characterization of the human isoforms revealed that either human EIF5A gene can complement growth of a yeast strain in which the yeast EIF5A genes were disrupted. This indicates functional similarity of the human isoforms in yeast and suggests that eIF5A‐2 has an important role in eukaryotic cell survival similar to that of the ubiquitous eIF5A‐1. Detectable structural differences were also noted, including lack of immunological cross‐reactivity, formation of different complexes with deoxyhypusine synthase, and Km values (1.5 ± 0.2 vs. 8.3 ± 1.4 µm for eIF5A‐1 and ‐2, respectively) as substrates for deoxyhypusine synthase in vitro. These physical characteristics and distinct amino acid sequences in the C‐terminal domain together with differences in gene expression patterns imply differentiated, tissue‐specific functions of the eIF5A‐2 isoform in the mammalian organism and in cancer.
The sex‐peptide DUP99B is expressed in the male ejaculatory duct and in the cardia of both sexes
Authors: Rexhepaj Albana , Liu Huanfa , Peng Jing , Choffat Yves , Kubli Eric
Published: October 2003

Mating elicits two postmating responses in many insect females: the egg laying rate increases and sexual receptivity is reduced. In Drosophila melanogaster, two peptides of the male genital tract, sex‐peptide and DUP99B, elicit these postmating responses when injected into virgin females. Here we show that the gene encoding DUP99B is expressed in the male ejaculatory duct and in the cardia of both sexes. The DUP99B that is synthesized in the ejaculatory duct is transferred, during mating, into the female genital tract. Expression of the gene is first seen in a late pupal stage. Males containing an intact ejaculatory duct, but lacking accessory glands, initiate the two postmating responses in their female partners [Xue, L. & Noll, M. (2000) Proc. Natl Acad. Sci. USA97, 3272–3275]. Although such males synthesize DUP99B in wild‐type quantities, they elicit only weak postmating responses in their mating partners. Males lacking the Dup99B gene elicit the two postmating responses to the same extent as wild‐type males. These results suggest that both sex‐peptide and DUP99B can elicit both responses in vivo. However, sex‐peptide seems to play the major role in eliciting the postmating responses, while DUP99B may have specialized for other, as yet unknown, functions.
Kinetics and thermodynamics of nick sealing by T4 DNA ligase
Authors: Cherepanov Alexey V. , de Vries Simon
Published: October 2003

T4 DNA ligase is an Mg2+‐dependent and ATP‐dependent enzyme that seals DNA nicks in three steps: it covalently binds AMP, transadenylates the nick phosphate, and catalyses formation of the phosphodiester bond releasing AMP. In this kinetic study, we further detail the reaction mechanism, showing that the overall ligation reaction is a superimposition of two parallel processes: a ‘processive’ ligation, in which the enzyme transadenylates and seals the nick without dissociating from dsDNA, and a ‘nonprocessive’ ligation, in which the enzyme takes part in the abortive adenylation cycle (covalent binding of AMP, transadenylation of the nick, and dissociation). At low concentrations of ATP (< 10 µm) and when the DNA nick is sealed with mismatching base pairs (e.g. five adjacent), this superimposition resolves into two kinetic phases, a burst ligation (≈ 0.2 min−1) and a subsequent slow ligation (≈ 2 × 10−3 min−1). The relative rate and extent of each phase depend on the concentrations of ATP and Mg2+. The activation energies of self‐adenylation (16.2 kcal·mol−1), transadenylation of the nick (0.9 kcal·mol−1), and nick‐sealing (16.3–18.8 kcal·mol−1) were determined for several DNA substrates. The low activation energy of transadenylation implies that the transfer of AMP to the terminal DNA phosphate is a spontaneous reaction, and that the T4 DNA ligase–AMP complex is a high‐energy intermediate. To summarize current findings in the DNA ligation field, we delineate a kinetic mechanism of T4 DNA ligase catalysis.
A selenium‐containing single‐chain abzyme with potent antioxidant activity
Authors: You Delin , Ren Xiaojun , Xue Yan , Luo Guimin , Yang Tongshu , Shen Jiacong
Published: October 2003

Reactive oxygen species (ROS) are products of normal metabolic activities and are thought to be the cause of many diseases. A selenium‐containing single‐chain abzyme 2F3 (Se‐2F3‐scFv) that imitates glutathione peroxidase has been produced which has the capacity to remove ROS. To evaluate the antioxidant ability of Se‐2F3‐scFv, we constructed a ferrous sulfate/ascorbate (Vc/Fe2+)‐induced mitochondrial damage model system and investigated the capacity of Se‐2F3‐scFv to protect mitochondria from oxidative damage. Se‐2F3‐scFv markedly decreased mitochondrial swelling, inhibited lipid peroxidation, and maintained the activity of cytochrome c oxidase, in comparison with Ebselen, a well‐studied glutathione peroxidase mimic, indicating that Se‐2F3‐scFv has potential for treating diseases mediated by ROS.
Fluorescence analysis of the Hansenula polymorpha peroxisomal targeting signal‐1 receptor, Pex5p
Authors: Boteva Raina , Koek Anne , Visser Nina V. , Visser Antonie J.W.G. , Krieger Elmar , Zlateva Theodora , Veenhuis Marten , van der Klei Ida
Published: October 2003

Correct sorting of newly synthesized peroxisomal matrix proteins is dependent on a peroxisomal targeting signal (PTS). So far two PTSs are known. PTS1 consists of a tripeptide that is located at the extreme C terminus of matrix proteins and is specifically recognized by the PTS1‐receptor Pex5p. We studied Hansenula polymorpha Pex5p (HpPex5p) using fluorescence spectroscopy. The intensity of Trp fluorescence of purified HpPex5p increased by 25% upon shifting the pH from pH 6.0 to pH 7.2. Together with the results of fluorescence quenching by acrylamide, these data suggest that the conformation of HpPex5p differs at these two pH values. Fluorescence anisotropy decay measurements revealed that the pH affected the oligomeric state of HpPex5p, possibly from monomers/dimers at pH 6.0 to larger oligomeric forms at pH 7.2. Addition of dansylated peptides containing a PTS1, caused some shortening of the average fluorescence lifetime of the Trp residues, which was most pronounced at pH 7.2. Our data are discussed in relation to a molecular model of HpPex5p based on the three‐dimensional structure of human Pex5p.
Purification and characterization of a sialic acid specific lectin from the hemolymph of the freshwater crab Paratelphusa jacquemontii
Authors: Denis Maghil , Palatty P. D. Mercy , Bai N. Renuka , Suriya S. Jeya
Published: October 2003

A naturally occurring hemagglutinin was detected in the serum of the freshwater crab, Paratelphusa jacquemontii (Rathbun). Hemagglutination activity with different mammalian erythrocytes suggested a strong affinity of the serum agglutinin for horse and rabbit erythrocytes. The most potent inhibitor of hemagglutination proved to be bovine submaxillary mucin. The lectin was purified by affinity chromatography using bovine submaxillary mucin‐coupled agarose. The molecular mass of the purified lectin was 34 kDa as determined by SDS/PAGE. The hemagglutination of purified lectin was inhibited by N‐acetylneuraminic acid but not by N‐glycolylneuraminic acid, even at a concentration of 100 mm. Bovine submaxillary mucin, which contains mainly 9‐O‐acetyl‐ and 8,9 di‐O‐acety‐N‐acetyl neuraminic acid was the most potent inhibitor of the lectin. Sialidase treatment and de‐O‐acetylation of bovine submaxillary mucin abolished its inhibitory capacity completely. Also, asialo‐rabbit erythrocytes lost there binding specificity towards the lectin. The findings indicated an O‐acetyl neuraminic acid specificity of the lectin.
Biochemical and molecular characterization of hazelnut (Corylus avellana) seed lipoxygenases
Authors: Santino Angelo , De Paolis Angelo , Gallo Antonia , Quarta Angela , Casey Rod , Mita Giovanni
Published: October 2003

Plant lipoxygenases (LOXs) are a class of dioxygenases which display diverse functions in several physiological processes such as growth, development and response to biotic and abiotic stresses. Even though LOXs have been characterized from several plant species, the physiological role of seed LOXs is still unclear. With the aim to better clarify the occurrence of LOXs and their influence on hazelnut seed quality, we carried out the biochemical and molecular characterization of the main LOX isoforms expressed during seed development. A genomic clone containing a complete LOX gene was isolated and fully characterized. The 9887 bp sequence reported contains an open reading frame of 5334 bp encoding a putative polypeptide of 99 kDa. Semiquantitative RT‐PCR carried out from RNAs extracted from seeds at different maturation stages showed that LOXs are mainly expressed at early developmental stages. These results were confirmed by LOX activity assays. Biochemical characterization of the reaction products of the hazelnut LOX indicated that it is a 9‐LOX. Two cDNAs were isolated by RT‐PCR carried out on total RNA from immature hazelnut seeds. Sequence analysis indicated that the two cDNAs are highly homologous (91.9% degree of identity) and one of these corresponded exactly to the genomic clone. The deduced amino acid sequences of the hazelnut LOXs showed that they are closely related to a previously reported almond LOX (79.5% identity) and, to a lesser extent, to some LOXs involved in plant responses to pathogens (cotton and tobacco LOXs, 75.5 and 74.6% identity, respectively). The physiological role of hazelnut LOXs and their role in influencing seed quality are also discussed.
Local changes in the catalytic site of mammalian histidine decarboxylase can affect its global conformation and stability
Authors: Rodríguez‐Caso Carlos , Rodríguez‐Agudo Daniel , Moya‐García Aurelio A. , Fajardo Ignacio , Medina Miguel Ángel , Subramaniam Vinod , Sánchez‐Jiménez Francisca
Published: October 2003

Mature, active mammalian histidine decarboxylase is a dimeric enzyme of carboxy‐truncated monomers (≈ 53 kDa). By using a biocomputational approach, we have generated a three‐dimensional model of a recombinant 1/512 fragment of the rat enzyme, which shows kinetic constants similar to those of the mature enzyme purified from rodent tissues. This model, together with previous spectroscopic data, allowed us to postulate that the occupation of the catalytic center by the natural substrate, or by substrate‐analogs, would induce remarkable changes in the conformation of the intact holoenzyme. To investigate the proposed conformational changes during catalysis, we have carried out electrophoretic, chromatographic and spectroscopic analyses of purified recombinant rat 1/512 histidine decarboxylase in the presence of the natural substrate or substrate‐analogs. Our results suggest that local changes in the catalytic site indeed affect the global conformation and stability of the dimeric protein. These results provide insights for new alternatives to inhibit histamine production efficiently in vivo.
α‐Fetoprotein positively regulates cytochrome c‐mediated caspase activation and apoptosome complex formation
Authors: Semenkova Lidia , Dudich Elena , Dudich Igor , Tokhtamisheva Natalie , Tatulov Edward , Okruzhnov Yury , Garcia‐Foncillas Jesus , Palop‐Cubillo Juan‐Antonio , Korpela Timo
Published: October 2003

Previous results have shown that the oncoembryonic marker α‐fetoprotein (AFP) is able to induce apoptosis in tumor cells through activation of caspase 3, bypassing Fas‐dependent and tumor necrosis factor receptor‐dependent signaling. In this study we further investigate the molecular interactions involved in the AFP‐mediated signaling of apoptosis. We show that AFP treatment of tumor cells is accompanied by cytosolic translocation of mitochondrial cytochrome c. In a cell‐free system, AFP mediates processing and activation of caspases 3 and 9 by synergistic enhancement of the low‐dose cytochrome c‐mediated signals. AFP was unable to regulate activity of caspase 3 in cell extracts depleted of cytochrome c or caspase 9. Using high‐resolution chromatography, we show that AFP positively regulates cytochrome c/dATP‐mediated apoptosome complex formation, enhances recruitment of caspases and Apaf‐1 into the complex, and stimulates release of the active caspases 3 and 9 from the apoptosome. By using a direct protein–protein interaction assay, we show that pure human AFP almost completely disrupts the association between processed caspases 3 and 9 and the cellular inhibitor of apoptosis protein (cIAP‐2), demonstrating its release from the complex. Our data suggest that AFP may regulate cell death by displacing cIAP‐2 from the apoptosome, resulting in promotion of caspase 3 activation and its release from the complex.
Molecular Characterization of a Strain ofSquash Leaf Curl China Virusfrom the Philippines
Authors: Kon T. , Dolores L. M. , Bajet N. B. , Hase S. , Takahashi H. , Ikegami M.
Published: October 2003

The complete nucleotide sequence of infectious cloned DNA components (A and B) of the causal agent of squash leaf curl disease in the Philippines was determined. DNA‐A and DNA‐B comprise 2739 and 2705 nucleotides, respectively; the common region is 174 bases in length. Five ORFs were found in DNA‐A and two in DNA‐B. Partial dimeric clones containing DNA‐A and DNA‐B, constructed in a binary vector and transformed intoAgrobacterium tumefaciens, induced systemic infection in agro‐inoculated pumpkin plants (Cucurbita moschata). The total DNA‐A sequence was most closely related to that ofSquash leaf curl China virus(SLCCNV) (88% identity), although the existence of B component of SLCCNV has not been reported. The deduced coat protein was like that of SLCCNV (98% amino acid sequence identity) and the Philippines virus has low sequence identity toSquash leaf curl virus(SLCV) andSquash mild leaf curl virus(SMLCV) (63 and 64% total nucleotide sequence identities, respectively). From these results, we propose that the Philippines virus be designatedSquash leaf curl China virus‐[Philippines] (SLCCNV‐[PH]).
Survival and Responses of Botrytis cinerea after Exposure to Ethanol and Heat
Authors: Lichter A. , Zhou H.‐W. , Vaknin M. , Dvir O. , Zutchi Y. , Kaplunov T. , Lurie S.
Published: October 2003

Dipping of table grapes in ethanol was previously shown to control bunch decay caused by Botrytis cinerea and further studies were conducted in order to quantify the effect of ethanol on the fungus. Different methods were used to determine that 30% ethanol prevented spore germination. Spores exposed to a range of temperatures, up to 49.5°C for 30 s, survived but were killed at the lethal heat dose of 46.3°C for 2 min. The combined application of ethanol and heat to ungerminated spores caused retardation of subsequent mycelial development. Ethanol or heat treatment of mature berries did not alter the expression of the gene that encodes the heat‐shock protein, HSP101 or grape berry alcohol dehydrogenase. Transcription of the heat‐shock gene, HSP104 of B. cinerea in mycelia was elevated 4 h after exposure to ethanol or heat, but not 18 h after such exposure. It is suggested that tolerance of spores to ethanol or heat may be correlated with their ability to repair damage through the action of chaperones such as HSP104.
The importance of social and cultural factors to nursing status
Authors: Bojtor Anna
Published: October 2003

The objective of the authors was to examine nursing staff workload. The examination is based on the workload model of Robert Karasek and on the status integration theory of P. Gibbs and Walter T. Martin. As a means of examination, an open questionnaire consisting of 55 items and 76 variables was given to nurses working in different fields of health care, studying in higher education instead, or to persons with a degree in nursing (n = 465). The main concern of the authors was to attempt to identify the inconsistencies in nursing status which might come from limited decision‐making freedom and high workplace requirements. In the process of statistical analysis of the data, considerable deviation was observed regarding some scales of the questionnaire. The authors emphasize that the reasons for the inconsistencies in nursing status might originate either from traditional cultural elements or from increased workload based on unclear competencies which are forced on the nurses.
Imbedding quality improvement into all aspects of nursing practice
Authors: Long Lesley E
Published: October 2003

Influencing nurses to accept and implement best practice and continuous quality improvement into everyday nursing activities and to change practice is a significant challenge for any nursing service. Quality improvement is often considered by clinicians to be yet another task to be added to their already busy workloads. This paper describes the attempts of one large metropolitan tertiary teaching hospital to imbed quality improvement and best practice across the spectrum of its nursing service and to foster a culture of quality within clinical practice and documentation. It outlines the methods used by one hospital to enshrine best practice into all nursing activities and to minimize variations in practice throughout the hospital through the use of its nursing clinical information system.
Quality improvement measures based on patient data: Some psychometric issues
Authors: Larsson Gerry , Larsson Bodil Wilde
Published: October 2003

Existing methods for handling patients’ opinions as a basis for quality improvement measures tend to be too sophisticated for practical nursing and/or weak psychometrically. An Index of Measures was developed by combining patients’ perceptions of actual care conditions with the subjective importance they ascribed to these conditions. Data from Swedish somatic inpatients (n = 4002) indicated that this index gave added value compared to patients’ ratings of actual care conditions only. Interpretation of the Index of Measures focused on the proportion of patients who receive the value ‘deficiency.’ This value is obtained if a given patient has a subjective importance rating on a given item which is higher than the mean subjective importance score for the whole patient group on that item, as well as a perceived reality rating on the item in question which is lower than the mean perceived reality rating for the whole group on that item. Guidelines are suggested on when improvement actions are necessary and when they are not necessary.
Evaluating the impact of a new high dependency unit
Authors: Armstrong Kiele , Young Jeanne , Hayburn Alison , Irish Benjamin , Nikoletti Sue
Published: October 2003

The impact of a new high dependency unit (HDU) that was developed from an adjoining ward was evaluated from both an organizational and consumer perspective. Satisfaction of patients and relatives with nursing care on the ward of origin was measured before and after the development of the HDU, and nurses were surveyed regarding the impact of the change on nursing practices. Patients’ perceptions of pain and pain management was compared between patients admitted to the ward prior to the development of the HDU, and a later group admitted to the HDU. Results showed that despite high satisfaction prior to the development of the HDU, satisfaction of patients and relatives improved on the ward after the development of the HDU. Nurses in both areas also described higher satisfaction levels after the change. Patients admitted to the HDU expressed higher levels of satisfaction with nurses’ response to analgesia requests and clinical data showed a trend toward lower pain scores in this group compared with pre‐HDU patients. The HDU was found to be effective in providing high level care and monitoring for a group of patients at risk of postoperative complications. It was also associated with additional benefits in the running of the adjacent ward.
Evidence‐based practice: To be or not to be, this is the question!
Authors: Zeitz Kathryn , McCutcheon Helen
Published: October 2003

Evidence‐based nursing is the current fashion. It is being touted as the mechanism to achieve best practice in the clinical setting. But while evidence‐based practice (EBP) is being presented in the literature, discussed at nursing practice forums, and evidence‐based centres of excellence have developed, there seems to be very little impact in the practice that nurses deliver on a daily basis. The case in point is the collection of vital signs. While not historically a nursing skill, over the last 60 years it has become an integral component of practice in the postoperative general surgical setting. The evidence to support these practices is scant. Policies and text purport traditional routine‐regulated practice without substantive evidence to support their claims. These polices are being used to control rather than support EBP. In conjunction with the traditional practice of vital sign collection and the culture of the clinical settings, the policies are limiting opportunities for clinicians to make individual decisions about care delivery based on the unique needs of each patient. Rather than focusing on EBP as the solution to the development of best practice, is it not time to change the focus to real strategies that will assist in achieving best practice? These include the creation of rigorous relevant evidence, the valuing of clinical expertise and the changing of the cultures in which nurses develop and practice.
Simplified 13C‐urea breath test with a new infrared spectrometer for diagnosis of Helicobacter pylori infection
Authors: CHEN TSENG‐SHING , CHANG FULL‐YOUNG , CHEN PANG‐CHI , HUANG THOMAS W , OU JONATHAN T , TSAI MING‐HUNG , WU MING‐SHIANG , LIN JAW‐TOWN
Published: October 2003

Background and Aim:  Infrared spectrometry has correlated excellently with mass spectrometry in detecting the ratio of 13CO2 to 12CO2 in breath samples. The present study aimed to evaluate the accuracy of the 13C‐urea breath test (13C‐UBT) using a new model of infrared analyzer.

Methods:  A total of 600 patients who were undergoing upper endoscopy without receiving eradication therapy were entered into the study. Culture, histology, and rapid urease test on biopsies from the antrum and corpus of the stomach were used for the determination of Helicobacter pylori infection. Breath samples were collected before and 20 min after drinking 100 mg 13C‐urea in 100 mL water. The optimal cutoff value was determined by the receiver operating characteristic curve.

Results:  Of the 586 patients who were eligible for analysis, 369 were positive for H. pylori infection, 185 were negative for H. pylori infection, and 32 were indeterminate. When the appropriate cutoff value was set at 3.5‰, a sensitivity of 97.8%, a specificity of 96.8% and an accuracy of 97.5% were obtained using the 13C‐UBT. The accuracy of the 13C‐UBT decreased when CO2 concentration in the breath sample was <2%, as compared with ≥2% (93.6%vs 97.7%), mainly because of a decrease in specificity (81.8%vs 97.7%). There were 2.7% of patients with Δ13CO2 values that fell between 3.0–4.5‰, in whom the risk of error was 47%.

Conclusions:  The 13C‐UBT performed with infrared spectrometry is a highly sensitive, specific, and non‐invasive method for the detection of H. pylori infection. The immediate availability of the test result and technical simplicity make it particularly effective in routine clinical practice.
Effect of Helicobacter pylori eradicated therapy on water gastric emptying in patients with active duodenal ulcer
Authors: CHANG FULL‐YOUNG , LU CHING‐LIANG , CHEN CHIH‐YEN , LUO JIING‐CHYUAN , JIUM KANG LIH , LEE SHOU‐DONG
Published: October 2003

Background and Aims:  It remains debatable if duodenal ulcer (DU) or Helicobacter pylori infection has a definite impact on human gastric emptying (GE). We explored the nature of water GE in active DU patients before and after ulcer healing and the influence of H. pylori eradication on GE.

Methods:  A home made applied potential tomography (APT) was used to measure liquid GE. Twelve electrodes were placed in a circular array around the upper abdomen of studied subjects. After drinking 500 mL of ion‐free water, paired electrodes injected electrical current and the remaining 10 electrodes recorded signals, one‐by‐one in a rotating order. Based on tomographical calculation, the serial changes of averaged signals from altered resistivities were constructed to display GE. Initially, 64 H. pylori infected active DU patients were enrolled. After APT measurement, one‐week triple therapy (omeprazole, amoxicillin and clarithromycin) was dispensed. Patients were asked back to determine ulcer/H. pylori status and GE on a scheduled date 3 months later. Finally, 58 patients finished the trial with valid and readable GE data obtained.

Results:  The ulcer healing and H. pylori eradicated rates were 91.4% and 82.8%, respectively. In general, liquid GE was prolonged in all DU patients at follow up. Of 48 eradicated patients, 35.4% manifested either enhanced or delayed GE before treatment, whereas only five (10.4%) had abnormal GE after treatment (P < 0.0001). In contrast, this characteristically normalized GE was not found in non‐eradicated patients.

Conclusions:  Water GE of active DU patients ranges from enhanced to delayed, while an effective H. pylori triple therapy is useful not only for healing ulcers, but also for restoring abnormal GE.
Differential diagnosis of intrahepatic bile duct dilatation without demonstrable mass on ultrasonography or CT: Benign versus malignancy
Authors: KIM HONG JOO , LEE KYU TAEK , KIM SEUNG HOON , LEE JONG KYUN , LIM JAE HOON , PAIK SEUNG WOON , RHEE JONG CHUL
Published: October 2003

Background:  The purpose of the present study was to define the differential diagnostic markers of benign and malignant bile duct strictures without demonstrable mass on ultrasonography (US) or computed tomography (CT) using aspects of clinical, laboratory and imaging findings.

Methods:  Between February 1995 and February 2001, 24 patients who underwent surgical resection for dilations of the bile duct (peripheral and hilar) without visible mass lesion on US or CT were included in our study. Hospital records, laboratory results, findings of imaging studies and pathological findings were reviewed retrospectively.

Results:  For laboratory results, levels of alkaline phosphatase (benign 163.9 ± 145.1 vs malignant 407.25 ± 481.7; p < 0.05) and CA 19–9 (benign 25.0 ± 41.1 vs malignant 614.6 ± 818.5; p < 0.05) levels were significantly elevated in the malignant group compared with that of the benign group. Findings such as thickening of the bile duct wall ≥ 5 mm (P < 0.05) on radiological examination, significant regional lymph node enlargement (> 1 cm, p = 0.01) on CT scan and abrupt cut‐off (P < 0.01), and separation of bile ducts (P < 0.05) on cholangiogram were differential diagnostic markers between the two groups.

Conclusions:  Preoperative laboratory data such as alkaline phosphatase and CA 19–9, and imaging findings such as significant wall thickening and regional lymph node enlargement on CT, abrupt cut‐off and separation of bile duct on cholangiogram are useful differential diagnostic markers for benign and malignant bile duct strictures without demonstrable mass on US or CT.
Elevation of serum and ascites cancer antigen 125 levels in patients with liver cirrhosis
Authors: XIAO WEN‐BIN , LIU YU‐LAN
Published: October 2003

Background and Aim:  Serum cancer antigen (CA) 125 elevation has been reported in patients with liver disease, but it is poorly characterized. The present study aimed to evaluate the range of serum and ascitic CA 125 levels in patients with liver cirrhosis and to explore possible factors associated with CA 125 elevation.

Methods:  A total of 70 patients were studied. Group I consisted of 30 patients with liver cirrhosis with or without ascites. Group II consisted of 30 patients with digestive malignant tumors with or without ascites. Group III consisted of 10 patients with benign ascites. The CA 125 levels were measured in the serum of all patients and also simultaneously in the ascitic fluid of 15 patients.

Results:  Serum CA125 levels in 80% of (24/30) patients from group I were elevated, particularly in those with ascites, irrespective of the etiology of cirrhosis. Serum CA 125 levels were correlated with Child–Pugh scores (r = 0.38), but not significantly (P = 0.06). All patients from group II with ascites and from group III had elevated serum CA 125 levels, but there was no difference in the serum CA 125 levels between patients with ascites from group I (275 ± 175 U/mL), group II (368 ± 190 U/mL) or group III (396 ± 287 U/mL), nor was there a significant difference in ascitic CA 125 levels (P > 0.05). The levels of serum CA 125 (198 ± 108 U/mL) were lower than, but correlated with that of ascites (460 ± 234 U/mL, r = 0.58, P = 0.026). The elevation of serum CA 125 accompanied by abnormalities of other tumor markers was more common in malignant ascites than in benign ascites (90% compared with 6%, P < 0.05).

Conclusion:  The elevation of serum CA 125 is common in patients with liver cirrhosis. It is related to the presence of ascites, and possibly to the insufficiency of liver function, but not the etiology of cirrhosis and ascites. Serum CA 125 probably comes from ascites. It usually predicts benign disease if the elevation of serum or ascites CA 125 is not accompanied by the abnormalities of other tumor markers.
Possible involvement of tetraspanin CO‐029 in hematogenous intrahepatic metastasis of liver cancer cells
Authors: KANETAKA KENGO , SAKAMOTO MICHIIE , YAMAMOTO YOSHIYA , TAKAMURA MASAAKI , KANEMATSU TAKASHI , HIROHASHI SETSUO
Published: October 2003

Background and Aim:  A correlation between overexpression of tetraspanin CO‐029 and the intrahepatic spread of hepatocellular carcinoma (HCC) has been observed in surgically resected specimens from humans. However, the cellular mechanisms involved in CO‐029 protein modulation of the metastatic phenotype are unknown. In the present study, CO‐029 cDNA was stably transfected into a non‐metastatic human HCC cell line to investigate whether it could directly promote metastasis.

Methods:  We constructed a human HCC cell line that stably overexpressed CO‐029 and mock transfectants. Using these transfectants, we examined cell proliferation characteristics in monolayer culture and the ability to adhere to culture plates coated with laminin, fibronectin, vitronectin and collagen type‐I or type‐IV. Orthotopic implantation of these transfectants to SCID mice was also performed.

Results:  Several clones of CO‐029 transfectants and mock transfectants were established. The growth rates and adhesive properties to the extracellular matrix did not differ between CO‐029 and mock transfectants. When orthotopically implanted, the size of the primary tumor in the liver did not differ between CO‐029 and the mock transfectants. However, only CO‐029 positive clones developed intrahepatic metastatic lesions.

Conclusion:  These results suggest that CO‐029 might be involved in hematogenous intrahepatic metastasis, although the precise cellular mechanisms involved remain unknown.
Rodent nutritional model of non‐alcoholic steatohepatitis: Species, strain and sex difference studies
Authors: KIRSCH RICHARD , CLARKSON VIVIAN , SHEPHARD ENID G , MARAIS DAVID A , JAFFER MOHAMED A , WOODBURNE VIVIENNE E , KIRSCH RALPH E , HALL PAULINE DE LA M
Published: October 2003

Background and Aim:  The methionine choline‐deficient (MCD) diet leads to steatohepatitis in rodents. The aim of the present study was to investigate species, strain and sex differences in this nutritional model of non‐alcoholic steatohepatitis (NASH).

Methods:  Male and female Wistar, Long–Evans and Sprague–Dawley rats, and C57/BL6 mice (n = 6 per group) were fed a MCD diet for 4 weeks. Control groups received an identical diet supplemented with choline bitartrate (0.2% w/w) and methionine (0.3% w/w). Liver pathology (steatosis and inflammation) and ultrastructure, liver lipid profile (total lipids, triglycerides, lipid peroxidation products), liver : body mass ratios and serum alanine aminotransferase (ALT) levels were compared between these groups.

Results:  The MCD diet‐fed male rats developed greater steatosis (P < 0.001), had higher liver lipid content (P < 0.05) and had higher serum ALT levels (P < 0.005) than did female rats. Wistar rats (both sexes) had higher liver lipid levels (P < 0.05), serum ALT levels (P < 0.05), and liver mass : body mass ratios (P < 0.025) than did Long–Evans and Sprague–Dawley rats. In female groups, Wistar rats showed greater fatty change than did the other two strains (P < 0.05). All rats fed the MCD diet developed hepatic steatosis, but necrosis and inflammation were minor features and fibrosis was absent. Compared with Wistar rats, male C57/BL6 mice showed a marked increase in inflammatory foci (P < 0.001), end products of lipid peroxidation (free thiobarbituric acid reactive substances) (P < 0.005), and mitochondrial injury, while showing less steatosis (P < 0.005), lower hepatic triglyceride levels, (P < 0.005) and lower early lipid peroxidation products (conjugated dienes and lipid hydroperoxides; P < 0.005 and P < 0.01, respectively).

Conclusions:  The Wistar strain and the male sex are associated with the greatest degree of steatosis in rats subjected to the MCD diet. Of the groups studied, male C57/BL6 mice develop the most inflammation and necrosis, lipid peroxidation, and ultrastructural injury, and best approximate the histological features of NASH.
Development of real‐time detection direct test for hepatitis B virus and comparison with two commercial tests using the WHO international standard
Authors: MUKAIDE MOTOKAZU , TANAKA YASUHITO , KATAYOSE SATOSHI , TANO HIROYUKI , MURATA MITSUHIRO , HIKATA MIKIO , FUJISE KIYOTAKA , SAKUGAWA HIROSHI , SUZUKI KAZUO , ZAUNDERS JOHN , NAGASAWA YOKO , TODA GOTARO , MIZOKAMI MASASHI
Published: October 2003

Aims:  A highly reproducible and sensitive hepatitis B virus real‐time detection direct (HBV RTD‐direct) test using DNA extraction by magnetic beads coated with polyclonal anti‐HBsAg, followed by the real‐time detection polymerase chain reaction (PCR) method, was developed for the detection of HBV DNA.

Methods:  The HBV DNA could be extracted from the HBsAg positive viral particles without resulting in viral DNA fragmentation. The HBV RTD‐direct test was validated using a serial dilution panel of the WHO standard HBV DNA 97/746 I.

Results:  The test had a dynamic range of 0.7–8.0 log10 international units (IU) per mL and the results were shown to be comparable to those obtained with two commercially available tests: the HBV DNA transcription‐mediated amplification‐hybridization protection assay and the Amplicor HBV Monitor test. In addition, the HBV RTD‐direct test, based on magnetic extraction, successfully eliminated PCR inhibitors in clinical specimens.

Conclusion:  We conclude that the HBV RTD‐direct test is an excellent alternative for monitoring patients undergoing antiviral treatment or for screening various clinical specimens.