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Crucial Step (crucial + step)
Selected AbstractsExperimental superficial candidiasis on tissue modelsMYCOSES, Issue 4 2010J. A. M. S. Jayatilake Summary Candida species are common pathogens causing superficial mycoses primarily affecting the mucosa and the skin in humans. Crucial steps during pathogenesis of superficial candidiasis comprise fungal adhesion, colonisation and subsequent penetration of the respective tissues. Exploring these pathological events and perhaps fungal and tissue responses towards drug treatment is imperative in the management of this infection. Unfortunately, pathological biopsies of superficial candidiasis do not exhibit the early changes in the host,pathogen interaction as the tissues are already invaded by the fungi. In vivo experimental assessments of pathological processes of superficial candidiasis are also limited because of the difficulties in providing reproducible and comparable conditions in the host environment. Conversely, in vitro models have helped studying fungal,host interactions under more defined and controlled conditions. Some common in vitro models used to simulate superficial candidiasis are chick chorioallantoic membrane, mucosal explants and single layer or multiple layer cell cultures. Interestingly, these experimental approaches share advantages as well as disadvantages when compared with in vivo conditions. Hence, this review intends to discuss about the experimental superficial candidiasis produced in various tissue models and their advantages as well as disadvantages with a particular reference to further improvement of validity and reliability of such experiments. [source] An EasyGrid portal for scheduling system-aware applications on computational GridsCONCURRENCY AND COMPUTATION: PRACTICE & EXPERIENCE, Issue 6 2006C. Boeres Abstract One of the objectives of computational Grids is to offer applications the collective computational power of distributed but typically shared heterogeneous resources. Unfortunately, efficiently harnessing the performance potential of such systems (i.e. how and where applications should execute on the Grid) is a challenging endeavor due principally to the very distributed, shared and heterogeneous nature of the resources involved. A crucial step towards solving this problem is the need to identify both an appropriate scheduling model and scheduling algorithm(s). This paper presents a tool to aid the design and evaluation of scheduling policies suitable for efficient execution of system-aware parallel applications on computational Grids. Copyright © 2005 John Wiley & Sons, Ltd. [source] Simultaneous flow cytometric detection of basophil activation marker CD63 and intracellular phosphorylated p38 mitogen-activated protein kinase in birch pollen allergy,CYTOMETRY, Issue 1 2009Nicolaas E. Aerts Abstract Background: Phosphorylation of p38 MAPK is a crucial step in IgE-receptor signaling in basophils. The relation of p38 MAPK to the well-validated diagnostic cell surface marker CD63 has not been evaluated in a clinical allergy model. Methods: Expression of CD63 and phosphorylation of p38 MAPK were analyzed flow cytometrically in anti-IgE-gated basophils from 18 birch pollen allergic patients, five grass pollen allergic patients, and five healthy individuals, after 3 and 20 min of stimulation with recombinant major birch pollen allergen (rBet v 1). Additional time points and the influence of p38 MAPK inhibitor SB203580 were studied in birch pollen allergic patients. Results: Phospho-p38 MAPK and CD63 were expressed dose-dependently in birch pollen allergic patient basophils within 1 minute of rBet v 1 stimulation. P38 MAPK phosphorylation was fastest and subsided gradually while CD63 expression remained elevated for at least 20 min. Inhibition of p38 MAPK significantly inhibited CD63 upregulation. With optimal stimulation of the cells (1 ,g/mL), sensitivity and specificity for the discrimination between patients and a group of control individuals (grass pollen allergic patients and healthy controls) were 94% and 100% for CD63 at 3 and 20 min and for phospho-p38 MAPK at 3 min. Conclusion: Antigen-induced p38 MAPK phosphorylation in human basophils essentially contributes to CD63 upregulation. It is a sensitive and specific intracellular marker for allergy diagnosis and offers new insight into the mechanisms of basophil activation. © 2008 Clinical Cytometry Society [source] Energy tax harmonization in the European Union: a proposal based on the internalization of environmental external costsENVIRONMENTAL POLICY AND GOVERNANCE, Issue 1 2002Susanna Dorigoni Energy tax harmonization represents a fundamental target within the European Union. In fact fiscal harmonization is a crucial step towards the creation of a single market. In this article the possibility of achieving such an objective is discussed. The paper consists of two sections. In the first the European taxation on energy products is analysed. This analysis is useful in showing the differences that exist between the European countries that account for the difficulties met so far in the process of harmonization. In this respect we comment on the recent proposal of the Directive of the European Union, which lays down the obligation of minimum levels of taxation in all European member states. In the second section, after simulating the effects related to the adoption of a common environmental taxation (a first best solution based on the internalization of environmental external costs), we propose, as a second best solution, an excise tax harmonization model taking into consideration the specificity of each country and being, as far as possible, coherent with the environmental objective. This model proposes: the introduction of a minimum level of taxation on all products equal to the external cost due to the greenhouse effect (a common carbon tax); the possibility, given to the member states, of deviating from such minimum levels, in accordance with their specific requirements, internalizing in the price of the different products, by means of taxes additional to that CO2 minimum, the external costs associated with other pollutant agents (the same in all countries); the opportunity, in case it should be necessary to exceed the entire external cost, for the member states to apply increases that are in accordance with the environmental objective. Copyright © 2002 John Wiley & Sons, Ltd. and ERP Environment [source] The crystal structure of pyruvate decarboxylase from Kluyveromyces lactisFEBS JOURNAL, Issue 18 2006Implications for the substrate activation mechanism of this enzyme The crystal structure of pyruvate decarboxylase from Kluyveromyces lactis has been determined to 2.26 Å resolution. Like other yeast enzymes, Kluyveromyces lactis pyruvate decarboxylase is subject to allosteric substrate activation. Binding of substrate at a regulatory site induces catalytic activity. This process is accompanied by conformational changes and subunit rearrangements. In the nonactivated form of the corresponding enzyme from Saccharomyces cerevisiae, all active sites are solvent accessible due to the high flexibility of loop regions 106,113 and 292,301. The binding of the activator pyruvamide arrests these loops. Consequently, two of four active sites become closed. In Kluyveromyces lactis pyruvate decarboxylase, this half-side closed tetramer is present even without any activator. However, one of the loops (residues 105,113), which are flexible in nonactivated Saccharomyces cerevisiae pyruvate decarboxylase, remains flexible. Even though the tetramer assemblies of both enzyme species are different in the absence of activating agents, their substrate activation kinetics are similar. This implies an equilibrium between the open and the half-side closed state of yeast pyruvate decarboxylase tetramers. The completely open enzyme state is favoured for Saccharomyces cerevisiae pyruvate decarboxylase, whereas the half-side closed form is predominant for Kluyveromyces lactis pyruvate decarboxylase. Consequently, the structuring of the flexible loop region 105,113 seems to be the crucial step during the substrate activation process of Kluyveromyces lactis pyruvate decarboxylase. [source] Accumulation of hydrogen peroxide is an early and crucial step for paclitaxel-induced cancer cell death both in vitro and in vivo,INTERNATIONAL JOURNAL OF CANCER, Issue 1 2006Jérôme Alexandre Abstract Intracellular events following paclitaxel binding to microtubules that lead to cell death remain poorly understood. Because reactive oxygen species (ROS) are involved in the cytotoxicity of anticancer agents acting through independent molecular targets, we explored the role of ROS in paclitaxel cytotoxicity. Within 15 min after in vitro exposure of A549 human lung cancer cells to paclitaxel, a concentration-dependent intracellular increase in O°2, and H2O2 levels was detected by spectrofluorometry. Addition of N -acetylcysteine (NAC) or glutathione, two H2O2 scavenger, induced a 4-fold increase in paclitaxel IC50. Delaying NAC co-incubation by 4 hr, resulted in a 3-fold reduction in cell protection. The glutathione synthesis inhibitor, buthionine sulfoximine significantly increased paclitaxel cytotoxicity and H2O2 accumulation, but did not modify O°2, levels. Co-incubation with diphenylene iodonium suggested that paclitaxel induced-O°2, production was in part associated with increased activity of cytoplasmic NADPH oxidase. Concomitant treatment with inhibitors of caspases 3 and 8 increased cell survival but did not prevent the early accumulation of H2O2. To evaluate the role of ROS in paclitaxel antitumoral activity, mice were injected with LLC1 lung cancer cells and treated with paclitaxel i.p. and/or NAC. The antitumoral activity of paclitaxel in mice was abolished by NAC. In conclusion, the accumulation of H2O2 is an early and crucial step for paclitaxel-induced cancer cell death before the commitment of the cells into apoptosis. These results suggest that ROS participate in vitro and in vivo to paclitaxel cytotoxicity. © 2006 Wiley-Liss, Inc. [source] Testing whether the epidural works: too time consuming?ACTA ANAESTHESIOLOGICA SCANDINAVICA, Issue 6 2010J. LARSSON Background: When using epidural anaesthesia (EDA) for pain relief after major surgery, a failure rate of 10% is common. A crucial step in improving the care of patients with EDA is to define the position of the epidural catheter. The aim of this study was to investigate how much time it takes to determine whether the block is sufficient by assessing the extent of loss of cold sensation before induction of anaesthesia. Methods: One hundred patients listed for abdominal surgery were included in the study. After an epidural catheter had been inserted and an intrathecal or an intravenous position had been made unlikely by the use of a test dose, the patient was given a bolus dose of local anaesthetic plus an opioid in the epidural catheter. The epidural block was tested every 2 min, starting at 5 min and ending at 15 min. When at least four segments were blocked bilaterally, the testing was stopped, the time was noted and the patient was anaesthetised. Results: An epidural block was demonstrated after 5,6 min in 37 patients, after 7,8 min in 43 additional patients and after 9,10 min in 15 patients. In one patient, it took 12 min and in three patients, it took 15 min. In two patients, no epidural block could be demonstrated. Conclusion: Testing an epidural anaesthetic before the induction of anaesthesia takes only 5,10 extra minutes. Knowing whether the catheter is correctly placed means better quality of care, giving the anaesthetist better prerequisites for taking care of the patient post-operatively. [source] On the small signal modeling of advanced microwave FETs: A comparative studyINTERNATIONAL JOURNAL OF RF AND MICROWAVE COMPUTER-AIDED ENGINEERING, Issue 5 2008Giovanni Crupi Abstract Although many successful techniques have been proposed in the last decades for extracting the small signal equivalent circuit for microwave transistors from scattering parameter measurements, small signal modeling is still object of intense research. Further improvement and development of the proposed methods are incessantly required to take into account the continuous and rapid evolution of the transistor technology. The purpose of this article is to facilitate the choice of the most appropriate strategy for each particular case. For that, we present a brief but thorough comparative study of analytical techniques developed for modeling different types of advanced microwave transistors: GaAs HEMTs, GaN HEMTs, and FinFETs. It will be shown that a crucial step for a successful modeling is to adapt accurately the small signal equivalent circuit topology under "cold" condition to each investigated technology. © 2008 Wiley Periodicals, Inc. Int J RF and Microwave CAE, 2008. [source] Efficient, Enantioselective Organocatalytic Synthesis of Trichostatin AADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 10-11 2006Shilei Zhang Abstract An efficient, highly stereocontrolled total synthesis of trichostatin A (1) has been achieved in 9 steps with 17.4,% overall yield and >99,% optical purity from readily available achiral starting materials. The key features of this synthesis include the L -proline-promoted, highly enantioselective cross-aldol reaction as a crucial step for the construction of the C-6 chiral center and the minimization of racemization by final step oxidation of the OH group to a ketone at position 7. [source] Expression of gangliosides in an immortalized neural progenitor/stem cell lineJOURNAL OF NEUROSCIENCE RESEARCH, Issue 5 2003Keiji Suetake Abstract Glycosphingolipids (GSLs) are known to play important roles in cellular growth and differentiation in the nervous system. The change in expression of gangliosides is correlated with crucial developmental events and is evolutionarily conserved among many vertebrate species. The emergence of neural progenitors represents a crucial step in neural development, but little is known about the exact composition and subcellular localization of gangliosides in neural progenitor cells. The C17.2 cell line was derived after v- myc transformation of neural progenitor cells isolated from neonatal mouse cerebellar cortex. The developmental potential of C17.2 cells is similar to that of endogenous neural progenitor/stem cells in that they are multipotential and capable of differentiating into all neural cell types. We characterized the GSL composition of C17.2 cells and found the presence of only a-series gangliosides. Subcellular localization studies revealed that GM1 and GD1a are localized mainly on the plasma membrane and partly in the cytoplasm, both as punctate clusters. Reverse transcription-polymerase chain reaction revealed the absence of ST-II transcripts in C17 cells, which most likely accounts for the lack of expression of b- and c-series complex gangliosides in this cell line. These data suggest that the divergence in ganglioside expression in C17.2 cells is regulated at the transcriptional level. © 2003 Wiley-Liss, Inc. [source] Evaluation and optimisation of five different extraction methods for soy DNA in chocolate and biscuits.JOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 11 2004Extraction of DNA as a first step in GMO analysis Abstract A method is described to discriminate between genetically modified (GM) and non-modified foodstuffs by detecting the presence of newly introduced genes at the protein or DNA level. Currently available methods operate almost exclusively at the DNA level and are based on the polymerase chain reaction (PCR). The first and most crucial step in this process is the isolation of DNA. In this study, five different methods for the isolation of DNA from chocolate and biscuits were evaluated, using four commercially available extraction kits and a non-commercial method for amplification of the soybean-specific lectin gene. The latter method involves the use of hot-start Taq polymerase, to prevent the formation of non-specific amplification products, and an increase in the number of cycles from 35 to 41. The performance of the non-commercial cetyl trimethylammonium bromide (CTAB)-based method was the best, taking into consideration the adaptations of the extraction procedure, although this method was more time-consuming than the others. Chocolate (white, milk and dark) and several biscuits generated positive amplification results using this PCR approach. Copyright © 2004 Society of Chemical Industry [source] The immunological monitoring of alloreactive responses in liver transplant recipients: A reviewLIVER TRANSPLANTATION, Issue 3 2006Raymond Reding The aim of this work is to review the current knowledge in the field of immunological monitoring of allogenic responsiveness in clinical liver transplantation. When compared to other solid-organ transplants, liver allografts are considered as immunologically privileged, and, accordingly, constitute a favorable setting to develop experimental as well as clinical strategies for minimization of immunosuppression and even induction of operational tolerance. The validation of simple, reliable, noninvasive assays exploring antidonor alloreactivity will constitute a crucial step toward implementing such approaches in the clinic. In contrast to research in rodents claiming the development of donor-specific tolerance in case of graft survivals of over 100 days without immunosuppression, it is impractical to confirm tolerance induction in this way in humans. Promising candidate assays include the detection of post-transplant immune deviation, of circulating precursors of dendritic cells subtypes, and of regulatory T cells. A conceptual framework for the development of tolerance assays in clinical liver transplantation is also proposed. Liver Transpl 12:373,383, 2006. © 2006 AASLD. [source] Multiple paternity in reptiles: patterns and processesMOLECULAR ECOLOGY, Issue 11 2008TOBIAS ULLER Abstract The evolution of female promiscuity poses an intriguing problem as benefits of mating with multiple males often have to arise via indirect, genetic, effects. Studies on birds have documented that multiple paternity is common in natural populations but strong evidence for selection via female benefits is lacking. In an attempt to evaluate the evidence more broadly, we review studies of multiple paternity in natural populations of all major groups of nonavian reptiles. Multiple paternity has been documented in all species investigated so far and commonly exists in over 50% of clutches, with particularly high levels in snakes and lizards. Marine turtles and lizards with prolonged pair-bonding have relatively low levels of multiple paternity but levels are nevertheless higher than in many vertebrates with parental care. There is no evidence that high levels of polyandry are driven by direct benefits to females and the evidence that multiple paternity arises from indirect genetic benefits is weak. Instead, we argue that the most parsimonious explanation for patterns of multiple paternity is that it represents the combined effect of mate-encounter frequency and conflict over mating rates between males and females driven by large male benefits and relatively small female costs, with only weak selection via indirect benefits. A crucial step for researchers is to move from correlative approaches to experimental tests of assumptions and predictions of theory under natural settings, using a combination of molecular techniques and behavioural observations. [source] Effect of isoflurane on monocyte adhesion molecule expression in human whole blood,ACTA ANAESTHESIOLOGICA SCANDINAVICA, Issue 5 2003L. W. De Rossi Background: Recruitment of monocytes to inflamed tissue is a crucial step in the acute inflammatory reaction. Adherence of monocytes to endothelial cells followed by transmigration depends on monocyte surface adhesion molecules, inflammatory cytokines and chemoattractant chemokines. In the present study, we determined the effect of isoflurane on monocyte adhesion receptor expression in vitro. Methods: Citrated whole blood was incubated for 60 min with either 0.5 or 1 MAC isoflurane. In unstimulated blood samples and after stimulation with N-formyl-methionyl-leucyl-phenylalanine (FMLP) monocyte cell-surface expression of the selectins PSGL-1 and L-selectin, and the ,2 -integrins CD11a and CD11b were evaluated by flow cytometry. Results: Isoflurane reduced significantly the expression of PSGL-1 on unstimulated monocytes, whereas the remaining selectins and ,2 -integrins were not affected. At both concentrations, the FMLP-induced removal of PSGL-1 from the monocyte surface was increased. Furthermore, at 1 MAC isoflurane the FMLP-induced increase in CD11a expression was significantly inhibited. The surface expression of L-selectin and CD11b was not affected following exposure to isoflurane. Conclusion: Isoflurane increases the removal of the selectin PSGL-1 from the monocyte surface. Since PSGL-1 is important during the initial step of monocyte adhesion to endothelial P-selectin, the decrease in monocyte surface PSGL-1 may have profound effects on monocyte,endothelial interactions. Furthermore, the effects of isoflurane on monocyte adhesion molecule expression are different from those reported for neutrophils. [source] Pesticide residues in food,acute dietary exposure,PEST MANAGEMENT SCIENCE (FORMERLY: PESTICIDE SCIENCE), Issue 4 2004Denis Hamilton Abstract Consumer risk assessment is a crucial step in the regulatory approval of pesticide use on food crops. Recently, an additional hurdle has been added to the formal consumer risk assessment process with the introduction of short-term intake or exposure assessment and a comparable short-term toxicity reference, the acute reference dose. Exposure to residues during one meal or over one day is important for short-term or acute intake. Exposure in the short term can be substantially higher than average because the consumption of a food on a single occasion can be very large compared with typical long-term or mean consumption and the food may have a much larger residue than average. Furthermore, the residue level in a single unit of a fruit or vegetable may be higher by a factor (defined as the variability factor, which we have shown to be typically ×3 for the 97.5th percentile unit) than the average residue in the lot. Available marketplace data and supervised residue trial data are examined in an investigation of the variability of residues in units of fruit and vegetables. A method is described for estimating the 97.5th percentile value from sets of unit residue data. Variability appears to be generally independent of the pesticide, the crop, crop unit size and the residue level. The deposition of pesticide on the individual unit during application is probably the most significant factor. The diets used in the calculations ideally come from individual and household surveys with enough consumers of each specific food to determine large portion sizes. The diets should distinguish the different forms of a food consumed, eg canned, frozen or fresh, because the residue levels associated with the different forms may be quite different. Dietary intakes may be calculated by a deterministic method or a probabilistic method. In the deterministic method the intake is estimated with the assumptions of large portion consumption of a ,high residue' food (high residue in the sense that the pesticide was used at the highest recommended label rate, the crop was harvested at the smallest interval after treatment and the residue in the edible portion was the highest found in any of the supervised trials in line with these use conditions). The deterministic calculation also includes a variability factor for those foods consumed as units (eg apples, carrots) to allow for the elevated residue in some single units which may not be seen in composited samples. In the probabilistic method the distribution of dietary consumption and the distribution of possible residues are combined in repeated probabilistic calculations to yield a distribution of possible residue intakes. Additional information such as percentage commodity treated and combination of residues from multiple commodities may be incorporated into probabilistic calculations. The IUPAC Advisory Committee on Crop Protection Chemistry has made 11 recommendations relating to acute dietary exposure. Copyright © 2004 Society of Chemical Industry [source] Initialization and logic gate operations of nuclear spin qubits using a submicron scale resistively-detected NMR devicePHYSICA STATUS SOLIDI (C) - CURRENT TOPICS IN SOLID STATE PHYSICS, Issue 1 2008T. Ota Abstract We demonstrate initialization of nuclear spin qubits using a resistively-detected nuclear magnetic resonance (NMR) device. In our device, nuclear spins are dynamically polarized in a sub-micron scale region defined by split gates. The population of each nuclear spin state is estimated from a resistively-detected NMR spectrum combined with numerical analysis. By applying radio frequency pulses to the polarized nuclear spins, we create two-qubit effective pure states, which is a crucial step toward NMR quantum computation. We also demonstrate simple logic gate operations such as controlled-NOT and SWAP gates in this two-qubit system. The obtained spectra are consistent with numerically calculated ones. (© 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Circadian rhythms and the evolution of photoperiodic timing in insectsPHYSIOLOGICAL ENTOMOLOGY, Issue 4 2009DAVID S. SAUNDERS Abstract. This review discusses possible evolutionary trends in insect photoperiodism, mainly from a chronobiological perspective. A crucial step was the forging of a link between the hormones regulating diapause and the systems of biological rhythms, circadian or circannual, which have independently evolved in eukaryotes to synchronize physiology and behaviour to the daily cycles of light and darkness. In many of these responses a central feature is that the circadian system resets to a constant phase at the beginning of the subjective night, and then ,measures' the duration of the next scotophase. In ,external coincidence', one version of such a clock, light now has a dual role. First, it serves to entrain the circadian system to the stream of pulses making up the light/dark cycle and, second, it regulates the nondiapause/diapause switch in development by illuminating/not illuminating a specific light sensitive phase falling at the end of the critical night length. Important work by A. D. Lees on the aphid Megoura viciae using so-called ,night interruption experiments' demonstrates that pulses falling early in the night lead to long-day effects that are reversible by a subsequent dark period longer than the critical night length and also show maximal sensitivity in the blue,green range of the spectrum. Pulses falling in the latter half of the night, however, produce long-day effects that are irreversible by a subsequent long-night and show a spectral sensitivity extending into the red. With movement to higher latitudes, insects develop genetic clines in various parameters, including critical night length, the number of long-night cycles needed for diapause induction, the strength of the response, and the ,depth' or intensity of the diapause thus induced. Evidence for these and other types of photoperiodic response suggests that they provided strong selective advantages for insect survival. [source] The L49F mutation in alpha erythroid spectrin induces local disorder in the tetramer association region: Fluorescence and molecular dynamics studies of free and bound alpha spectrinPROTEIN SCIENCE, Issue 9 2009Yuanli Song Abstract The bundling of the N-terminal, partial domain helix (Helix C,) of human erythroid ,-spectrin (,I) with the C-terminal, partial domain helices (Helices A, and B,) of erythroid ,-spectrin (,I) to give a spectrin pseudo structural domain (triple helical bundle A,B,C,) has long been recognized as a crucial step in forming functional spectrin tetramers in erythrocytes. We have used apparent polarity and Stern,Volmer quenching constants of Helix C, of ,I bound to Helices A, and B, of ,I, along with previous NMR and EPR results, to propose a model for the triple helical bundle. This model was used as the input structure for molecular dynamics simulations for both wild type (WT) and ,I mutant L49F. The simulation output structures show a stable helical bundle for WT, but not for L49F. In WT, four critical interactions were identified: two hydrophobic clusters and two salt bridges. However, in L49F, the region downstream of Helix C, was unable to assume a helical conformation and one critical hydrophobic cluster was disrupted. Other molecular interactions critical to the WT helical bundle were also weakened in L49F, possibly leading to the lower tetramer levels observed in patients with this mutation-induced blood disorder. [source] Peroxidative aggregation of ,-synuclein requires tyrosinesPROTEIN SCIENCE, Issue 11 2004Alina Olteanu Abstract ,-Synuclein is the main component of the intracellular protein aggregates in neurons of patients with Parkinson's disease. The occurrence of the disease is associated with oxidative damage. Although it is known that peroxidative chemistry leads to the aggregation of ,-synuclein in vitro, the specific amino acid types of ,-synuclein involved in this type of aggregation have not been identified. We show, using human cytochrome c plus H2O2 as the source oxidative stress, that the tyrosines of ,-synuclein are required for aggregation. The studies reveal the chemical basis for a crucial step in the aggregation process. [source] A novel wavelet-based thresholding method for the pre-processing of mass spectrometry data that accounts for heterogeneous noisePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 15 2008Deukwoo Kwon Dr. Abstract In recent years there has been an increased interest in using protein mass spectroscopy to discriminate diseased from healthy individuals with the aim of discovering molecular markers for disease. A crucial step before any statistical analysis is the pre-processing of the mass spectrometry data. Statistical results are typically strongly affected by the specific pre-processing techniques used. One important pre-processing step is the removal of chemical and instrumental noise from the mass spectra. Wavelet denoising techniques are a standard method for denoising. Existing techniques, however, do not accommodate errors that vary across the mass spectrum, but instead assume a homogeneous error structure. In this paper we propose a novel wavelet denoising approach that deals with heterogeneous errors by incorporating a variance change point detection method in the thresholding procedure. We study our method on real and simulated mass specrometry data and show that it improves on performances of peak detection methods. [source] Membrane microdomains and proteomics: Lessons from tetraspanin microdomains and comparison with lipid raftsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 24 2006François Le Naour Dr. Abstract Biological membranes are compartmentalized into microdomains that exhibit particular lipid and protein compositions. Membrane microdomains, such as tetraspanin-enriched microdomains and lipid rafts, have been suggested to play a role in a variety of physiological and pathological processes. Therefore, the characterization of the protein compositions of these microdomains, which is the focus of this review, appears to be a crucial step to better understanding their function. Proteomics has recently allowed the characterization of tetraspanin-enriched microdomains in colon cancer cells. This demonstrated the presence of different categories of membrane proteins and suggested a variation in the composition of tetraspanin-enriched microdomains during tumor progression. On the other hand, proteomics has permitted the identification of hundreds of proteins in lipid rafts of different origins. However, the diversity of methodologies in sample preparation and of strategies in protein identification led to a broad variability in the data obtained. These methodological issues are discussed. Moreover, proteomics has revealed that different sets of proteins were present in tetraspanin-enriched microdomains as compared with lipid rafts, strengthening the idea that these microdomains are distinct structures. [source] Improved detection of reactive metabolites with a bromine-containing glutathione analog using mass defect and isotope pattern matchingRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 9 2010André LeBlanc Drug bioactivation leading to the formation of reactive species capable of covalent binding to proteins represents an important cause of drug-induced toxicity. Reactive metabolite detection using invitro microsomal incubations is a crucial step in assessing potential toxicity of pharmaceutical compounds. The most common method for screening the formation of these unstable, electrophilic species is by trapping them with glutathione (GSH) followed by liquid chromatography/mass spectrometry (LC/MS) analysis. The present work describes the use of a brominated analog of glutathione, N -(2-bromocarbobenzyloxy)-GSH (GSH-Br), for the invitro screening of reactive metabolites by LC/MS. This novel trapping agent was tested with four drug compounds known to form reactive metabolites, acetaminophen, fipexide, trimethoprim and clozapine. Invitro rat microsomal incubations were performed with GSH and GSH-Br for each drug with subsequent analysis by liquid chromatography/high-resolution mass spectrometry on an electrospray time-of-flight (ESI-TOF) instrument. A generic LC/MS method was used for data acquisition, followed by drug-specific processing of accurate mass data based on mass defect filtering and isotope pattern matching. GSH and GSH-Br incubations were compared to control samples using differential analysis (Mass Profiler) software to identify adducts formed via the formation of reactive metabolites. In all four cases, GSH-Br yielded improved results, with a decreased false positive rate, increased sensitivity and new adducts being identified in contrast to GSH alone. The combination of using this novel trapping agent with powerful processing routines for filtering accurate mass data and differential analysis represents a very reliable method for the identification of reactive metabolites formed in microsomal incubations. Copyright © 2010 John Wiley & Sons, Ltd. [source] OC1 Effects of Cryopreservation on the Expression of Glut-3, Glut-5 and As-A Proteins in Iberian Boar Sperm MembranesREPRODUCTION IN DOMESTIC ANIMALS, Issue 2006S Sancho In order to determine the injure produced in boar spermatozoa through cryopreservation process, we analyzed the expression of the hexose transporters Glut-3 and Glut-5 and the zona pellucida binding protein As-A (P68) in three different steps of the freezing-thawed protocol: at 17°C (fresh BTS-diluted semen, 1 : 2 v/v, step 1), at 5°C (after glycerol addition; step 2), and post-thawing (step 3). All sperm analyses were carried out with immunogold techniques under electronic microscopy. For this study eight healthy post-pubertal Iberian boars were submitted to a collection of twice per week through 3 months, evaluating two ejaculates from each boar. Glut-3 maintains the expression in the acrosome region post-thawing but not along the tail where is reduced. The expression of Glut-5 and As-A is majority located at the post-acrosome region of the spermatozoa at step 1, but in step 2 and step 3 this expression is relocated to sperm tail area. In conclusion, while cryopreservation affects the localization and the expression of Glut-3 and Glut-5, its fertilizing capacity is not significantly reduced. The stabilization of boar semen at 5°C was found to be the most crucial step for sperm survival. [source] ORIGINAL RESEARCH,INTERSEX AND GENDER IDENTITY DISORDERS: A Report from a Single Institute's 14-Year Experience in Treatment of Male-to-Female TranssexualsTHE JOURNAL OF SEXUAL MEDICINE, Issue 10 2009Ciro Imbimbo MD ABSTRACT Introduction., Gender identity disorder or transsexualism is a complex clinical condition, and prevailing social context strongly impacts the form of its manifestations. Sex reassignment surgery (SRS) is the crucial step of a long and complex therapeutic process starting with preliminary psychiatric evaluation and culminating in definitive gender identity conversion. Aim., The aim of our study is to arrive at a clinical and psychosocial profile of male-to-female transsexuals in Italy through analysis of their personal and clinical experience and evaluation of their postsurgical satisfaction levels SRS. Methods., From January 1992 to September 2006, 163 male patients who had undergone gender-transforming surgery at our institution were requested to complete a patient satisfaction questionnaire. Main Outcome Measures., The questionnaire consisted of 38 questions covering nine main topics: general data, employment status, family status, personal relationships, social and cultural aspects, presurgical preparation, surgical procedure, and postsurgical sex life and overall satisfaction. Results., Average age was 31 years old. Seventy-two percent had a high educational level, and 63% were steadily employed. Half of the patients had contemplated suicide at some time in their lives before surgery and 4% had actually attempted suicide. Family and colleague emotional support levels were satisfactory. All patients had been adequately informed of surgical procedure beforehand. Eighty-nine percent engaged in postsurgical sexual activities. Seventy-five percent had a more satisfactory sex life after SRS, with main complications being pain during intercourse and lack of lubrication. Seventy-eight percent were satisfied with their neovagina's esthetic appearance, whereas only 56% were satisfied with depth. Almost all of the patients were satisfied with their new sexual status and expressed no regrets. Conclusions., Our patients' high level of satisfaction was due to a combination of a well-conducted preoperative preparation program, competent surgical skills, and consistent postoperative follow-up. Imbimbo C, Verze P, Palmieri A, Longo N, Fusco F, Arcaniolo D, and Mirone V. A Report from a single institute's fourteen year experience in treatment of male-to-female transsexuals. J Sex Med 2009;6:2736,2745. [source] Dynamic changes in gene expression during vitellogenic stages of the white shrimp: Fenneropenaeus merguiensis de ManAQUACULTURE RESEARCH, Issue 6 2009Monwadee Wonglapsuwan Abstract Ovarian maturation is a crucial step for shrimp brood stock. A suppressive subtractive hybridization was used to identify differentially expressed genes in the ovaries during vitellogenesis of Fenneropenaeus merguiensis. Three- to sevenfold up-regulated genes were selected. A blast search identified nine unique genes. The genes that may be involved in ovarian maturation, namely translationally controlled tumour protein (TCTP), heat shock protein 70 (HSP70), H-L(3)MBT-LIKE, shrimp ovarian peritrophin (SOP), vitellin (Vn), thrombospondin (TSP) and ribosomal protein L10a (RPL10a), were further studied. The transcripts of HSP70, TCTP, SOP and RPL10a in the ovary showed their highest expression in the early stage and declined in the later stages. In contrast, the transcripts of the H-L(3)MBT-LIKE, TSP and Vn genes increased from the early stage to be significantly up-regulated during the late stage. A comparison of gene expression among organs during the vitellogenesis showed that the transcripts of HSP70, SOP, H-L(3)MBT-LIKE and TSP were down-regulated in the brain, intestine, hepatopancreas and lymphoid (except for TSP) when compared with their expression in shrimp with non-developed ovaries. The mRNA of TCTP and RPL10a was significantly over-expressed in the lymphoid and heart, whereas TCTP transcripts were significantly down-regulated in the brain during the vitellogenesis. The molecular behaviour of the transcripts in this study may, in the future, lead to an ability to stimulate the ovarian development in shrimp. [source] Habitat suitability analysis for lacustrine brown trout (Salmo trutta) in Lake Walchensee, Germany: implications for the conservation of an endangered flagship speciesAQUATIC CONSERVATION: MARINE AND FRESHWATER ECOSYSTEMS, Issue 1 2010Marco Denic Abstract 1.The lacustrine brown trout (Salmo trutta) is endangered and of high conservation importance. In the only spawning habitat of the population in the Bavarian Lake Walchensee, the River Obernach, a substantial decrease in spawning runs has been reported. In this study, the present ecological state of the spawning stream was analysed with the objective of identifying life-stage specific limitations to successful recruitment attributable to deficiencies in (i) spawning migration, (ii) spawning habitat quality, and (iii) habitat quality for juveniles. 2.Structural stream analysis showed that discharge and several migration barriers , particularly near the river outlet into the lake , prevent successful spawning migrations at normal water levels. Migration barriers are probably the main limiting factor for reproduction of lacustrine brown trout, whereas structural variability of the Obernach meets the habitat requirements of both spawners and juveniles. 3.Spawning site quality was suitable for trout, as indicated by stream substratum texture and high exchange rates between free-flowing water and the interstitial zone in physico-chemical parameters (redox potential, dissolved oxygen, pH, temperature and conductivity). 4.Analyses of fish community structure revealed dominance of lithophilic species, in particular of riverine brown trout (Salmo trutta). Its density and intact demographic population structure suggest that spawning and juvenile habitat quality for salmonids is not limiting. Recapture of stocked lacustrine trout juveniles also indicates habitat suitability for the juvenile stage. 5.In conclusion, the results show that the methodology used in this study is suitable for the identification of life-stage specific habitat deficiencies in lacustrine brown trout and other fish species. Availability of habitat data throughout the species' distribution range is a first crucial step for the development of an effective recovery plan. Copyright © 2009 John Wiley & Sons, Ltd. [source] Escherichia coli tRNAArg acceptor-stem isoacceptors: comparative crystallization and preliminary X-ray diffraction analysisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2009André Eichert The aminoacylation of tRNA is a crucial step in cellular protein biosynthesis. Recognition of the cognate tRNA by the correct aminoacyl-tRNA synthetase is ensured by tRNA identity elements. In tRNAArg, the identity elements consist of the anticodon, parts of the D-loop and the discriminator base. The minor groove of the aminoacyl stem interacts with the arginyl-tRNA synthetase. As a consequence of the redundancy of the genetic code, six tRNAArg isoacceptors exist. In the present work, three different Escherichia coli tRNAArg acceptor-stem helices were crystallized. Two of them, the tRNAArg microhelices RR-1660 and RR-1662, were examined by X-ray diffraction analysis and diffracted to 1.7 and 1.8,Å resolution, respectively. The tRNAArg RR-1660 helix crystallized in space group P1, with unit-cell parameters a = 26.28, b = 28.92, c = 29.00,Å, , = 105.74, , = 99.01, , = 97.44°, whereas the tRNAArg RR-1662 helix crystallized in space group C2, with unit-cell parameters a = 33.18, b = 46.16, c = 26.04,Å, , = 101.50°. [source] High-throughput system for determining dissolution kinetics of inclusion bodiesBIOTECHNOLOGY JOURNAL, Issue 5 2009Astrid Dürauer Dr. Abstract Efficient solubilization is a crucial step during inclusion body processing and dissolving conditions were usually empirically established. Here we describe a new methodology for rapid screening of solubilization conditions and evaluation of dissolution kinetics in microtiter plates. Increase of protein in solution over time was directly related to decrease of turbidity measured by absorbance at 600 nm. Dissolution kinetics of inclusion bodies were described by a first-order reaction kinetics, which was used for drug dissolution modeling. Reaction constants were in the range of 0.01,0.03 s,1 for buffer conditions providing sufficient solubilization power. This method is not limited to the screening of optimal buffer conditions for solubilization and can be applied for studying other parameters involved in the solubility of IBs, such as pI of the protein, influence of fermentation conditions, influence of initial protein concentration, and more. [source] Prevention of medication errors: detection and auditBRITISH JOURNAL OF CLINICAL PHARMACOLOGY, Issue 6 2009Germana Montesi 1. Medication errors have important implications for patient safety, and their identification is a main target in improving clinical practice errors, in order to prevent adverse events. 2. Error detection is the first crucial step. Approaches to this are likely to be different in research and routine care, and the most suitable must be chosen according to the setting. 3. The major methods for detecting medication errors and associated adverse drug-related events are chart review, computerized monitoring, administrative databases, and claims data, using direct observation, incident reporting, and patient monitoring. All of these methods have both advantages and limitations. 4. Reporting discloses medication errors, can trigger warnings, and encourages the diffusion of a culture of safe practice. Combining and comparing data from various and encourages the diffusion of a culture of safe practice sources increases the reliability of the system. 5. Error prevention can be planned by means of retroactive and proactive tools, such as audit and Failure Mode, Effect, and Criticality Analysis (FMECA). Audit is also an educational activity, which promotes high-quality care; it should be carried out regularly. In an audit cycle we can compare what is actually done against reference standards and put in place corrective actions to improve the performances of individuals and systems. 6. Patient safety must be the first aim in every setting, in order to build safer systems, learning from errors and reducing the human and fiscal costs. [source] Fallen Women and Rescued Girls: Social Stigma and Media Narratives of the Sex Industry in Victoria, B.C., from 1980 to 2005,CANADIAN REVIEW OF SOCIOLOGY/REVUE CANADIENNE DE SOCIOLOGIE, Issue 3 2006HELGA KRISTIN HALLGRIMSDOTTIR Cet article compare les portraits médiatiques de personnes qui travaillent dans l'industrie du sexe avec les représentations de soi de ces travailleurs, représentations qui incluent leurs origines personnelles et leurs expériences vécues au quotidien. Notre objectif est de jauger la distance empirique entre les descriptions médiatiques et la réalité vécue de ces travailleurs puis de comprendre comment les médias contribuent à cons-truire, reproduire et approfondir les stigmates sociaux associés au travail dans l'industrie du sexe. Nous avangons que le fait de distinguer la variabilité historique et spatiale de ces stigmates ainsi que le fait d'expli-quer leurs racines dans les activités de sens et de pratiques des auteurs et autorités médiatiques représentent une avancée cruciale pour la compréhension de leur construction sociale. This paper compares media portrayals of people who work in the sex industry with these workers' self-reports of their personal backgrounds and experiences of what they do for a living. Our aim is to first, gauge the empirical distance between media depictions and workers' lived reality, and second, to understand how the media contributes to constructing, reproducing and deepening the social stigmas associated with working in the sex industry. We argue that pulling apart the historical and spatial variability of these stigmas and explicating their roots in the meaning-making activities of media authors and authorities is a crucial step towards understanding their social construction. [source] | ||||||||||||||||||||||