bluuebunny/crossref_metadata_2025 huggingface dataset:
Leptin levels in gingival crevicular fluid in periodontal health and disease
Authors: Karthikeyan B. V. , Pradeep A. R.
Published: December 2006

Background and Objective:  A high concentration of leptin is associated with healthy gingival tissue, and the concentration of leptin decreases as periodontal disease progresses. However, to date, the leptin concentration in gingival crevicular fluid has not been documented. Hence, the present study was carried out to explore the presence of leptin in gingival crevicular fluid in periodontal health and disease, and to probe further into its possible role in periodontal disease progression.

Material and Methods:  A total of 45 adult patients were selected, based on their body mass index, for the study. They were categorized into three groups of 15 patients each, based on their periodontal tissue status, as follows: group I (clinically healthy gingiva with no loss of attachment); group II (chronic gingivitis with no loss of attachment); and group III (chronic periodontitis). Gingival crevicular fluid samples of 1 µL were collected extracrevicularly using white color‐coded 1–5 µL calibrated volumetric microcapillary pipettes from one site in each person, and samples were analyzed for leptin using a commercially available enzyme‐linked immunosorbent assay kit.

Results:  The concentration of leptin in gingival crevicular fluid of patients in group I (2292.69 pg/mL) was statistically higher (p < 0.05) than in those of groups II (1409.95 pg/mL) and III (1071.89 pg/mL). This suggests a negative correlation of gingival crevicular fluid leptin concentration with clinical attachment loss (p < 0.05).

Conclusion:  As periodontal tissue destruction increased, there was a substantial decrease in gingival crevicular fluid leptin concentration. This observation extends our knowledge of the protective role of leptin in periodontal health.
Comparison of experimental gingivitis with persistent gingivitis: differences in clinical parameters and cytokine concentrations
Authors: Deinzer R. , Weik U. , Kolb‐Bachofen V. , Herforth A.
Published: December 2006

Background and Objective:  Experimental gingivitis has been studied extensively as a well‐controlled laboratory model of gingivitis. It is unclear, however, how experimental gingivitis compares with persistent plaque and gingivitis in more naturalistic settings. The present study compares both conditions in a randomized controlled design.

Material and Methods:  Twenty‐six students suffering from plaque and gingivitis were randomly assigned to either a persistent gingivitis or an experimental gingivitis condition. Subjects with persistent gingivitis continued their habitual (i.e. insufficient) oral hygiene behaviour, resulting in persistence of plaque and gingivitis. Experimental gingivitis consisted of initial prophylaxis and subsequent total neglect of oral hygiene. Crevicular interleukin‐1β and interleukin‐8 and clinical data were assessed weekly.

Results:  After 4 wk, subjects with experimental gingivitis showed significantly more plaque accumulation (p = 0.005), higher interleukin‐1β (p = 0.037), and lower interleukin‐8 (p = 0.043) concentrations than subjects with persistent gingivitis. Whereas in experimental gingivitis we observed considerable fluctuations in clinical and immunological parameters over the 4‐wk period, persistent gingivitis was characterized by little fluctuation, indicating that we were monitoring an inflammatory steady state.

Conclusion:  The data indicate that conditions observed after 4 wk of experimental gingivitis are not comparable with persistent gingival inflammation in a naturalistic setting. Results are discussed with respect to current studies, indicating that chronic inflammation may reflect a stage of down‐regulated pro‐inflammatory response.
Reduced brain‐derived neurotrophic factor is associated with a loss of serotonergic innervation in the hippocampus of aging mice
Authors: Luellen B. A. , Bianco L. E. , Schneider L. M. , Andrews A. M.
Published: December 2006

Brain‐derived neurotrophic factor (BDNF) regulates monoamine neuronal growth, survival and function in development and throughout adulthood. At 18 months of age, mice with constitutive reductions in BDNF expression show decreased serotonin innervation in the hippocampus compared with age‐matched wildtype mice. It is not known, however, whether age‐accelerated loss of serotonergic innervation in BDNF+/− mice occurs in other brain regions, advances beyond 18 months or is associated with alterations in other neurotransmitter systems. In this study, immunocytochemistry was used to assess serotonergic and catecholaminergic innervation in 26‐month‐old BDNF+/− mice. Age‐related loss of serotonin axons in the hippocampus was potentiated in BDNF+/− mice compared with wildtype mice at this late age, particularly in the CA1 subregion. By contrast, aging BDNF+/− mice showed increased serotonin innervation of the basomedial nucleus of the amygdala. In the noradrenergic system, BDNF+/− mice showed reduced numbers of cell bodies and fibers in the locus coeruleus compared with age‐matched wildtype mice, whereas no changes were observed in dopaminergic innervation with respect to genotype. In vivo zero net flux microdialysis in awake mice showed a significant decrease in extracellular serotonin levels in the hippocampus in BDNF+/− mice at 20 months of age. Thus, reduced BDNF is associated with altered serotonergic and noradrenergic innervation in aging mice and, in particular, with accelerated loss of serotonergic innervation to the hippocampus that is manifest as a decrease in basal neurotransmission.
Adoption of a Ventilator‐Associated Pneumonia Clinical Practice Guideline
Authors: Abbott Cynthia A. , Dremsa Theresa , Stewart Della W. , Mark Debra D. , Swift Caren C.
Published: December 2006

Conceptual Framework:The Academic Center for Evidence‐based Practice (ACE) Star Model was used to implement an evidence‐based clinical practice guideline (CPG) in order to decrease ventilator‐associated pneumonia (VAP) incidence rates and ventilator days. The goal was to interrupt person‐to‐person transmission of bacteria and bacterial colonization using low‐cost, evidence‐based strategies to prevent VAP.

Discovery:Two geographically proximate medical centers, inclusive of five intensive care units located in the southwestern region of the United States had significant variations in their VAP rates.

Evidence summary:Using the U.S. Preventive Services Task Force grading criteria, the results of 69 studies were used to establish a clinical practice guideline to prevent ventilator‐associated pneumonia.

Translation:A clinical practice guideline was developed for the prevention of VAP and included five nursing activities: (a) head‐of‐bed elevation; (b) oral care; (c) ventilator tubing condensate removal; (d) hand hygiene; and (e) glove use. The effect of the CPG, inclusive of an educational intervention, was measured using an observational, prospective, quasi‐experimental design.

Integration:A multidisciplinary education team developed a self‐learning packet, educational materials, and storyboards for the staff as dissemination strategies. Strategies also included e‐mail, one‐on‐one teaching with clinicians, and feedback on guideline adoption and VAP rate reports.

Evaluation:Observation data were collected to evaluate adoption of the CPG while caring for 106 ventilated patients. VAP rates changed at both hospitals although the change was not statistically significant. Additionally, the ICU length of stay declined at both facilities, causing cost savings.

Discussion:These results support the idea that adoption of evidence‐based practices contributes to decreased VAP rates. For a successful program, ICU leaders should emphasize strategies that routinize adoption of evidence‐based CPGs.
Understanding Work Contextual Factors: A Short‐Cut to Evidence‐Based Practice?
Authors: Wallin Lars , Ewald Uwe , Wikblad Karin , Scott‐Findlay Shannon , Arnetz Bengt B.
Published: December 2006

Background: It has become increasingly clear that workplace contextual factors make an important contribution to provider and patient outcomes. The potential for health care professionals of using research in practice is also linked to such factors, although the exact factors or mechanisms for enhancing this potential are not understood. From a perspective of implementing evidence‐based nursing practice, the authors of this article report on a study examining contextual factors.

Aim: The objective of this study was to identify predictors of organizational improvement by measuring staff perceptions of work contextual factors.

Method: The Quality Work Competence questionnaire was used in a repeated measurement survey with a 1‐year break between the two periods of data collection. The sample consisted of 134 employees from four neonatal units in Sweden.

Findings: Over the study period significant changes occurred among staff perceptions, both within and between units, on various factors. Changes in staff perceptions on skills development and participatory management were the major predictors of enhanced potential of overall organizational improvement. Perceived improvement in skills development and performance feedback predicted improvement in leadership. Change in commitment was predicted by perceived decreases in work tempo and work‐related exhaustion.

Conclusions and Implications: These findings indicate the potential for organizational improvement by developing a learning and supportive professional environment as well as by involving staff in decision‐making at the unit level. Such initiatives are also likely to be of importance for enhanced use of research in practice and evidence‐based nursing. On the other hand, high levels of work tempo and burnout appear to have negative consequences on staff commitment for improving care and the work environment. A better understanding of workplace contextual factors is necessary for improving the organizational potential of getting research into practice and should be considered in future implementation projects.
Health Promotion in the Practice Setting: Findings from a Review of Clinical Issues
Authors: Whitehead Dean
Published: December 2006

Background:Many agencies over the last few decades have called for the explicit inclusion and application of health promotion in all aspects of nursing practice. This article reviews the body of empirical (research‐based) and theoretical (conceptual) literature addressing health promotion in nursing clinical practice, to uncover its nature, intention, and impact.

Approach:The aim of this review was to identify from the nursing literature what is reported on in relation to health promotion activity, both theoretical and clinical in the practice setting. An extensive review identified that literature pertaining to these issues clustered around the categories of: theorising and conceptualising the meaning and purpose of health promotion in nursing practice, the development and testing of tools and models for practice, and the exploration of attitudes, behaviours, roles and functions of nurses and health professionals in practice.

Findings:Against more recent trends in health promotion, the literature shows that broader and wide‐ranging health promotion reform has yet to have universal impact, whereas conventional health education activities remain the mainstay for the majority of “health promotion” activities in nursing practice. Where nurses purport to be conducting “health promotion” they are often conducting behavioural, lifestyle, and risk‐orientated health education. The findings of this review therefore support the notion that much of nursing‐related health practice is firmly located in reactive ill health and disease risk‐specific preventive locations, instead of incorporating and instilling broader societal, economic, ecological, and political dimensions of health promotion. Recommendations for wider reform, developed from personal opinion and supported by the literature, are presented.

Conclusions:It is argued that nursing bodies, as well as individual nurses themselves, should seek current health promotion locations in nursing practice, and find effective ways to implement health promotion strategies and, subsequently, equip practitioners with the necessary resources and skills to engage with other health promotion agencies. Future research should be aimed at implementing and evaluating broad health promotion processes.
Leishmania donovani bisubunit topoisomerase I gene fusion leads to an active enzyme with conserved type IB enzyme function
Authors: Das Benu B. , Bose Dasgupta Somdeb , Ganguly Agneyo , Mazumder Saumyabrata , Roy Amit , Majumder Hemanta K.
Published: December 2006

All eukaryotic topoisomerase I enzymes are monomeric enzymes, whereas the kinetoplastid family (Trypanosoma and Leishmania) possess an unusual bisubunit topoisomerase I. To determine what happens to the enzyme architecture and catalytic property if the two subunits are fused, and to explore the functional relationship between the two subunits, we describe here in vitro gene fusion of Leishmania bisubunit topoisomerase I into a single ORF encoding a new monomeric topoisomerase I (LdTOPIL‐fus‐S). It was found that LdTOPIL‐fus‐S is active. Gene fusion leads to a significant modulation of in vitro topoisomerase I activity compared to the wild‐type heterodimeric enzyme (LdTOPILS). Interestingly, an N‐terminal truncation mutant (1–210 amino acids) of the small subunit, when fused to the intact large subunit [LdTOPIL‐fus‐Δ(1–210)S], showed reduced topoisomerase I activity and camptothecin sensitivity in comparison to LdTOPIL‐fus‐S. Investigation of the reduction in enzyme activity indicated that the nonconserved 1–210 residues of LdTOPIS probably act as a ‘pseudolinker’ domain between the core and catalytic domain of the fused Leishmania enzyme, whereas mutational analysis of conserved His453 in the core DNA‐binding domain (LdTOPIL) strongly suggested that its role is to stabilize the enzyme–DNA transition state through hydrogen bonding to one of the nonbridging oxygens. Taken together, our findings provide an insight into the details of the unusual structure of bisubunit topoisomerase I of Leishmania donovani.
Comparative studies of endonuclease I from cold‐adapted Vibrio salmonicida and mesophilic Vibrio cholerae
Authors: Altermark Bjørn , Niiranen Laila , Willassen Nils P. , Smalås Arne O. , Moe Elin
Published: December 2006

Endonuclease I is a periplasmic or extracellular enzyme present in many different Proteobacteria. The endA gene encoding endonuclease I from the psychrophilic and mildly halophilic bacterium Vibrio salmonicida and from the mesophilic brackish water bacterium Vibrio cholerae have been cloned, over‐expressed in Escherichia coli, and purified. A comparison of the enzymatic properties shows large differences in NaCl requirements, optimum pH, temperature stability and catalytic efficiency of the two proteins. The V. salmonicida EndA shows typical cold‐adapted features such as lower unfolding temperature, lower temperature optimum for activity, and higher specific activity than V. cholerae EndA. The thermodynamic activation parameters confirm the psychrophilic nature of V. salmonicida EndA with a much lower activation enthalpy. The optimal conditions for enzymatic activity coincide well with the corresponding optimal requirements for growth of the organisms, and the enzymes function predominantly as DNases at physiological concentrations of NaCl. The periplasmic or extracellular localization of the enzymes, which renders them constantly exposed to the outer environment of the cell, may explain this fine‐tuning of biochemical properties.
Elongation of polyunsaturated fatty acids in trypanosomatids
Authors: Livore Verónica I. , Tripodi Karina E. J. , Uttaro Antonio D.
Published: December 2006

Leishmania major synthesizes polyunsaturated fatty acids by using Δ6, Δ5 and Δ4 front‐end desaturases, which have recently been characterized [Tripodi KE, Buttigliero LV, Altabe SG & Uttaro AD (2006) FEBS J273, 271–280], and two predicted elongases specific for C18 Δ6 and C20 Δ5 polyunsaturated fatty acids, respectively. Trypanosoma brucei and Trypanosoma cruzi lack Δ6 and Δ5 desaturases but contain Δ4 desaturases, implying that trypanosomes use exogenous polyunsaturated fatty acids to produce C22 Δ4 fatty acids. In order to identify putative precursors of these C22 fatty acids and to completely describe the pathways for polyunsaturated fatty acid biosynthesis in trypanosomatids, we have performed a search in the three genomes and identified four different elongase genes in T. brucei, five in T. cruzi and 14 in L. major. After a phylogenetic analysis of the encoded proteins together with elongases from a variety of other organisms, we selected four candidate polyunsaturated fatty acid elongases. Leishmania major CAJ02037, T. brucei AAX69821 and T. cruzi XP_808770 share 57–52% identity, and group together with C20 Δ5 polyunsaturated fatty acid elongases from algae. The predicted activity was corroborated by functional characterization after expression in yeast. T. brucei elongase was also able to elongate Δ8 and Δ11 C20 polyunsaturated fatty acids. L. major CAJ08636, which shares 33% identity with Mortierella alpinaΔ6 elongase, showed a high specificity for C18 Δ6 polyunsaturated fatty acids. In all cases, a preference for n6 polyunsaturated fatty acids was observed. This indicates that L. major has, as predicted, Δ6 and Δ5 elongases and a complete pathway for polyunsaturated fatty acid biosynthesis. Trypanosomes contain only Δ5 elongases, which, together with Δ4 desaturases, allow them to use eicosapentaenoic acid and arachidonic acid, a precursor that is relatively abundant in the host, for C22 polyunsaturated fatty acid biosynthesis.
An investigation of the substrate specificity of the xyloglucanase Cel74A from Hypocrea jecorina
Authors: Desmet Tom , Cantaert Tineke , Gualfetti Peter , Nerinckx Wim , Gross Laurie , Mitchinson Colin , Piens Kathleen
Published: December 2006

The substrate specificity of the xyloglucanase Cel74A from Hypocrea jecorina (Trichoderma reesei) was examined using several polysaccharides and oligosaccharides. Our results revealed that xyloglucan chains are hydrolyzed at substituted Glc residues, in contrast to the action of all known xyloglucan endoglucanases (EC 3.2.1.151). The building block of xyloglucan, XXXG (where X is a substituted Glc residue, and G is an unsubstituted Glc residue), was rapidly degraded to XX and XG (kcat = 7.2 s−1 and Km = 120 µm at 37 °C and pH 5), which has only been observed before with the oligoxyloglucan‐reducing‐end‐specific cellobiohydrolase from Geotrichum (EC 3.2.1.150). However, the cellobiohydrolase can only release XG from XXXGXXXG, whereas Cel74A hydrolyzed this substrate at both chain ends, resulting in XGXX. Differences in the length of a specific loop at subsite + 2 are discussed as being the basis for the divergent specificity of these xyloglucanases.
Inhibition of pneumococcal choline‐binding proteins and cell growth by esters of bicyclic amines
Authors: Maestro Beatriz , González Ana , García Pedro , Sanz Jesús M
Published: December 2006

Streptococcus pneumoniae is one of the major pathogens worldwide. The use of currently available antibiotics to treat pneumococcal diseases is hampered by increasing resistance levels; also, capsular polysaccharide‐based vaccination is of limited efficacy. Therefore, it is desirable to find targets for the development of new antimicrobial drugs specifically designed to fight pneumococcal infections. Choline‐binding proteins are a family of polypeptides, found in all S. pneumoniae strains, that take part in important physiologic processes of this bacterium. Among them are several murein hydrolases whose enzymatic activity is usually inhibited by an excess of choline. Using a simple chromatographic procedure, we have identified several choline analogs able to strongly interact with the choline‐binding module (C‐LytA) of the major autolysin of S. pneumoniae. Two of these compounds (atropine and ipratropium) display a higher binding affinity to C‐LytA than choline, and also increase the stability of the protein. CD and fluorescence spectroscopy analyses revealed that the conformational changes of C‐LytA upon binding of these alkaloids are different to those induced by choline, suggesting a different mode of binding. In vitro inhibition assays of three pneumococcal, choline‐dependent cell wall lytic enzymes also demonstrated a greater inhibitory efficiency of those molecules. Moreover, atropine and ipratropium strongly inhibited in vitro pneumococcal growth, altering cell morphology and reducing cell viability, a very different response than that observed upon addition of an excess of choline. These results may open up the possibility of the development of bicyclic amines as new antimicrobials for use against pneumococcal pathologies.
Crystal structures of a bacterial 6‐phosphogluconate dehydrogenase reveal aspects of specificity, mechanism and mode of inhibition by analogues of high‐energy reaction intermediates
Authors: Sundaramoorthy Ramasubramanian , Iulek Jorge , Barrett Michael P. , Bidet Olivier , Ruda Gian Filippo , Gilbert Ian H. , Hunter William N.
Published: December 2006

Crystal structures of recombinant Lactococcus lactis 6‐phosphogluconate dehydrogenase (LlPDH) in complex with substrate, cofactor, product and inhibitors have been determined. LlPDH shares significant sequence identity with the enzymes from sheep liver and the protozoan parasite Trypanosoma brucei for which structures have been reported. Comparisons indicate that the key residues in the active site are highly conserved, as are the interactions with the cofactor and the product ribulose 5‐phosphate. However, there are differences in the conformation of the substrate 6‐phosphogluconate which may reflect distinct states relevant to catalysis. Analysis of the complex formed with the potent inhibitor 4‐phospho‐d‐erythronohydroxamic acid, suggests that this molecule does indeed mimic the high‐energy intermediate state that it was designed to. The analysis also identified, as a contaminant by‐product of the inhibitor synthesis, 4‐phospho‐d‐erythronamide, which binds in similar fashion. LlPDH can now serve as a model system for structure‐based inhibitor design targeting the enzyme from Trypanosoma species.
Histidine‐rich glycoprotein exerts antibacterial activity
Authors: Rydengård Victoria , Olsson Anna‐Karin , Mörgelin Matthias , Schmidtchen Artur
Published: December 2006

Histidine‐rich glycoprotein (HRGP), an abundant heparin‐binding protein found in plasma and thrombocytes, exerts antibacterial effects against Gram‐positive bacteria (Enterococcus faecalis and Staphylococcus aureus) and Gram‐negative bacteria (Escherichia coli and Pseudomonas aeruginosa). Fluorescence studies and electron microscopy to assess membrane permeation showed that HRGP induces lysis of E. faecalisbacteria in the presence of Zn2+ or at low pH. Heparin blocked binding of the protein to E. faecalis and abolished antibacterial activity. Furthermore, truncated HRGP, devoid of the heparin‐binding and histidine‐rich domain, was not antibacterial. It has previously been shown that peptides containing consensus heparin‐binding sequences (Cardin and Weintraub motifs) are antibacterial. Thus, the peptide (GHHPH)4, derived from the histidine‐rich region of HRGP and containing such a heparin‐binding motif, was antibacterial for E. faecalis in the presence of Zn2+ or at low pH. The results show a previously undisclosed antibacterial activity of HRGP and suggest that the histidine‐rich and heparin‐binding domain of HRGP mediates the antibacterial activity of the protein.
A polyketide synthase of Plumbago indica that catalyzes the formation of hexaketide pyrones
Authors: Springob Karin , Samappito Supachai , Jindaprasert Aphacha , Schmidt Jürgen , Page Jonathan E , De‐Eknamkul Wanchai , Kutchan Toni M
Published: December 2006

Plumbago indica L. contains naphthoquinones that are derived from six acetate units. To characterize the enzyme catalyzing the first step in the biosynthesis of these metabolites, a cDNA encoding a type III polyketide synthase (PKS) was isolated from roots of P. indica. The translated polypeptide shared 47–60% identical residues with PKSs from other plant species. Recombinant P. indica PKS expressed in Escherichia coli accepted acetyl‐CoA as starter and carried out five decarboxylative condensations with malonyl coenzyme A (‐CoA). The resulting hexaketide was not folded into a naphthalene derivative. Instead, an α‐pyrone, 6‐(2′,4′‐dihydroxy‐6′‐methylphenyl)‐4‐hydroxy‐2‐pyrone, was produced. In addition, formation of α‐pyrones with linear keto side chains derived from three to six acetate units was observed. As phenylpyrones could not be detected in P. indica roots, we propose that the novel PKS is involved in the biosynthesis of naphthoquinones, and additional cofactors are probably required for the biosynthesis of these secondary metabolites in vivo.
Purification, microsequencing and cloning of spinach ATP‐dependent phosphofructokinase link sequence and function for the plant enzyme
Authors: Winkler Christian , Delvos Britta , Martin William , Henze Katrin
Published: December 2006

Despite its importance in plant metabolism, no sequences of higher plant ATP‐dependent phosphofructokinase (EC 2.7.1.11) are annotated in the databases. We have purified the enzyme from spinach leaves 309‐fold to electrophoretic homogeneity. The purified enzyme was a homotetramer of ∼52 kDa subunits with a specific activity of 600 mU·mg−1 and a Km value for ATP of 81 µm. The purified enzyme was not activated by phosphate, but slightly inhibited instead, suggesting that it was the chloroplast isoform. The inclusion of adenosine 5′‐(β,γ‐imido)triphosphate was conducive to enzyme activitiy during the purification protocol. The sequences of eight tryptic peptides from the final protein preparation, which did not utilize pyrophosphate as a phosphoryl donor, were determined and an exactly corresponding cDNA was cloned. The sequence of enzymatically active spinach ATP‐dependent phosphofructokinase suggests that a large family of genomics‐derived higher plant sequences currently annotated in the databases as putative pyrophosphate‐dependent phosphofructokinases according to sequence similarity is misannotated with respect to the cosubstrate.
cGMP transport by vesicles from human and mouse erythrocytes
Authors: de Wolf Cornelia J. F. , Yamaguchi Hiroaki , van der Heijden Ingrid , Wielinga Peter R. , Hundscheid Stefanie L. , Ono Nobuhito , Scheffer George L. , de Haas Marcel , Schuetz John D. , Wijnholds Jan , Borst Piet
Published: December 2006

cGMP secretion from cells can be mediated by ATP‐binding cassette (ABC) transporters ABCC4, ABCC5, and ABCC11. Indirect evidence suggests that ABCC4 and ABCC5 contribute to cGMP transport by erythrocytes. We have re‐investigated the issue using erythrocytes from wild‐type and transporter knockout mice. Murine wild‐type erythrocyte vesicles transported cGMP with an apparentKmthat was 100‐fold higher than their human counterparts, the apparentVmaxbeing similar. Whereas cGMP transport into human vesicles was efficiently inhibited by the ABCC4‐specific substrate prostaglandin E1, cGMP transport into mouse vesicles was inhibited equally by Abcg2 and Abcc4 inhibitors/substrates. Similarly, cGMP transport into vesicles fromAbcc4–/–andAbcg2–/–mice was 42% and 51% of that into wild‐type mouse vesicles, respectively, whereas cGMP transport into vesicles fromAbcc4–/–/Abcg2–/–mice was near background. The knockout mice were used to show that Abcg2‐mediated cGMP transport occurred with lower affinity but higherVmaxthan Abcc4‐mediated transport. Involvement of Abcg2 in cGMP transport byAbcc4–/–erythrocyte vesicles was supported by higher transport at pH 5.5 than at pH 7.4, a characteristic of Abcg2‐mediated transport. The relative contribution of ABCC4/Abcc4 and ABCG2/Abcg2 in cGMP transport was confirmed with a new inhibitor of ABCC4 transport, the protease inhibitor 4‐(2‐aminoethyl)benzenesulfonyl fluoride.
Purification and characterization of the cysteine proteinases in the latex of Vasconcellea spp.
Authors: Kyndt Tina , Van Damme Els J. M. , Van Beeumen Jozef , Gheysen Godelieve
Published: December 2006

Latex of all Vasconcellea species analyzed to date exhibits higher proteolytic amidase activities, generally attributed to cysteine proteinases, than the latex of Carica papaya. In the present study, we show that this higher activity is correlated with a higher concentration of enzymes in the latex of Vasconcellea fruits, but in addition also results from the presence of other cysteine proteinases or isoforms. In contrast to the cysteine proteinases present in papaya latex, which have been extensively studied, very little is known about the cysteine proteinases of Vasconcellea spp. In this investigation, several cDNA sequences coding for cysteine proteinases in Vasconcellea × heilbornii and Vasconcellea stipulata were determined using primers based on conserved sequences. In silico translation showed that they hold the characteristic features of all known papain‐class cysteine proteinases, and a phylogenetic analysis revealed the existence of several papain and chymopapain homologues in these species. Ion‐exchange chromatography and gel filtration procedures were applied on latex of V. × heilbornii in order to characterize its cysteine proteinases at the protein level. Five major protein fractions (VXH‐I–VXH‐V) revealing very high amidase activities (between 7.5 and 23.3 nkat·mg protein−1) were isolated. After further purification, three of them were N‐terminally sequenced. The observed microheterogeneity in the N‐terminal and cDNA sequences reveals the presence of several distinct cysteine proteinase isoforms in the latex of Vasconcellea spp.
Authentic interdomain communication in an RNA helicase reconstituted by expressed protein ligation of two helicase domains
Authors: Karow Anne R. , Theissen Bettina , Klostermeier Dagmar
Published: December 2006

RNA helicases mediate structural rearrangements of RNA or RNA–protein complexes at the expense of ATP hydrolysis. Members of the DEAD box helicase family consist of two flexibly connected helicase domains. They share nine conserved sequence motifs that are involved in nucleotide binding and hydrolysis, RNA binding, and helicase activity. Most of these motifs line the cleft between the two helicase domains, and extensive communication between them is required for RNA unwinding. The two helicase domains of the Bacillus subtilis RNA helicase YxiN were produced separately as intein fusions, and a functional RNA helicase was generated by expressed protein ligation. The ligated helicase binds adenine nucleotides with very similar affinities to the wild‐type protein. Importantly, its intrinsically low ATPase activity is stimulated by RNA, and the Michaelis–Menten parameters are similar to those of the wild‐type. Finally, ligated YxiN unwinds a minimal RNA substrate to an extent comparable to that of the wild‐type helicase, confirming authentic interdomain communication.
Effect of oculopharyngeal muscular dystrophy‐associated extension of seven alanines on the fibrillation properties of the N‐terminal domain of PABPN1
Authors: Lodderstedt Grit , Hess Simone , Hause Gerd , Scheuermann Till , Scheibel Thomas , Schwarz Elisabeth
Published: December 2006

Oculopharyngeal muscular dystrophy (OPMD) is an autosomal dominant disease that usually manifests itself within the fifth decade. The most prominent symptoms are progressive ptosis, dysphagia, and proximal limb muscle weakness. The disorder is caused by trinucleotide (GCG) expansions in the N‐terminal part of the poly(A)‐binding protein 1 (PABPN1) that result in the extension of a 10‐alanine segment by up to seven more alanines. In patients, biopsy material displays intranuclear inclusions consisting primarily of PABPN1. Poly l‐alanine‐dependent fibril formation was studied using the recombinant N‐terminal domain of PABPN1. In the case of the protein fragment with the expanded poly l‐alanine sequence [N‐(+7)Ala], fibril formation could be induced by low amounts of fragmented fibrils serving as seeds. Besides homologous seeds, seeds derived from fibrils of the wild‐type fragment (N‐WT) also accelerated fibril formation of N‐(+7)Ala in a concentration‐dependent manner. Seed‐induced fibrillation of N‐WT was considerably slower than that of N‐(+7)Ala. Using atomic force microscopy, differences in fibril morphologies between N‐WT and N‐(+7)Ala were detected. Furthermore, fibrils of N‐WT showed a lower resistance against solubilization with the chaotropic agent guanidinium thiocyanate than those from N‐(+7)Ala. Our data clearly reveal biophysical differences between fibrils of the two variants that are likely caused by divergent fibril structures.
All‐trans‐retinoic acid inhibits collapsin response mediator protein‐2 transcriptional activity during SH‐SY5Y neuroblastoma cell differentiation
Authors: Fontán‐Gabás Lorena , Oliemuller Erik , Martínez‐Irujo Juan José , de Miguel Carlos , Rouzaut Ana
Published: December 2006

Neurons are highly polarized cells composed of two structurally and functionally distinct parts, the axon and the dendrite. The establishment of this asymmetric structure is a tightly regulated process. In fact, alterations in the proteins involved in the configuration of the microtubule lattice are frequent in neuro‐oncologic diseases. One of these cytoplasmic mediators is the protein known as collapsin response mediator protein‐2, which interacts with and promotes tubulin polymerization. In this study, we investigated collapsin response mediator protein‐2 transcriptional regulation during all‐trans‐retinoic acid‐induced differentiation of SH‐SY5Y neuroblastoma cells. All‐trans‐retinoic acid is considered to be a potential preventive and therapeutic agent, and has been extensively used to differentiate neuroblastoma cells in vitro. Therefore, we first demonstrated that collapsin response mediator protein‐2 mRNA levels are downregulated during the differentiation process. After completion of deletion construct analysis and mutagenesis and mobility shift assays, we concluded that collapsin response mediator protein‐2 basal promoter activity is regulated by the transcription factors AP‐2 and Pax‐3, whereas E2F, Sp1 and NeuroD1 seem not to participate in its regulation. Furthermore, we finally established that reduced expression of collapsin response mediator protein‐2 after all‐trans‐retinoic acid exposure is associated with impaired Pax‐3 and AP‐2 binding to their consensus sequences in the collapsin response mediator protein‐2 promoter. Decreased attachment of AP‐2 is a consequence of its accumulation in the cytoplasm. On the other hand, Pax‐3 shows lower binding due to all‐trans‐retinoic acid‐mediated transcriptional repression. Unraveling the molecular mechanisms behind the action of all‐trans‐retinoic acid on neuroblastoma cells may well offer new perspectives for its clinical application.
Attenuation of Oxidative Stress in Plasma and Tissues of Rats with Experimentally Induced Hyperthyroidism by Caffeic Acid Phenylethyl Ester
Authors: Mohamadin Ahmed M. , Hammad Lamiaa N. A. , El‐Bab Mohamed Fath , Abdel Gawad Hala S.
Published: December 2006

Abstract:   Increased oxidative stress with high free radical generation has been described previously in animal models of hyperthyroidism. The present study was designed to investigate the protective effects of caffeic acid phenylethyl ester (CAPE) on oxidative damage in rats with experimentally induced hyperthyroidism. The study was conducted on 32 male Sprague‐Dawley rats. The experimental animals were divided into four groups (control, CAPE alone, hyperthyroidism, and hyperthyroidism + CAPE). Hyperthyroidism was induced by intraperitoneal administration of 0.3 mg/kg/day l‐thyroxine for 4 weeks. CAPE (10 µg/kg) was administered intraperitoneally for 4 weeks. At the end of the experimental period, blood samples and various organs (liver, heart and brain) of rats were taken for the determination of thiobarbituric acid reactive substances (TBARS), reduced glutathione (GSH), oxidized glutathione, vitamin C and superoxide dismutase (SOD) levels and concentrations of triiodothyronine (T3), thyroxine (T4) and thyroxine‐stimulating hormone (TSH). Our results indicate that TBARS, oxidized glutathione, SOD levels and concentrations of T3 and T4 were higher in plasma and tissues of the hyperthyroid group compared to controls. Vitamin C, GSH and TSH levels were decreased significantly in the hyperthyroid group when compared to the control group. CAPE treatment decreased the elevated TBARS, SOD, T3 and T4 levels and increased the lowered GSH, vitamin C and TSH levels to control levels in rats with hyperthyroidism. In conclusion, our results indicate that CAPE is beneficial as a protective agent against oxidative stress induced by hyperthyroidism in rats. The protection is probably due to multiple mechanisms involving free radical scavenger properties, attenuating lipid peroxidation and increasing the antioxidant status.
Vanilloid (Subtype 1) Receptor‐Modulatory Drugs Inhibit [3H]Batrachotoxinin‐A 20‐α‐benzoate Binding to Na+Channels
Authors: Duan Yin , Zheng Jian , Nicholson Russell A.
Published: December 2006

Abstract: This investigation was conducted to provide further insight into the effects of vanilloid (subtype 1) receptor (VR1) drugs at voltage‐gated sodium channels and examine the potential of this interaction to influence release of neurotransmitters from synaptosomes prepared from mammalian brain. The VR1 modulatory drugs capsaicin, olvanil and capsazepine inhibited the binding of batrachotoxinin‐A 20‐α‐benzoate ([3H]BTX‐B) to receptor site 2 of voltage‐gated sodium channels. All drugs reduced the affinity of radioligand for sodium channels, and capsazepine also decreased the number of [3H]BTX‐B binding sites. In kinetic experiments, no reduction in radioligand association rate was found, but capsaicin, olvanil and capsazepine all enhanced the dissociation rate of [3H]BTX‐B. All drugs inhibited veratridine‐evoked release ofl‐glutamic acid, γ‐amino butyric acid andl‐aspartic acid from synaptosomes; however, their inhibitory effects on transmitter release were much weaker when 35 mM potassium chloride was used to depolarize synaptosomes. The study compounds, in common with other central nervous system depressants, interact with a region on the voltage‐gated sodium channel that permits negative allosteric coupling with receptor site 2 and this mechanism likely accounts for blockade of sodium channel‐activated transmitter release.
Role of Ellagic Acid against Cisplatin‐Induced Nephrotoxicity and Oxidative Stress in Rats
Authors: Ateşşahín Ahmet , Çeríbaşi Ali Osman , Yuce Abdurrauf , Bulmus Özgür , Çikim Gürkan
Published: December 2006

Abstract:  The aim of this study was to investigate the possible protective role of antioxidant treatment with ellagic acid on cisplatin‐induced nephrotoxicity using biochemical and histopatological approaches. Adult male Sprague‐Dawley rats were randomly divided into four groups. The control group received 0.9% saline; animals in the ellagic acid group received only ellagic acid (10 mg/kg); animals in the cisplatin group received only cisplatin (7 mg/kg); animals in the cisplatin + ellagic acid group received ellagic acid for 10 days after cisplatin. The effects of ellagic acid on cisplatin‐induced nephrotoxicity were evaluated by plasma creatinine, urea, sodium and calcium concentrations; kidney tissue malondialdehyde, reduced glutathione (GSH), glutathione peroxidase (GSH peroxidase) and catalase activities and histopatological examinations. Administration of cisplatin to rats induced a marked renal failure, characterized by significant increases in plasma creatinine, urea and calcium concentrations. Cisplatin also induced oxidative stress, as indicated by increased kidney tissue concentrations of malondialdehyde, and reduced activities of GSH peroxidase and catalase. Furthermore, treatment with cisplatin caused a marked tubular necrosis, degeneration and desquamation, luminal cast formation, karyomegaly, tubular dilatation, interstitial mononuclear cell infiltration and inter‐tubular haemorrhagia. Ellagic acid markedly reduced elevated plasma creatinine, urea and calcium levels and counteracted the deleterious effects of cisplatin on oxidative stress markers. In the same way, ellagic acid ameliorated cisplatin‐induced pathological changes including tubular necrosis, degeneration, karyomegaly, tubular dilatation when compared to the cisplatin alone group. These results indicate that the antioxidant ellagic acid might have a protective effect against cisplatin‐induced nephrotoxicity and oxidative stress in rat, but not enough to inhibit cisplatin‐induced renal dysfunction.
Additive Cytotoxic Effect of Apoptin and Chemotherapeutic Agents Paclitaxel and Etoposide on Human Tumour Cells
Authors: Olijslagers Sharon J. , Zhang Ying‐Hui , Backendorf Claude , Noteborn Mathieu H. M.
Published: December 2006

Abstract:  Gene therapy experiments in animal models have shown that apoptin expression results in tumour regression without any significant side effects. Therefore, apoptin is regarded as a potential anticancer drug for clinical applications. In this study, we analysed whether chemotherapeutic agents combined with apoptin treatment could result in enhanced cytotoxicity in human tumour cell cultures. Combined treatment with recombinant adenovirus AdAptVP3 expressing apoptin and etoposide clearly showed an additive cytotoxic effect on human osteosarcoma U2OS cells. Paclitaxel treatment combined with apoptin expression significantly inhibited the survival of p53‐positive human osteosarcoma U2OS and non‐small lung carcinoma A549 cells, p53‐negative human osteosarcoma Saos‐2 cells and p53‐mutant human prostate cancer Du145 cells, already at low doses of the chemotherapeutic agent. Our results indicate that the cytotoxicity‐enhancing action by the tumour‐specific apoptin in combination with chemotherapeutic agents might offer an effective and safe antitumour therapeutics.
Diatomaceous earths as alternatives to chemical insecticides in stored grain
Authors: NIKPAY AMIN
Published: December 2006

Abstract
Diatomaceous earth (DE) is a natural inert dust used to control insect pests in stored grain as an alternative to synthetic residual insecticides. Various DE formulations are now registered as a grain protectant or for structural treatment in many different countries throughout the world. The mode of action of DE is through the absorption of cuticular waxes in the insect cuticle, and insect death occurs from desiccation. The main advantages of using DE are its low mammalian toxicity and its stability. The main limitations to widespread commercial use of DE are reduction of the bulk density and flowability of grain, irritant hazards during application and reduction in efficacy at high moisture contents. This paper is an updated review of published results of researches related to the use of DEs and discusses their potential use in large‐scale, commercial storage and in small scale applications.
Sodium channel gene expression in mosquitoes, Aedes albopictus (S.)
Authors: LIU NANNAN , XU QIANG , ZHANG LEE
Published: December 2006

Abstract
A mosquito strain of Aedes albopictus, HAmAalG0, from Huntsville, Alabama, USA, showed a normal susceptibility and low tolerance to permethrin and resmethrin (pyrethroid insecticides) compared to a susceptible Ikaken strain, even though these pyrethroid insecticides have been used in the field for a long period of time in Alabama. Recently, we treated HAmAalG0 in the laboratory with permethrin for five generations and detected no significant change in the level of resistance to permethrin in the selected mosquitoes, HAmAalG5, compared with the parental strain HAmAalG0. We then examined the allelic expression at the L‐to‐F kdr site of the sodium channel gene in the Aedes mosquitoes to address our hypothesis that the L‐to‐F kdr mutation was not present in HAmAalG0 and HAmAalG5 mosquitoes. We found that every tested individual in Ikaken, HAmAalG0, and HAmAalG5 populations expressed a codon of CTA at the L‐to‐F kdr site encoding Leu, strongly corresponding to their susceptibility to insecticides.
Genetic differentiation of Helicoverpa armigera (Hübner) and H. assulta (Guenée) (Lepidoptera: Noctuidae) based on AFLP markers
Authors: MING QING‐LEI , WANG CHEN‐ZHU
Published: December 2006

Abstract
Here we use amplified fragment length polymorphism (AFLP) to assess genetic differentiation of Helicoverpa armigera and H. assulta. The results indicated that both species‐specific fingerprints and cluster analysis showed the ability of AFLP technique to discriminate the two sibling species; among a total 1963 AFLP markers amplified from nine primer combinations: 777 (39.6%) were H. armigera‐specific, 602 (30.7%) were H. assulta‐specific, and 584 (29.7%) were common bands. The mean number of H. armigera‐specific bands was significantly more than that of H. assulta‐specific bands for nine primer combinations (P < 0.05); the intraspecific distance of H. armigera and H. assulta was 0.123 0 and 0.110 7 respectively, and the interspecific distance was 0.178 3. In addition, the percentage of polymorphic loci and estimated average heterozygosity were used to estimate genetic diversity of the two species. This study therefore demonstrates that AFLP analysis is a sensitive and reliable technique to study genetic differentiation and genetic relationships between species and provides sufficient molecular markers for future linkage map construction, location and eventual cloning of genes involved in traits differentiation.
Circadian rhythms of feeding, oviposition, and emergence of the boll weevil (Coleoptera: Curculionidae)
Authors: GREENBERG SHOIL M. , ARMSTRONG J. SCOTT , SÉTAMOU MAMOUDOU , SAPPINGTON THOMAS W. , COLEMAN RANDY J. , LIU TONG‐XIAN
Published: December 2006

Abstract
Circadian rhythm of feeding, oviposition, and emergence of boll weevil adults were determined at five different photophases (24, 14, 12, 10, and 0 hours) and a constant 27°C temperature, 65% RH in the laboratory. Squares from Petri dishes, where they were exposed to boll weevil females, were removed and examined for feeding and oviposition punctures every 4 hours during daylight (0700–1900 h) and every 12 h at night (1900–0700 h) over eight consecutive days. Cohorts of randomly selected egg‐punctured squares were sampled from ovipositing females at 0700, 1100, 1500, and 1900 during 24 hours and under different photophase treatments, and maintained in Petri dishes at 27 ± 1°C, 65% RH. Dishes were observed twice daily (1900 and 0700 h) for adults emerging at day or night. Circadian rhythm of oviposition was not affected by the length of the photophase. The boll weevil has round‐the‐clock circadian rhythm of oviposition, with a daytime preference. We observed that 82.4%‐86.0% of the boll weevil eggs were deposited between 0700 and 1900 h, and 14.0%‐17.6% between 1900 and 0700 h during a 24‐h period. Feeding of boll weevil females in photoperiods 24: 0 h (complete light) and 0: 24 h (complete darkness) did not significantly change between 0700–1900 h versus 1900–0700 h, while the daily cycle of light and darkness in other photoperiods significantly increased the feeding punctures from 0700–1900 compared with 1900–0700 h. The circadian rhythm of emergence depended significantly on the time of oviposition and the length of the photophase. Investigation of boll weevil circadian rhythm provides a better understanding of boll weevil ecology and reveals potential weak links for improving control technologies targeting their reproductive strategies.