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Sample records for reliable isoform quantification

  1. Network-Based Isoform Quantification with RNA-Seq Data for Cancer Transcriptome Analysis.

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    2015-12-01

    Full Text Available High-throughput mRNA sequencing (RNA-Seq is widely used for transcript quantification of gene isoforms. Since RNA-Seq data alone is often not sufficient to accurately identify the read origins from the isoforms for quantification, we propose to explore protein domain-domain interactions as prior knowledge for integrative analysis with RNA-Seq data. We introduce a Network-based method for RNA-Seq-based Transcript Quantification (Net-RSTQ to integrate protein domain-domain interaction network with short read alignments for transcript abundance estimation. Based on our observation that the abundances of the neighboring isoforms by domain-domain interactions in the network are positively correlated, Net-RSTQ models the expression of the neighboring transcripts as Dirichlet priors on the likelihood of the observed read alignments against the transcripts in one gene. The transcript abundances of all the genes are then jointly estimated with alternating optimization of multiple EM problems. In simulation Net-RSTQ effectively improved isoform transcript quantifications when isoform co-expressions correlate with their interactions. qRT-PCR results on 25 multi-isoform genes in a stem cell line, an ovarian cancer cell line, and a breast cancer cell line also showed that Net-RSTQ estimated more consistent isoform proportions with RNA-Seq data. In the experiments on the RNA-Seq data in The Cancer Genome Atlas (TCGA, the transcript abundances estimated by Net-RSTQ are more informative for patient sample classification of ovarian cancer, breast cancer and lung cancer. All experimental results collectively support that Net-RSTQ is a promising approach for isoform quantification. Net-RSTQ toolbox is available at http://compbio.cs.umn.edu/Net-RSTQ/.

  2. Exact reliability quantification of highly reliable systems with maintenance

    Energy Technology Data Exchange (ETDEWEB)

    Bris, Radim, E-mail: radim.bris@vsb.c [VSB-Technical University Ostrava, Faculty of Electrical Engineering and Computer Science, Department of Applied Mathematics, 17. listopadu 15, 70833 Ostrava-Poruba (Czech Republic)

    2010-12-15

    When a system is composed of highly reliable elements, exact reliability quantification may be problematic, because computer accuracy is limited. Inaccuracy can be due to different aspects. For example, an error may be made when subtracting two numbers that are very close to each other, or at the process of summation of many very different numbers, etc. The basic objective of this paper is to find a procedure, which eliminates errors made by PC when calculations close to an error limit are executed. Highly reliable system is represented by the use of directed acyclic graph which is composed from terminal nodes, i.e. highly reliable input elements, internal nodes representing subsystems and edges that bind all of these nodes. Three admissible unavailability models of terminal nodes are introduced, including both corrective and preventive maintenance. The algorithm for exact unavailability calculation of terminal nodes is based on merits of a high-performance language for technical computing MATLAB. System unavailability quantification procedure applied to a graph structure, which considers both independent and dependent (i.e. repeatedly occurring) terminal nodes is based on combinatorial principle. This principle requires summation of a lot of very different non-negative numbers, which may be a source of an inaccuracy. That is why another algorithm for exact summation of such numbers is designed in the paper. The summation procedure uses benefits from a special number system with the base represented by the value 2{sup 32}. Computational efficiency of the new computing methodology is compared with advanced simulation software. Various calculations on systems from references are performed to emphasize merits of the methodology.

  3. Review of some aspects of human reliability quantification

    International Nuclear Information System (INIS)

    Lydell, B.O.Y.; Spurgin, A.J.; Hannaman, G.W.; Lukic, Y.D.

    1986-01-01

    An area in systems reliability considered to be weak, is the characterization and quantification of the role of the operations and maintenance staff in combatting accidents. Several R and D programs are underway to improve the modeling of human interactions and some progress has been made. This paper describes a specific aspect of human reliability analysis which is referred to as modeling of cognitive processes. In particular, the basis for the so- called Human Cognitive Reliability (HCR) model is described and the focus is on its validation and on its benefits and limitations

  4. Reliability and discriminatory power of methods for dental plaque quantification

    Directory of Open Access Journals (Sweden)

    Daniela Prócida Raggio

    2010-04-01

    Full Text Available OBJECTIVE: This in situ study evaluated the discriminatory power and reliability of methods of dental plaque quantification and the relationship between visual indices (VI and fluorescence camera (FC to detect plaque. MATERIAL AND METHODS: Six volunteers used palatal appliances with six bovine enamel blocks presenting different stages of plaque accumulation. The presence of plaque with and without disclosing was assessed using VI. Images were obtained with FC and digital camera in both conditions. The area covered by plaque was assessed. Examinations were done by two independent examiners. Data were analyzed by Kruskal-Wallis and Kappa tests to compare different conditions of samples and to assess the inter-examiner reproducibility. RESULTS: Some methods presented adequate reproducibility. The Turesky index and the assessment of area covered by disclosed plaque in the FC images presented the highest discriminatory powers. CONCLUSION: The Turesky index and images with FC with disclosing present good reliability and discriminatory power in quantifying dental plaque.

  5. An architectural model for software reliability quantification: sources of data

    International Nuclear Information System (INIS)

    Smidts, C.; Sova, D.

    1999-01-01

    Software reliability assessment models in use today treat software as a monolithic block. An aversion towards 'atomic' models seems to exist. These models appear to add complexity to the modeling, to the data collection and seem intrinsically difficult to generalize. In 1997, we introduced an architecturally based software reliability model called FASRE. The model is based on an architecture derived from the requirements which captures both functional and nonfunctional requirements and on a generic classification of functions, attributes and failure modes. The model focuses on evaluation of failure mode probabilities and uses a Bayesian quantification framework. Failure mode probabilities of functions and attributes are propagated to the system level using fault trees. It can incorporate any type of prior information such as results of developers' testing, historical information on a specific functionality and its attributes, and, is ideally suited for reusable software. By building an architecture and deriving its potential failure modes, the model forces early appraisal and understanding of the weaknesses of the software, allows reliability analysis of the structure of the system, provides assessments at a functional level as well as at a systems' level. In order to quantify the probability of failure (or the probability of success) of a specific element of our architecture, data are needed. The term element of the architecture is used here in its broadest sense to mean a single failure mode or a higher level of abstraction such as a function. The paper surveys the potential sources of software reliability data available during software development. Next the mechanisms for incorporating these sources of relevant data to the FASRE model are identified

  6. Quantification of human reliability in probabilistic safety assessment

    International Nuclear Information System (INIS)

    Hirschberg, S.; Dankg, Vinh N.

    1996-01-01

    Human performance may substantially influence the reliability and safety of complex technical systems. For this reason, Human Reliability Analysis (HRA) constitutes an important part of Probabilistic Safety Assessment (PSAs) or Quantitative Risk Analyses (QRAs). The results of these studies as well as analyses of past accidents and incidents clearly demonstrate the importance of human interactions. The contribution of human errors to the core damage frequency (CDF), as estimated in the Swedish nuclear PSAs, are between 15 and 88%. A survey of the FRAs in the Swiss PSAs shows that also for the Swiss nuclear power plants the estimated HE contributions are substantial (49% of the CDF due to internal events in the case of Beznau and 70% in the case of Muehleberg; for the total CDF, including external events, 25% respectively 20%). Similar results can be extracted from the PSAs carried out for French, German, and US plants. In PSAs or QRAs, the adequate treatment of the human interactions with the system is a key to the understanding of accident sequences and their relative importance to overall risk. The main objectives of HRA are: first, to ensure that the key human interactions are systematically identified and incorporated into the safety analysis in a traceable manner, and second, to quantify the probabilities of their success and failure. Adopting a structured and systematic approach to the assessment of human performance makes it possible to provide greater confidence that the safety and availability of human-machine systems is not unduly jeopardized by human performance problems. Section 2 discusses the different types of human interactions analysed in PSAs. More generally, the section presents how HRA fits in the overall safety analysis, that is, how the human interactions to be quantified are identified. Section 3 addresses the methods for quantification. Section 4 concludes the paper by presenting some recommendations and pointing out the limitations of the

  7. Identification and quantification of metallothionein isoforms and superoxide dismutase in spiked liver extracts using HPLC-ESI-MS offline coupling and HPLC-ICP-MS online coupling.

    Science.gov (United States)

    Nischwitz, V; Michalke, B; Kettrup, A

    2003-01-01

    A two-dimensional chromatographic method for the characterization of metallothionein isoforms (MT) and superoxide dismutase (SOD) in spiked liver extracts was developed for the optimization of extraction procedures from liver samples. Element-specific detection (ICP-MS) and molecule-specific detection (ESI-MS) were applied for maximum species information. A special focus was laid on the quantitative data evaluation (species stoichiometry, calibration with and without matrix, recovery), which is neglected in most MT/SOD publications with hyphenated techniques. Linearity, precision (residual standard deviation of calibration curves <10%), and detection limits (<0.6 mg L(-1) for MT isoforms and 13 mg L(-1) for SOD) prove the suitability of the method for quantification. An alternative quantification is proposed for the extension towards other lesser or even unknown trace element species, especially the native porcine MT and SOD.

  8. Probabilistic risk assessment course documentation. Volume 5. System reliability and analysis techniques Session D - quantification

    International Nuclear Information System (INIS)

    Lofgren, E.V.

    1985-08-01

    This course in System Reliability and Analysis Techniques focuses on the probabilistic quantification of accident sequences and the link between accident sequences and consequences. Other sessions in this series focus on the quantification of system reliability and the development of event trees and fault trees. This course takes the viewpoint that event tree sequences or combinations of system failures and success are available and that Boolean equations for system fault trees have been developed and are available. 93 figs., 11 tabs

  9. Practical reliability and uncertainty quantification in complex systems : final report.

    Energy Technology Data Exchange (ETDEWEB)

    Grace, Matthew D.; Ringland, James T.; Marzouk, Youssef M. (Massachusetts Institute of Technology, Cambridge, MA); Boggs, Paul T.; Zurn, Rena M.; Diegert, Kathleen V. (Sandia National Laboratories, Albuquerque, NM); Pebay, Philippe Pierre; Red-Horse, John Robert (Sandia National Laboratories, Albuquerque, NM)

    2009-09-01

    The purpose of this project was to investigate the use of Bayesian methods for the estimation of the reliability of complex systems. The goals were to find methods for dealing with continuous data, rather than simple pass/fail data; to avoid assumptions of specific probability distributions, especially Gaussian, or normal, distributions; to compute not only an estimate of the reliability of the system, but also a measure of the confidence in that estimate; to develop procedures to address time-dependent or aging aspects in such systems, and to use these models and results to derive optimal testing strategies. The system is assumed to be a system of systems, i.e., a system with discrete components that are themselves systems. Furthermore, the system is 'engineered' in the sense that each node is designed to do something and that we have a mathematical description of that process. In the time-dependent case, the assumption is that we have a general, nonlinear, time-dependent function describing the process. The major results of the project are described in this report. In summary, we developed a sophisticated mathematical framework based on modern probability theory and Bayesian analysis. This framework encompasses all aspects of epistemic uncertainty and easily incorporates steady-state and time-dependent systems. Based on Markov chain, Monte Carlo methods, we devised a computational strategy for general probability density estimation in the steady-state case. This enabled us to compute a distribution of the reliability from which many questions, including confidence, could be addressed. We then extended this to the time domain and implemented procedures to estimate the reliability over time, including the use of the method to predict the reliability at a future time. Finally, we used certain aspects of Bayesian decision analysis to create a novel method for determining an optimal testing strategy, e.g., we can estimate the 'best' location to

  10. Quantification of total apolipoprotein E and its specific isoforms in cerebrospinal fluid and blood in Alzheimer’s disease and other neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    Melinda Rezeli

    2015-09-01

    Full Text Available A targeted mass spectrometric assay was developed for identification and quantification of apoE isoforms (apoE2, E3 and E4, and it was utilized for screening of samples from AD patients (n = 39 and patients with other neurodegenerative disorders (n = 38. The assay showed good linearity with LOQ corresponds to total apoE concentration of 0.8 and 40 ng/mL in CSF and plasma/serum, respectively. We identified apoE phenotypes with 100% accuracy in clinical samples. We found strong association between genotypes of the individuals and their apoE levels in blood; ϵ4 allele carriers had significantly lower apoE levels in blood than non-carriers.

  11. Multiplicity of expression of Na+,K+-ATPase alpha-subunit isoforms in the gill of Atlantic salmon: quantification and cellular localisation in response to salinity

    DEFF Research Database (Denmark)

    Madsen, Steffen; Kiilerich, Pia; Tipsmark, Christian Kølbæk

    2009-01-01

    but occasionally also on lamellae. Overall, the salinity-induced variation in labelling pattern and intensity matched the quantification data. In conclusion, the predominant switching of Na+,K+-ATPase -subunit isoform mRNA during salinity acclimation reflects a marked remodelling of mitochondrion-rich cells (MRCs...

  12. Evaluation of the reliability of maize reference assays for GMO quantification.

    Science.gov (United States)

    Papazova, Nina; Zhang, David; Gruden, Kristina; Vojvoda, Jana; Yang, Litao; Buh Gasparic, Meti; Blejec, Andrej; Fouilloux, Stephane; De Loose, Marc; Taverniers, Isabel

    2010-03-01

    A reliable PCR reference assay for relative genetically modified organism (GMO) quantification must be specific for the target taxon and amplify uniformly along the commercialised varieties within the considered taxon. Different reference assays for maize (Zea mays L.) are used in official methods for GMO quantification. In this study, we evaluated the reliability of eight existing maize reference assays, four of which are used in combination with an event-specific polymerase chain reaction (PCR) assay validated and published by the Community Reference Laboratory (CRL). We analysed the nucleotide sequence variation in the target genomic regions in a broad range of transgenic and conventional varieties and lines: MON 810 varieties cultivated in Spain and conventional varieties from various geographical origins and breeding history. In addition, the reliability of the assays was evaluated based on their PCR amplification performance. A single base pair substitution, corresponding to a single nucleotide polymorphism (SNP) reported in an earlier study, was observed in the forward primer of one of the studied alcohol dehydrogenase 1 (Adh1) (70) assays in a large number of varieties. The SNP presence is consistent with a poor PCR performance observed for this assay along the tested varieties. The obtained data show that the Adh1 (70) assay used in the official CRL NK603 assay is unreliable. Based on our results from both the nucleotide stability study and the PCR performance test, we can conclude that the Adh1 (136) reference assay (T25 and Bt11 assays) as well as the tested high mobility group protein gene assay, which also form parts of CRL methods for quantification, are highly reliable. Despite the observed uniformity in the nucleotide sequence of the invertase gene assay, the PCR performance test reveals that this target sequence might occur in more than one copy. Finally, although currently not forming a part of official quantification methods, zein and SSIIb

  13. mRNA Quantification of NIPBL Isoforms A and B in Adult and Fetal Human Tissues, and a Potentially Pathological Variant Affecting Only Isoform A in Two Patients with Cornelia de Lange Syndrome

    Directory of Open Access Journals (Sweden)

    Beatriz Puisac

    2017-02-01

    Full Text Available Cornelia de Lange syndrome (CdLS is a congenital developmental disorder characterized by craniofacial dysmorphia, growth retardation, limb malformations, and intellectual disability. Approximately 60% of patients with CdLS carry a recognizable pathological variant in the NIPBL gene, of which two isoforms, A and B, have been identified, and which only differ in the C-terminal segment. In this work, we describe the distribution pattern of the isoforms A and B mRNAs in tissues of adult and fetal origin, by qPCR (quantitative polymerase chain reaction. Our results show a higher gene expression of the isoform A, even though both seem to have the same tissue distribution. Interestingly, the expression in fetal tissues is higher than that of adults, especially in brain and skeletal muscle. Curiously, the study of fibroblasts of two siblings with a mild CdLS phenotype and a pathological variant specific of the isoform A of NIPBL (c.8387A > G; P.Tyr2796Cys, showed a similar reduction in both isoforms, and a normal sensitivity to DNA damage. Overall, these results suggest that the position of the pathological variant at the 3´ end of the NIPBL gene affecting only isoform A, is likely to be the cause of the atypical mild phenotype of the two brothers.

  14. Reliable quantification of phthalates in environmental matrices (air, water, sludge, sediment and soil): a review.

    Science.gov (United States)

    Net, Sopheak; Delmont, Anne; Sempéré, Richard; Paluselli, Andrea; Ouddane, Baghdad

    2015-05-15

    Because of their widespread application, phthalates or phthalic acid esters (PAEs) are ubiquitous in the environment. Their presence has attracted considerable attention due to their potential impacts on ecosystem functioning and on public health, so their quantification has become a necessity. Various extraction procedures as well as gas/liquid chromatography and mass spectrometry detection techniques are found as suitable for reliable detection of such compounds. However, PAEs are ubiquitous in the laboratory environment including ambient air, reagents, sampling equipment, and various analytical devices, that induces difficult analysis of real samples with a low PAE background. Therefore, accurate PAE analysis in environmental matrices is a challenging task. This paper reviews the extensive literature data on the techniques for PAE quantification in natural media. Sampling, sample extraction/pretreatment and detection for quantifying PAEs in different environmental matrices (air, water, sludge, sediment and soil) have been reviewed and compared. The concept of "green analytical chemistry" for PAE determination is also discussed. Moreover useful information about the material preparation and the procedures of quality control and quality assurance are presented to overcome the problem of sample contamination and these encountered due to matrix effects in order to avoid overestimating PAE concentrations in the environment. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Review of advances in human reliability analysis of errors of commission-Part 2: EOC quantification

    International Nuclear Information System (INIS)

    Reer, Bernhard

    2008-01-01

    In close connection with examples relevant to contemporary probabilistic safety assessment (PSA), a review of advances in human reliability analysis (HRA) of post-initiator errors of commission (EOCs), i.e. inappropriate actions under abnormal operating conditions, has been carried out. The review comprises both EOC identification (part 1) and quantification (part 2); part 2 is presented in this article. Emerging HRA methods in this field are: ATHEANA, MERMOS, the EOC HRA method developed by Gesellschaft fuer Anlagen- und Reaktorsicherheit (GRS), the MDTA method and CREAM. The essential advanced features are on the conceptual side, especially to envisage the modeling of multiple contexts for an EOC to be quantified (ATHEANA, MERMOS and MDTA), in order to explicitly address adverse conditions. There is promising progress in providing systematic guidance to better account for cognitive demands and tendencies (GRS, CREAM), and EOC recovery (MDTA). Problematic issues are associated with the implementation of multiple context modeling and the assessment of context-specific error probabilities. Approaches for task or error opportunity scaling (CREAM, GRS) and the concept of reference cases (ATHEANA outlook) provide promising orientations for achieving progress towards data-based quantification. Further development work is needed and should be carried out in close connection with large-scale applications of existing approaches

  16. Uncertainty Quantification in the Reliability and Risk Assessment of Generation IV Reactors: Final Scientific/Technical Report

    International Nuclear Information System (INIS)

    Vierow, Karen; Aldemir, Tunc

    2009-01-01

    The project entitled, 'Uncertainty Quantification in the Reliability and Risk Assessment of Generation IV Reactors', was conducted as a DOE NERI project collaboration between Texas A and M University and The Ohio State University between March 2006 and June 2009. The overall goal of the proposed project was to develop practical approaches and tools by which dynamic reliability and risk assessment techniques can be used to augment the uncertainty quantification process in probabilistic risk assessment (PRA) methods and PRA applications for Generation IV reactors. This report is the Final Scientific/Technical Report summarizing the project.

  17. Uncertainty Quantification in the Reliability and Risk Assessment of Generation IV Reactors: Final Scientific/Technical Report

    Energy Technology Data Exchange (ETDEWEB)

    Vierow, Karen; Aldemir, Tunc

    2009-09-10

    The project entitled, “Uncertainty Quantification in the Reliability and Risk Assessment of Generation IV Reactors”, was conducted as a DOE NERI project collaboration between Texas A&M University and The Ohio State University between March 2006 and June 2009. The overall goal of the proposed project was to develop practical approaches and tools by which dynamic reliability and risk assessment techniques can be used to augment the uncertainty quantification process in probabilistic risk assessment (PRA) methods and PRA applications for Generation IV reactors. This report is the Final Scientific/Technical Report summarizing the project.

  18. Reliability Quantification Method for Safety Critical Software Based on a Finite Test Set

    International Nuclear Information System (INIS)

    Shin, Sung Min; Kim, Hee Eun; Kang, Hyun Gook; Lee, Seung Jun

    2014-01-01

    Software inside of digitalized system have very important role because it may cause irreversible consequence and affect the whole system as common cause failure. However, test-based reliability quantification method for some safety critical software has limitations caused by difficulties in developing input sets as a form of trajectory which is series of successive values of variables. To address these limitations, this study proposed another method which conduct the test using combination of single values of variables. To substitute the trajectory form of input using combination of variables, the possible range of each variable should be identified. For this purpose, assigned range of each variable, logical relations between variables, plant dynamics under certain situation, and characteristics of obtaining information of digital device are considered. A feasibility of the proposed method was confirmed through an application to the Reactor Protection System (RPS) software trip logic

  19. Assessment of ALWR passive safety system reliability. Phase 1: Methodology development and component failure quantification

    International Nuclear Information System (INIS)

    Hake, T.M.; Heger, A.S.

    1995-04-01

    Many advanced light water reactor (ALWR) concepts proposed for the next generation of nuclear power plants rely on passive systems to perform safety functions, rather than active systems as in current reactor designs. These passive systems depend to a great extent on physical processes such as natural circulation for their driving force, and not on active components, such as pumps. An NRC-sponsored study was begun at Sandia National Laboratories to develop and implement a methodology for evaluating ALWR passive system reliability in the context of probabilistic risk assessment (PRA). This report documents the first of three phases of this study, including methodology development, system-level qualitative analysis, and sequence-level component failure quantification. The methodology developed addresses both the component (e.g. valve) failure aspect of passive system failure, and uncertainties in system success criteria arising from uncertainties in the system's underlying physical processes. Traditional PRA methods, such as fault and event tree modeling, are applied to the component failure aspect. Thermal-hydraulic calculations are incorporated into a formal expert judgment process to address uncertainties in selected natural processes and success criteria. The first phase of the program has emphasized the component failure element of passive system reliability, rather than the natural process uncertainties. Although cursory evaluation of the natural processes has been performed as part of Phase 1, detailed assessment of these processes will take place during Phases 2 and 3 of the program

  20. Modeling and Quantification of Team Performance in Human Reliability Analysis for Probabilistic Risk Assessment

    Energy Technology Data Exchange (ETDEWEB)

    Jeffrey C. JOe; Ronald L. Boring

    2014-06-01

    Probabilistic Risk Assessment (PRA) and Human Reliability Assessment (HRA) are important technical contributors to the United States (U.S.) Nuclear Regulatory Commission’s (NRC) risk-informed and performance based approach to regulating U.S. commercial nuclear activities. Furthermore, all currently operating commercial NPPs in the U.S. are required by federal regulation to be staffed with crews of operators. Yet, aspects of team performance are underspecified in most HRA methods that are widely used in the nuclear industry. There are a variety of "emergent" team cognition and teamwork errors (e.g., communication errors) that are 1) distinct from individual human errors, and 2) important to understand from a PRA perspective. The lack of robust models or quantification of team performance is an issue that affects the accuracy and validity of HRA methods and models, leading to significant uncertainty in estimating HEPs. This paper describes research that has the objective to model and quantify team dynamics and teamwork within NPP control room crews for risk informed applications, thereby improving the technical basis of HRA, which improves the risk-informed approach the NRC uses to regulate the U.S. commercial nuclear industry.

  1. High level issues in reliability quantification of safety-critical software

    International Nuclear Information System (INIS)

    Kim, Man Cheol

    2012-01-01

    For the purpose of developing a consensus method for the reliability assessment of safety-critical digital instrumentation and control systems in nuclear power plants, several high level issues in reliability assessment of the safety-critical software based on Bayesian belief network modeling and statistical testing are discussed. Related to the Bayesian belief network modeling, the relation between the assessment approach and the sources of evidence, the relation between qualitative evidence and quantitative evidence, how to consider qualitative evidence, and the cause-consequence relation are discussed. Related to the statistical testing, the need of the consideration of context-specific software failure probabilities and the inability to perform a huge number of tests in the real world are discussed. The discussions in this paper are expected to provide a common basis for future discussions on the reliability assessment of safety-critical software. (author)

  2. Quantification of Wave Model Uncertainties Used for Probabilistic Reliability Assessments of Wave Energy Converters

    DEFF Research Database (Denmark)

    Ambühl, Simon; Kofoed, Jens Peter; Sørensen, John Dalsgaard

    2015-01-01

    Wave models used for site assessments are subjected to model uncertainties, which need to be quantified when using wave model results for probabilistic reliability assessments. This paper focuses on determination of wave model uncertainties. Four different wave models are considered, and validation...... data are collected from published scientific research. The bias and the root-mean-square error, as well as the scatter index, are considered for the significant wave height as well as the mean zero-crossing wave period. Based on an illustrative generic example, this paper presents how the quantified...... uncertainties can be implemented in probabilistic reliability assessments....

  3. Identification of Reliable Reference Genes for Quantification of MicroRNAs in Serum Samples of Sulfur Mustard-Exposed Veterans.

    Science.gov (United States)

    Gharbi, Sedigheh; Shamsara, Mehdi; Khateri, Shahriar; Soroush, Mohammad Reza; Ghorbanmehr, Nassim; Tavallaei, Mahmood; Nourani, Mohammad Reza; Mowla, Seyed Javad

    2015-01-01

    In spite of accumulating information about pathological aspects of sulfur mustard (SM), the precise mechanism responsible for its effects is not well understood. Circulating microRNAs (miRNAs) are promising biomarkers for disease diagnosis and prognosis. Accurate normalization using appropriate reference genes, is a critical step in miRNA expression studies. In this study, we aimed to identify appropriate reference gene for microRNA quantification in serum samples of SM victims. In this case and control experimental study, using quantitative real-time polymerase chain reaction (qRT-PCR), we evaluated the suitability of a panel of small RNAs including SNORD38B, SNORD49A, U6, 5S rRNA, miR-423-3p, miR-191, miR-16 and miR-103 in sera of 28 SM-exposed veterans of Iran-Iraq war (1980-1988) and 15 matched control volunteers. Different statistical algorithms including geNorm, Normfinder, best-keeper and comparative delta-quantification cycle (Cq) method were employed to find the least variable reference gene. miR-423-3p was identified as the most stably expressed reference gene, and miR- 103 and miR-16 ranked after that. We demonstrate that non-miRNA reference genes have the least stabil- ity in serum samples and that some house-keeping miRNAs may be used as more reliable reference genes for miRNAs in serum. In addition, using the geometric mean of two reference genes could increase the reliability of the normalizers.

  4. Reliability of smartphone-based gait measurements for quantification of physical activity/inactivity levels.

    Science.gov (United States)

    Ebara, Takeshi; Azuma, Ryohei; Shoji, Naoto; Matsukawa, Tsuyoshi; Yamada, Yasuyuki; Akiyama, Tomohiro; Kurihara, Takahiro; Yamada, Shota

    2017-11-25

    Objective measurements using built-in smartphone sensors that can measure physical activity/inactivity in daily working life have the potential to provide a new approach to assessing workers' health effects. The aim of this study was to elucidate the characteristics and reliability of built-in step counting sensors on smartphones for development of an easy-to-use objective measurement tool that can be applied in ergonomics or epidemiological research. To evaluate the reliability of step counting sensors embedded in seven major smartphone models, the 6-minute walk test was conducted and the following analyses of sensor precision and accuracy were performed: 1) relationship between actual step count and step count detected by sensors, 2) reliability between smartphones of the same model, and 3) false detection rates when sitting during office work, while riding the subway, and driving. On five of the seven models, the inter-class correlations coefficient (ICC (3,1) ) showed high reliability with a range of 0.956-0.993. The other two models, however, had ranges of 0.443-0.504 and the relative error ratios of the sensor-detected step count to the actual step count were ±48.7%-49.4%. The level of agreement between the same models was ICC (3,1) : 0.992-0.998. The false detection rates differed between the sitting conditions. These results suggest the need for appropriate regulation of step counts measured by sensors, through means such as correction or calibration with a predictive model formula, in order to obtain the highly reliable measurement results that are sought in scientific investigation.

  5. Development of a reliable extraction and quantification method for glucosinolates in Moringa oleifera.

    Science.gov (United States)

    Förster, Nadja; Ulrichs, Christian; Schreiner, Monika; Müller, Carsten T; Mewis, Inga

    2015-01-01

    Glucosinolates are the characteristic secondary metabolites of plants in the order Brassicales. To date the common DIN extraction 'desulfo glucosinolates' method remains the common procedure for determination and quantification of glucosinolates. However, the desulfation step in the extraction of glucosinolates from Moringa oleifera leaves resulted in complete conversion and degradation of the naturally occurring glucosinolates in this plant. Therefore, a method for extraction of intact Moringa glucosinolates was developed and no conversion and degradation of the different rhamnopyranosyloxy-benzyl glucosinolates was found. Buffered eluents (0.1 M ammonium acetate) were necessary to stabilize 4-α-rhamnopyranosyloxy-benzyl glucosinolate (Rhamno-Benzyl-GS) and acetyl-4-α-rhamnopyranosyloxy-benzyl glucosinolate isomers (Ac-Isomers-GS) during HPLC analysis. Due to the instability of intact Moringa glucosinolates at room temperature and during the purification process of single glucosinolates, influences of different storage (room temperature, frozen, thawing and refreezing) and buffer conditions on glucosinolate conversion were analysed. Conversion and degradations processes were especially determined for the Ac-Isomers-GS III. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Reliable Assessment and Quantification of the Fluorescence-Labeled Antisense Oligonucleotides In Vivo

    Directory of Open Access Journals (Sweden)

    Maria Chiara Munisso

    2014-01-01

    Full Text Available The availability of fluorescent dyes and the advances in the optical systems for in vivo imaging have stimulated an increasing interest in developing new methodologies to study and quantify the biodistribution of labeled agents. However, despite these great achievements, we are facing significant challenges in determining if the observed fluorescence does correspond to the quantity of the dye in the tissues. In fact, although the far-red and near-infrared lights can propagate through several centimetres of tissue, they diffuse within a few millimetres as consequence of the elastic scattering of photons. In addition, when dye-labeled oligonucleotides form stable complex with cationic carriers, a large change in the fluorescence intensity of the dye is observed. Therefore, the measured fluorescence intensity is altered by the tissue heterogeneity and by the fluctuation of dye intensity. Hence, in this study a quantification strategy for fluorescence-labeled oligonucleotides was developed to solve these disadvantageous effects. Our results proved that upon efficient homogenization and dilution with chaotropic agents, such as guanidinium thiocyanate, it is possible to achieve a complete fluorescence intensity recovery. Furthermore, we demonstrated that this method has the advantage of good sensitivity and reproducibility, as well as easy handling of the tissue samples.

  7. Quantification of colour Doppler activity in the wrist in patients with rheumatoid arthritis - the reliability of different methods for image selection and evaluation

    DEFF Research Database (Denmark)

    Ellegaard, K.; Torp-Pedersen, S.; Lund, H.

    2008-01-01

    measurements in the wrist of patients with rheumatoid arthritis (RA) using different selection and quantification methods. Materials and Methods: 14 patients with RA had their wrist scanned twice by the same investigator with an interval of 30 Minutes, The images for analysis were selected either......Purpose: The amount Of colour Doppler activity in the inflamed synovium is used to quantity inflammatory activity. The measurements may vary due to image selection, quantification method, and point in cardiac cycle. This study investigated the test-retest reliability Of ultrasound colour Doppler...... was obtained when the images were selected guided by colour Doppler and the Subsequent quantification was (done in an area defined by anatomical Structures. With this method, the intra-class coefficient ICC (2.1) was 0.95 and the within-subject SD (SW) was 0.017, indicating good reliability. In contrast, poor...

  8. Latest scientific and technological knowledge of human-reliability quantification - December 1991

    International Nuclear Information System (INIS)

    Berg, H.P.; Schott, H.

    1992-02-01

    Again an again real incidents and accidents show that human factors may seriously affect the safety of plants. This is also true for, e.g. nuclear facilities. The major methods which are used to quantify the reliability of humans are described. These methods are applied in the framework of German and international risk analyses. Since in probabilistic safety analyses data bases are of great importance of the, however, naturally very difficult quantitative evaluation of human errors, the study also discusses the present limits to the treatment of human misbehavior in safety analyses. (orig.) [de

  9. Study on Performance Shaping Factors (PSFs) Quantification Method in Human Reliability Analysis (HRA)

    International Nuclear Information System (INIS)

    Kim, Ar Ryum; Jang, Inseok Jang; Seong, Poong Hyun; Park, Jinkyun; Kim, Jong Hyun

    2015-01-01

    The purpose of HRA implementation is 1) to achieve the human factor engineering (HFE) design goal of providing operator interfaces that will minimize personnel errors and 2) to conduct an integrated activity to support probabilistic risk assessment (PRA). For these purposes, various HRA methods have been developed such as technique for human error rate prediction (THERP), simplified plant analysis risk human reliability assessment (SPAR-H), cognitive reliability and error analysis method (CREAM) and so on. In performing HRA, such conditions that influence human performances have been represented via several context factors called performance shaping factors (PSFs). PSFs are aspects of the human's individual characteristics, environment, organization, or task that specifically decrements or improves human performance, thus respectively increasing or decreasing the likelihood of human errors. Most HRA methods evaluate the weightings of PSFs by expert judgment and explicit guidance for evaluating the weighting is not provided. It has been widely known that the performance of the human operator is one of the critical factors to determine the safe operation of NPPs. HRA methods have been developed to identify the possibility and mechanism of human errors. In performing HRA methods, the effect of PSFs which may increase or decrease human error should be investigated. However, the effect of PSFs were estimated by expert judgment so far. Accordingly, in order to estimate the effect of PSFs objectively, the quantitative framework to estimate PSFs by using PSF profiles is introduced in this paper

  10. A reliable and validated LC-MS/MS method for the simultaneous quantification of 4 cannabinoids in 40 consumer products.

    Directory of Open Access Journals (Sweden)

    Qingfang Meng

    Full Text Available In the past 50 years, Cannabis sativa (C. sativa has gone from a substance essentially prohibited worldwide to one that is gaining acceptance both culturally and legally in many countries for medicinal and recreational use. As additional jurisdictions legalize Cannabis products and the variety and complexity of these products surpass the classical dried plant material, appropriate methods for measuring the biologically active constituents is paramount to ensure safety and regulatory compliance. While there are numerous active compounds in C. sativa the primary cannabinoids of regulatory and safety concern are (--Δ⁹-tetrahydrocannabinol (THC, cannabidiol (CBD, and their respective acidic forms THCA-A and CBDA. Using the US Food and Drug Administration (FDA bioanalytical method validation guidelines we developed a sensitive, selective, and accurate method for the simultaneous analysis CBD, CBDA, THC, and THCA-A in oils and THC & CBD in more complex matrices. This HPLC-MS/MS method was simple and reliable using standard sample dilution and homogenization, an isocratic chromatographic separation, and a triple quadrupole mass spectrometer. The lower limit of quantification (LLOQ for analytes was 0.195 ng/mL over a 0.195-50.0 ng/mL range of quantification with a coefficient of correlation of >0.99. Average intra-day and inter-day accuracies were 94.2-112.7% and 97.2-110.9%, respectively. This method was used to quantify CBD, CBDA, THC, and THCA-A in 40 commercial hemp products representing a variety of matrices including oils, plant materials, and creams/cosmetics. All products tested met the federal regulatory restrictions on THC content in Canada (1,000 (an oil-based product. Overall, the method proved amenable to the analysis of various commercial products including oils, creams, and plant material and may be diagnostically indicative of adulteration with non-hemp C. sativa, specialized hemp cultivars, or unique manufacturing methods.

  11. Quantification of the occurrence of common-mode faults in highly reliable protective systems

    International Nuclear Information System (INIS)

    Aitken, A.

    1978-10-01

    The report first covers the investigation, definition and classification of common mode failure (CMF) based on an extensive study of the nature of CMF. A new classification of CMF is proposed, based on possible causes of failures. This is used as a basis for analysing data from reported failures of reactor safety systems and aircraft systems. Design and maintenance errors are shown to be predominant cause of CMF. The estimated CMF rates for the highly reliable nuclear power plant automatic protection system (APS) and for the emergency core cooling system (ECCS) are 2.8.10 -2 CMF/sub-system-year and 3.3.10 -2 CMF/sub-system-year respectively. For comparison, the data from the aircraft accident records have shown a CMF rate for total flight control system (FCS), 2.1.10 -5 CMF/sub-system-year. The analysis has laid the grounds for work on relating CMF modelling and defences

  12. Reliability

    OpenAIRE

    Condon, David; Revelle, William

    2017-01-01

    Separating the signal in a test from the irrelevant noise is a challenge for all measurement. Low test reliability limits test validity, attenuates important relationships, and can lead to regression artifacts. Multiple approaches to the assessment and improvement of reliability are discussed. The advantages and disadvantages of several different approaches to reliability are considered. Practical advice on how to assess reliability using open source software is provided.

  13. Improving the reliability of POD curves in NDI methods using a Bayesian inversion approach for uncertainty quantification

    Science.gov (United States)

    Ben Abdessalem, A.; Jenson, F.; Calmon, P.

    2016-02-01

    This contribution provides an example of the possible advantages of adopting a Bayesian inversion approach to uncertainty quantification in nondestructive inspection methods. In such problem, the uncertainty associated to the random parameters is not always known and needs to be characterised from scattering signal measurements. The uncertainties may then correctly propagated in order to determine a reliable probability of detection curve. To this end, we establish a general Bayesian framework based on a non-parametric maximum likelihood function formulation and some priors from expert knowledge. However, the presented inverse problem is time-consuming and computationally intensive. To cope with this difficulty, we replace the real model by a surrogate one in order to speed-up the model evaluation and to make the problem to be computationally feasible for implementation. The least squares support vector regression is adopted as metamodelling technique due to its robustness to deal with non-linear problems. We illustrate the usefulness of this methodology through the control of tube with enclosed defect using ultrasonic inspection method.

  14. A reliable and validated LC-MS/MS method for the simultaneous quantification of 4 cannabinoids in 40 consumer products.

    Science.gov (United States)

    Meng, Qingfang; Buchanan, Beth; Zuccolo, Jonathan; Poulin, Mathieu-Marc; Gabriele, Joseph; Baranowski, David Charles

    2018-01-01

    In the past 50 years, Cannabis sativa (C. sativa) has gone from a substance essentially prohibited worldwide to one that is gaining acceptance both culturally and legally in many countries for medicinal and recreational use. As additional jurisdictions legalize Cannabis products and the variety and complexity of these products surpass the classical dried plant material, appropriate methods for measuring the biologically active constituents is paramount to ensure safety and regulatory compliance. While there are numerous active compounds in C. sativa the primary cannabinoids of regulatory and safety concern are (-)-Δ⁹-tetrahydrocannabinol (THC), cannabidiol (CBD), and their respective acidic forms THCA-A and CBDA. Using the US Food and Drug Administration (FDA) bioanalytical method validation guidelines we developed a sensitive, selective, and accurate method for the simultaneous analysis CBD, CBDA, THC, and THCA-A in oils and THC & CBD in more complex matrices. This HPLC-MS/MS method was simple and reliable using standard sample dilution and homogenization, an isocratic chromatographic separation, and a triple quadrupole mass spectrometer. The lower limit of quantification (LLOQ) for analytes was 0.195 ng/mL over a 0.195-50.0 ng/mL range of quantification with a coefficient of correlation of >0.99. Average intra-day and inter-day accuracies were 94.2-112.7% and 97.2-110.9%, respectively. This method was used to quantify CBD, CBDA, THC, and THCA-A in 40 commercial hemp products representing a variety of matrices including oils, plant materials, and creams/cosmetics. All products tested met the federal regulatory restrictions on THC content in Canada (CBD, the majority of analyzed products contained low CBD levels and a CBD: CBDA ratio of CBD and a CBD: CBDA ratio of >1,000 (an oil-based product). Overall, the method proved amenable to the analysis of various commercial products including oils, creams, and plant material and may be diagnostically indicative of

  15. Zeta-potential data reliability of gold nanoparticle biomolecular conjugates and its application in sensitive quantification of surface absorbed protein.

    Science.gov (United States)

    Wang, Wenjie; Ding, Xiaofan; Xu, Qing; Wang, Jing; Wang, Lei; Lou, Xinhui

    2016-12-01

    Zeta potentials (ZP) of gold nanoparticle bioconjugates (AuNP-bios) provide important information on surface charge that is critical for many applications including drug delivery, biosensing, and cell imaging. The ZP measurements (ZPMs) are conducted under an alternative electrical field at a high frequency under laser irradiation, which may strongly affect the status of surface coating of AuNP-bios and generate unreliable data. In this study, we systemically evaluated the ZP data reliability (ZPDR) of citrate-, thiolated single stranded DNA-, and protein-coated AuNPs mainly according to the consistence of ZPs in the repeated ZPMs and the changes of the hydrodynamic size before and after the ZPMs. We found that the ZPDR was highly dependent on both buffer conditions and surface modifications. Overall, the higher ionic strength of the buffer and the lower affinity of surface bounders were related with the worse ZPDR. The ZPDR of citrate-coated AuNP was good in water, but bad in 10mM phosphate buffer (PB), showing substantially decrease of the absolute ZP values after each measurement, probably due to the electrical field facilitated adsorption of negatively charged phosphate ions on AuNPs. The significant desorption of DNAs from AuNP was observed in the PB containing medium concentration of NaCl, but not in PB. The excellent ZPDR of bovine serum albumin (BSA)-coated AuNP was observed at high salt concentrations and low surface coverage, enabling ZPM as an ultra-sensitive tool for protein quantification on the surface of AuNPs with a single molecule resolution. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Overall Key Performance Indicator to Optimizing Operation of High-Pressure Homogenizers for a Reliable Quantification of Intracellular Components in Pichia pastoris.

    Science.gov (United States)

    Garcia-Ortega, Xavier; Reyes, Cecilia; Montesinos, José Luis; Valero, Francisco

    2015-01-01

    The most commonly used cell disruption procedures may present lack of reproducibility, which introduces significant errors in the quantification of intracellular components. In this work, an approach consisting in the definition of an overall key performance indicator (KPI) was implemented for a lab scale high-pressure homogenizer (HPH) in order to determine the disruption settings that allow the reliable quantification of a wide sort of intracellular components. This innovative KPI was based on the combination of three independent reporting indicators: decrease of absorbance, release of total protein, and release of alkaline phosphatase activity. The yeast Pichia pastoris growing on methanol was selected as model microorganism due to it presents an important widening of the cell wall needing more severe methods and operating conditions than Escherichia coli and Saccharomyces cerevisiae. From the outcome of the reporting indicators, the cell disruption efficiency achieved using HPH was about fourfold higher than other lab standard cell disruption methodologies, such bead milling cell permeabilization. This approach was also applied to a pilot plant scale HPH validating the methodology in a scale-up of the disruption process. This innovative non-complex approach developed to evaluate the efficacy of a disruption procedure or equipment can be easily applied to optimize the most common disruption processes, in order to reach not only reliable quantification but also recovery of intracellular components from cell factories of interest.

  17. Accurate and reliable quantification of total microalgal fuel potential as fatty acid methyl esters by in situ transesterification

    Energy Technology Data Exchange (ETDEWEB)

    Laurens, Lieve M.L.; Quinn, Matthew; Wychen, Stefanie van; Templeton, David W.; Wolfrum, Edward J. [National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO (United States)

    2012-04-15

    In the context of algal biofuels, lipids, or better aliphatic chains of the fatty acids, are perhaps the most important constituents of algal biomass. Accurate quantification of lipids and their respective fuel yield is crucial for comparison of algal strains and growth conditions and for process monitoring. As an alternative to traditional solvent-based lipid extraction procedures, we have developed a robust whole-biomass in situ transesterification procedure for quantification of algal lipids (as fatty acid methyl esters, FAMEs) that (a) can be carried out on a small scale (using 4-7 mg of biomass), (b) is applicable to a range of different species, (c) consists of a single-step reaction, (d) is robust over a range of different temperature and time combinations, and (e) tolerant to at least 50% water in the biomass. Unlike gravimetric lipid quantification, which can over- or underestimate the lipid content, whole biomass transesterification reflects the true potential fuel yield of algal biomass. We report here on the comparison of the yield of FAMEs by using different catalysts and catalyst combinations, with the acid catalyst HCl providing a consistently high level of conversion of fatty acids with a precision of 1.9% relative standard deviation. We investigate the influence of reaction time, temperature, and biomass water content on the measured FAME content and profile for 4 different samples of algae (replete and deplete Chlorella vulgaris, replete Phaeodactylum tricornutum, and replete Nannochloropsis sp.). We conclude by demonstrating a full mass balance closure of all fatty acids around a traditional lipid extraction process. (orig.)

  18. Reliability of video-based quantification of the knee- and hip angle at foot strike during running

    DEFF Research Database (Denmark)

    Damsted, Camma; Nielsen, R.O.; Larsen, Lars Henrik

    2015-01-01

    INTRODUCTION: In clinical practice, joint kinematics during running are primarily quantified by two-dimensional (2D) video recordings and motion-analysis software. The applicability of this approach depends on the clinicians' ability to quantify kinematics in a reliable manner. The reliability...... motion analysis system prior to the recordings and conclusions should take measurement variations (3-8 degrees and 9-14 degrees for within and between day, respectively) into account. LEVEL OF EVIDENCE: 3....

  19. Reliability of video-based quantification of the knee- and hip angle at foot strike during running

    DEFF Research Database (Denmark)

    Damsted, Camma; Oestergaard Nielsen, Rasmus; Larsen, Lars Henrik

    2014-01-01

    INTRODUCTION: In clinical practice, joint kinematics during running are primarily quantified by two-dimensional (2D) video recordings and motion-analysis software. The applicability of this approach depends on the clinicians' ability to quantify kinematics in a reliable manner. The reliability...... motion analysis system prior to the recordings and conclusions should take measurement variations (3-8 degrees and 9-14 degrees for within and between day, respectively) into account....

  20. A reliable method of quantification of trace copper in beverages with and without alcohol by spectrophotometry after cloud point extraction

    Directory of Open Access Journals (Sweden)

    Ramazan Gürkan

    2013-01-01

    Full Text Available A new cloud point extraction (CPE method was developed for the separation and preconcentration of copper (II prior to spectrophotometric analysis. For this purpose, 1-(2,4-dimethylphenyl azonapthalen-2-ol (Sudan II was used as a chelating agent and the solution pH was adjusted to 10.0 with borate buffer. Polyethylene glycol tert-octylphenyl ether (Triton X-114 was used as an extracting agent in the presence of sodium dodecylsulphate (SDS. After phase separation, based on the cloud point of the mixture, the surfactant-rich phase was diluted with acetone, and the enriched analyte was spectrophotometrically determined at 537 nm. The variables affecting CPE efficiency were optimized. The calibration curve was linear within the range 0.285-20 µg L-1 with a detection limit of 0.085 µg L-1. The method was successfully applied to the quantification of copper in different beverage samples.

  1. DNA microsatellite region for a reliable quantification of soft wheat adulteration in durum wheat-based foodstuffs by real-time PCR.

    Science.gov (United States)

    Sonnante, Gabriella; Montemurro, Cinzia; Morgese, Anita; Sabetta, Wilma; Blanco, Antonio; Pasqualone, Antonella

    2009-11-11

    Italian industrial pasta and durum wheat typical breads must be prepared using exclusively durum wheat semolina. Previously, a microsatellite sequence specific of the wheat D-genome had been chosen for traceability of soft wheat in semolina and bread samples, using qualitative and quantitative Sybr green-based real-time experiments. In this work, we describe an improved method based on the same soft wheat genomic region by means of a quantitative real-time PCR using a dual-labeled probe. Standard curves based on dilutions of 100% soft wheat flour, pasta, or bread were constructed. Durum wheat semolina, pasta, and bread samples were prepared with increasing amounts of soft wheat to verify the accuracy of the method. Results show that reliable quantifications were obtained especially for the samples containing a lower amount of soft wheat DNA, fulfilling the need to verify labeling of pasta and typical durum wheat breads.

  2. Design and performance testing of a DNA extraction assay for sensitive and reliable quantification of acetic acid bacteria directly in red wine using real time PCR

    Directory of Open Access Journals (Sweden)

    Cédric eLONGIN

    2016-06-01

    Full Text Available Although strategies exist to prevent AAB contamination, the increased interest for wines with low sulfite addition leads to greater AAB spoilage. Hence there is a real need for a rapid, specific, sensitive and reliable method for detecting these spoilage bacteria. All these requirements are met by real time Polymerase Chain Reaction (or quantitative PCR; qPCR. Here, we compare existing methods of isolating DNA and their adaptation to a red wine matrix. Two different protocols for isolating DNA and three PCR mix compositions were tested to select the best method. The addition of insoluble polyvinylpolypyrrolidone (PVPP at 1% (v/v during DNA extraction using a protocol succeeded in eliminating PCR inhibitors from red wine. We developed a bacterial internal control which was efficient in avoiding false negative results due to decreases in the efficiency of DNA isolation and/or amplification. The specificity, linearity, repeatability and reproducibility of the method were evaluated. A standard curve was established for the enumeration of AAB inoculated into red wines. The limit of quantification in red wine was 3.7 log AAB/mL and about 2.8 log AAB/mL when the volume of the samples was increased from 1 mL to 10 mL. Thus the DNA extraction method developed in this paper allows sensitive and reliable AAB quantification without underestimation thanks to the presence of an internal control. Moreover, monitoring of both the AAB population and the amount of acetic acid in ethanol medium and red wine highlighted that a minimum about 6.0 log cells/mL of AAB is needed to significantly increase the production of acetic acid leading to spoilage.

  3. Assessment of the human factor in the quantification of technical system reliability taking into consideration cognitive-causal aspects. Partial project 2. Modeling of the human behavior for reliability considerations. Final report

    International Nuclear Information System (INIS)

    Jennerich, Marco; Imbsweiler, Jonas; Straeter, Oliver; Arenius, Marcus

    2015-03-01

    This report presents the findings of the project for the consideration of human factor in the quantification of the reliability of technical systems, taking into account cognitive-causal aspects concerning the modeling of human behavior of reliability issues (funded by the Federal Ministry of Economics and Technology; grant number 15014328). This project is part of a joint project with the University of Applied Sciences Zittau / Goerlitz for assessing the human factor in the quantification of the reliability of technical systems. The concern of the University of Applied Sciences Zittau / Goerlitz is the mathematical modeling of human reliability by means of a fuzzy set approach (grant number 1501432A). The part of the project presented here provides the necessary data basis for the evaluation of the mathematical modeling using fuzzy set approach. At the appropriate places in this report, the interfaces and data bases between the two projects are outlined accordingly. HRA-methods (Human Reliability Analysis) are an essential component to analyze the reliability of socio-technical systems. Various methods have been established and are used in different areas of application. The established HRA methods have been checked on their congruence. In particular the underlying models and their parameters such as performance-influencing factors and situational influences have been investigated. The elaborated parameters were combined into a hierarchical class structure. Cross-domain incidents were studied. The specific performance-influencing factors have been worked out and have been integrated into a cross-domain database. The dominant (critical) situational factors and their interactions within the event data were identified using the CAHR method (connectionism Assessment of Human Reliability). Task dependent cognitive load profiles have been defined. Within these profiles qualitative and quantitative data of the possibility of emergence of errors have been acquired. This

  4. Quantification of the reliability of personnel actions from the evaluation of actual German operational experience. Final report

    International Nuclear Information System (INIS)

    Preischl, W.; Fassmann, W.

    2013-07-01

    The results and their uncertainty bounds of PSA studies are considerably impacted by the assessment of human reliability. But the amount of available, generic data is not sufficient to evaluate all human actions considered in a modern PSA study adequately. Further the data are not sufficiently validated and rely as well as the proposed uncertainty bounds on expert judgement. This research project as well as the preceding project /GRS 10/ validated data recommended by the German PSA Guidelines and enlarged the amount of available data. The findings may contribute to an update of the German PSA Guidelines. In a first step of the project information about reportable events in German nuclear power plants with observed human errors (event reports, expert statements, technical documents, interviews and plant walk downs with subject matter experts from the plants) were analysed. The investigation resulted in 67 samples describing personal activities, performance conditions, the number of observed errors and the number of action performance. In a second step a new methodology was developed and applied in a pilot plant. The objective was to identify undoubtedly error free safety relevant actions, their performance conditions, and frequency as well as to prove and demonstrate that probabilistic data can be derived from that operational experience (OE). The application in the pilot plant resulted in 18 ''error free'' samples characterizing human reliability. All available samples were evaluated by use of the method of Bayes. That commonly accepted methodology was applied in order to derive probabilistic data based on samples taken from operational experience. A thorough analysis of the obtained results shows that both data sources (OE reportable events, OE with undoubtedly error free action performance) provide data with comparable quality and validity. At the end of the research project the following products are available. - Methods to select samples

  5. Reliable Quantification of the Potential for Equations Based on Spot Urine Samples to Estimate Population Salt Intake

    DEFF Research Database (Denmark)

    Huang, Liping; Crino, Michelle; Wu, Jason Hy

    2016-01-01

    to a standard format. Individual participant records will be compiled and a series of analyses will be completed to: (1) compare existing equations for estimating 24-hour salt intake from spot urine samples with 24-hour urine samples, and assess the degree of bias according to key demographic and clinical......BACKGROUND: Methods based on spot urine samples (a single sample at one time-point) have been identified as a possible alternative approach to 24-hour urine samples for determining mean population salt intake. OBJECTIVE: The aim of this study is to identify a reliable method for estimating mean...... population salt intake from spot urine samples. This will be done by comparing the performance of existing equations against one other and against estimates derived from 24-hour urine samples. The effects of factors such as ethnicity, sex, age, body mass index, antihypertensive drug use, health status...

  6. APPRIS 2017: principal isoforms for multiple gene sets

    Science.gov (United States)

    Rodriguez-Rivas, Juan; Di Domenico, Tomás; Vázquez, Jesús; Valencia, Alfonso

    2018-01-01

    Abstract The APPRIS database (http://appris-tools.org) uses protein structural and functional features and information from cross-species conservation to annotate splice isoforms in protein-coding genes. APPRIS selects a single protein isoform, the ‘principal’ isoform, as the reference for each gene based on these annotations. A single main splice isoform reflects the biological reality for most protein coding genes and APPRIS principal isoforms are the best predictors of these main proteins isoforms. Here, we present the updates to the database, new developments that include the addition of three new species (chimpanzee, Drosophila melangaster and Caenorhabditis elegans), the expansion of APPRIS to cover the RefSeq gene set and the UniProtKB proteome for six species and refinements in the core methods that make up the annotation pipeline. In addition APPRIS now provides a measure of reliability for individual principal isoforms and updates with each release of the GENCODE/Ensembl and RefSeq reference sets. The individual GENCODE/Ensembl, RefSeq and UniProtKB reference gene sets for six organisms have been merged to produce common sets of splice variants. PMID:29069475

  7. reliability reliability

    African Journals Online (AJOL)

    eobe

    Corresponding author, Tel: +234-703. RELIABILITY .... V , , given by the code of practice. However, checks must .... an optimization procedure over the failure domain F corresponding .... of Concrete Members based on Utility Theory,. Technical ...

  8. Human reliability

    International Nuclear Information System (INIS)

    Embrey, D.E.

    1987-01-01

    Concepts and techniques of human reliability have been developed and are used mostly in probabilistic risk assessment. For this, the major application of human reliability assessment has been to identify the human errors which have a significant effect on the overall safety of the system and to quantify the probability of their occurrence. Some of the major issues within human reliability studies are reviewed and it is shown how these are applied to the assessment of human failures in systems. This is done under the following headings; models of human performance used in human reliability assessment, the nature of human error, classification of errors in man-machine systems, practical aspects, human reliability modelling in complex situations, quantification and examination of human reliability, judgement based approaches, holistic techniques and decision analytic approaches. (UK)

  9. Quantification in emission tomography

    International Nuclear Information System (INIS)

    Buvat, Irene

    2011-11-01

    The objective of this lecture is to understand the possibilities and limitations of the quantitative analysis of single photon emission computed tomography (SPECT) and positron emission tomography (PET) images. It is also to identify the conditions to be fulfilled to obtain reliable quantitative measurements from images. Content: 1 - Introduction: Quantification in emission tomography - definition and challenges; quantification biasing phenomena 2 - Main problems impacting quantification in PET and SPECT: problems, consequences, correction methods, results (Attenuation, scattering, partial volume effect, movement, un-stationary spatial resolution in SPECT, fortuitous coincidences in PET, standardisation in PET); 3 - Synthesis: accessible efficiency, know-how, Precautions, beyond the activity measurement

  10. Statistical modeling of isoform splicing dynamics from RNA-seq time series data.

    Science.gov (United States)

    Huang, Yuanhua; Sanguinetti, Guido

    2016-10-01

    Isoform quantification is an important goal of RNA-seq experiments, yet it remains problematic for genes with low expression or several isoforms. These difficulties may in principle be ameliorated by exploiting correlated experimental designs, such as time series or dosage response experiments. Time series RNA-seq experiments, in particular, are becoming increasingly popular, yet there are no methods that explicitly leverage the experimental design to improve isoform quantification. Here, we present DICEseq, the first isoform quantification method tailored to correlated RNA-seq experiments. DICEseq explicitly models the correlations between different RNA-seq experiments to aid the quantification of isoforms across experiments. Numerical experiments on simulated datasets show that DICEseq yields more accurate results than state-of-the-art methods, an advantage that can become considerable at low coverage levels. On real datasets, our results show that DICEseq provides substantially more reproducible and robust quantifications, increasing the correlation of estimates from replicate datasets by up to 10% on genes with low or moderate expression levels (bottom third of all genes). Furthermore, DICEseq permits to quantify the trade-off between temporal sampling of RNA and depth of sequencing, frequently an important choice when planning experiments. Our results have strong implications for the design of RNA-seq experiments, and offer a novel tool for improved analysis of such datasets. Python code is freely available at http://diceseq.sf.net G.Sanguinetti@ed.ac.uk Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. Accident sequence quantification with KIRAP

    International Nuclear Information System (INIS)

    Kim, Tae Un; Han, Sang Hoon; Kim, Kil You; Yang, Jun Eon; Jeong, Won Dae; Chang, Seung Cheol; Sung, Tae Yong; Kang, Dae Il; Park, Jin Hee; Lee, Yoon Hwan; Hwang, Mi Jeong.

    1997-01-01

    The tasks of probabilistic safety assessment(PSA) consists of the identification of initiating events, the construction of event tree for each initiating event, construction of fault trees for event tree logics, the analysis of reliability data and finally the accident sequence quantification. In the PSA, the accident sequence quantification is to calculate the core damage frequency, importance analysis and uncertainty analysis. Accident sequence quantification requires to understand the whole model of the PSA because it has to combine all event tree and fault tree models, and requires the excellent computer code because it takes long computation time. Advanced Research Group of Korea Atomic Energy Research Institute(KAERI) has developed PSA workstation KIRAP(Korea Integrated Reliability Analysis Code Package) for the PSA work. This report describes the procedures to perform accident sequence quantification, the method to use KIRAP's cut set generator, and method to perform the accident sequence quantification with KIRAP. (author). 6 refs

  12. Accident sequence quantification with KIRAP

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Tae Un; Han, Sang Hoon; Kim, Kil You; Yang, Jun Eon; Jeong, Won Dae; Chang, Seung Cheol; Sung, Tae Yong; Kang, Dae Il; Park, Jin Hee; Lee, Yoon Hwan; Hwang, Mi Jeong

    1997-01-01

    The tasks of probabilistic safety assessment(PSA) consists of the identification of initiating events, the construction of event tree for each initiating event, construction of fault trees for event tree logics, the analysis of reliability data and finally the accident sequence quantification. In the PSA, the accident sequence quantification is to calculate the core damage frequency, importance analysis and uncertainty analysis. Accident sequence quantification requires to understand the whole model of the PSA because it has to combine all event tree and fault tree models, and requires the excellent computer code because it takes long computation time. Advanced Research Group of Korea Atomic Energy Research Institute(KAERI) has developed PSA workstation KIRAP(Korea Integrated Reliability Analysis Code Package) for the PSA work. This report describes the procedures to perform accident sequence quantification, the method to use KIRAP`s cut set generator, and method to perform the accident sequence quantification with KIRAP. (author). 6 refs.

  13. A risk-informed approach of quantification of epistemic uncertainty for the long-term radioactive waste disposal. Improving reliability of expert judgements with an advanced elicitation procedure

    International Nuclear Information System (INIS)

    Sugiyama, Daisuke; Chida, Taiji; Fujita, Tomonari; Tsukamoto, Masaki

    2011-01-01

    A quantification methodology of epistemic uncertainty by expert judgement based on the risk-informed approach is developed to assess inevitable uncertainty for the long-term safety assessment of radioactive waste disposal. The proposed method in this study employs techniques of logic tree, by which options of models and/or scenarios are identified, and Evidential Support Logic (ESL), by which possibility of each option is quantified. In this report, the effect of a feedback process of discussion between experts and input of state-of-the-art knowledge in the proposed method is discussed to estimate alteration of the distribution of expert judgements which is one of the factors causing uncertainty. In a preliminary quantification experiment of uncertainty of degradation of the engineering barrier materials in a tentative sub-surface disposal using the proposed methodology, experts themselves modified questions appropriately to facilitate sound judgements and to correlate those with scientific evidences clearly. The result suggests that the method effectively improves confidence of expert judgement. Also, the degree of consensus of expert judgement was sort of improved in some cases, since scientific knowledge and information of expert judgement in other fields became common understanding. It is suggested that the proposed method could facilitate consensus on uncertainty between interested persons. (author)

  14. Characterization of ductal and lobular breast carcinomas using novel prolactin receptor isoform specific antibodies

    Directory of Open Access Journals (Sweden)

    Heger Christopher D

    2010-12-01

    Full Text Available Abstract Background Prolactin is a polypeptide hormone responsible for proliferation and differentiation of the mammary gland. More recently, prolactin's role in mammary carcinogenesis has been studied with greater interest. Studies from our laboratory and from others have demonstrated that three specific isoforms of the prolactin receptor (PRLR are expressed in both normal and cancerous breast cells and tissues. Until now, reliable isoform specific antibodies have been lacking. We have prepared and characterized polyclonal antibodies against each of the human PRLR isoforms that can effectively be used to characterize human breast cancers. Methods Rabbits were immunized with synthetic peptides of isoform unique regions and immune sera affinity purified prior to validation by Western blot and immunohistochemical analyses. Sections of ductal and lobular carcinomas were stained with each affinity purified isoform specific antibody to determine expression patterns in breast cancer subclasses. Results We show that the rabbit antibodies have high titer and could specifically recognize each isoform of PRLR. Differences in PRLR isoform expression levels were observed and quantified using histosections from xenografts of established human breast cancer cells lines, and ductal and lobular carcinoma human biopsy specimens. In addition, these results were verified by real-time PCR with isoform specific primers. While nearly all tumors contained LF and SF1b, the majority (76% of ductal carcinoma biopsies expressed SF1a while the majority of lobular carcinomas lacked SF1a staining (72% and 27% had only low levels of expression. Conclusions Differences in the receptor isoform expression profiles may be critical to understanding the role of PRL in mammary tumorigenesis. Since these antibodies are specifically directed against each PRLR isoform, they are valuable tools for the evaluation of breast cancer PRLR content and have potential clinical importance in

  15. Simultaneous Detection of Human C-Terminal p53 Isoforms by Single Template Molecularly Imprinted Polymers (MIPs) Coupled with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS)-Based Targeted Proteomics.

    Science.gov (United States)

    Jiang, Wenting; Liu, Liang; Chen, Yun

    2018-03-06

    Abnormal expression of C-terminal p53 isoforms α, β, and γ can cause the development of cancers including breast cancer. To date, much evidence has demonstrated that these isoforms can differentially regulate target genes and modulate their expression. Thus, quantification of individual isoforms may help to link clinical outcome to p53 status and to improve cancer patient treatment. However, there are few studies on accurate determination of p53 isoforms, probably due to sequence homology of these isoforms and also their low abundance. In this study, a targeted proteomics assay combining molecularly imprinted polymers (MIPs) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed for simultaneous quantification of C-terminal p53 isoforms. Isoform-specific surrogate peptides (i.e., KPLDGEYFTLQIR (peptide-α) for isoform α, KPLDGEYFTLQDQTSFQK (peptide-β) for isoform β, and KPLDGEYFTLQMLLDLR (peptide-γ) for isoform γ) were first selected and used in both MIPs enrichment and mass spectrometric detection. The common sequence KPLDGEYFTLQ of these three surrogate peptides was used as single template in MIPs. In addition to optimization of imprinting conditions and characterization of the prepared MIPs, binding affinity and cross-reactivity of the MIPs for each surrogate peptide were also evaluated. As a result, a LOQ of 5 nM was achieved, which was >15-fold more sensitive than that without MIPs. Finally, the assay was validated and applied to simultaneous quantitative analysis of C-terminal p53 isoforms α, β, and γ in several human breast cell lines (i.e., MCF-10A normal cells, MCF-7 and MDA-MB-231 cancer cells, and drug-resistant MCF-7/ADR cancer cells). This study is among the first to employ single template MIPs and cross-reactivity phenomenon to select isoform-specific surrogate peptides and enable simultaneous quantification of protein isoforms in LC-MS/MS-based targeted proteomics.

  16. The test-retest reliability of anatomical co-ordinate axes definition for the quantification of lower extremity kinematics during running.

    Science.gov (United States)

    Sinclair, Jonathan; Taylor, Paul John; Greenhalgh, Andrew; Edmundson, Christopher James; Brooks, Darrell; Hobbs, Sarah Jane

    2012-12-01

    Three-dimensional (3-D) kinematic analyses are used widely in both sport and clinical examinations. However, this procedure depends on reliable palpation of anatomical landmarks and mal-positioning of markers between sessions may result in improperly defined segment co-ordinate system axes which will produce in-consistent joint rotations. This had led some to question the efficacy of this technique. The aim of the current investigation was to assess the reliability of the anatomical frame definition when quantifying 3-D kinematics of the lower extremities during running. Ten participants completed five successful running trials at 4.0 m·s(-1) ± 5%. 3-D angular joint kinematics parameters from the hip, knee and ankle were collected using an eight camera motion analysis system. Two static calibration trials were captured. The first (test) was conducted prior to the running trials following which anatomical landmarks were removed. The second was obtained following completion of the running trials where anatomical landmarks were re-positioned (retest). Paired samples t-tests were used to compare 3-D kinematic parameters quantified using the two static trials, and intraclass correlations were employed to examine the similarities between the sagittal, coronal and transverse plane waveforms. The results indicate that no significant (p>0.05) differences were found between test and retest 3-D kinematic parameters and strong (R(2)≥0.87) correlations were observed between test and retest waveforms. Based on the results obtained from this investigation, it appears that the anatomical co-ordinate axes of the lower extremities can be defined reliably thus confirming the efficacy of studies using this technique.

  17. Quantification of intervertebral displacement with a novel MRI-based modeling technique: Assessing measurement bias and reliability with a porcine spine model.

    Science.gov (United States)

    Mahato, Niladri K; Montuelle, Stephane; Goubeaux, Craig; Cotton, John; Williams, Susan; Thomas, James; Clark, Brian C

    2017-05-01

    The purpose of this study was to develop a novel magnetic resonance imaging (MRI)-based modeling technique for measuring intervertebral displacements. Here, we present the measurement bias and reliability of the developmental work using a porcine spine model. Porcine lumbar vertebral segments were fitted in a custom-built apparatus placed within an externally calibrated imaging volume of an open-MRI scanner. The apparatus allowed movement of the vertebrae through pre-assigned magnitudes of sagittal and coronal translation and rotation. The induced displacements were imaged with static (T 1 ) and fast dynamic (2D HYCE S) pulse sequences. These images were imported into animation software, in which these images formed a background 'scene'. Three-dimensional models of vertebrae were created using static axial scans from the specimen and then transferred into the animation environment. In the animation environment, the user manually moved the models (rotoscoping) to perform model-to-'scene' matching to fit the models to their image silhouettes and assigned anatomical joint axes to the motion-segments. The animation protocol quantified the experimental translation and rotation displacements between the vertebral models. Accuracy of the technique was calculated as 'bias' using a linear mixed effects model, average percentage error and root mean square errors. Between-session reliability was examined by computing intra-class correlation coefficients (ICC) and the coefficient of variations (CV). For translation trials, a constant bias (β 0 ) of 0.35 (±0.11) mm was detected for the 2D HYCE S sequence (p=0.01). The model did not demonstrate significant additional bias with each mm increase in experimental translation (β 1 Displacement=0.01mm; p=0.69). Using the T 1 sequence for the same assessments did not significantly change the bias (p>0.05). ICC values for the T 1 and 2D HYCE S pulse sequences were 0.98 and 0.97, respectively. For rotation trials, a constant bias (

  18. A construction of standardized near infrared hyper-spectral teeth database: a first step in the development of reliable diagnostic tool for quantification and early detection of caries

    Science.gov (United States)

    Bürmen, Miran; Usenik, Peter; Fidler, Aleš; Pernuš, Franjo; Likar, Boštjan

    2011-03-01

    Dental caries is a disease characterized by demineralization of enamel crystals leading to the penetration of bacteria into the dentin and pulp. If left untreated, the disease can lead to pain, infection and tooth loss. Early detection of enamel demineralization resulting in increased enamel porosity, commonly known as white spots, is a difficult diagnostic task. Several papers reported on near infrared (NIR) spectroscopy to be a potentially useful noninvasive spectroscopic technique for early detection of caries lesions. However, the conducted studies were mostly qualitative and did not include the critical assessment of the spectral variability of the sound and carious dental tissues and influence of the water content. Such assessment is essential for development and validation of reliable qualitative and especially quantitative diagnostic tools based on NIR spectroscopy. In order to characterize the described spectral variability, a standardized diffuse reflectance hyper-spectral database was constructed by imaging 12 extracted human teeth with natural lesions of various degrees in the spectral range from 900 to 1700 nm with spectral resolution of 10 nm. Additionally, all the teeth were imaged by digital color camera. The influence of water content on the acquired spectra was characterized by monitoring the teeth during the drying process. The images were assessed by an expert, thereby obtaining the gold standard. By analyzing the acquired spectra we were able to accurately model the spectral variability of the sound dental tissues and identify the advantages and limitations of NIR hyper-spectral imaging.

  19. Inter- and intra-rater reliability of patellofemoral kinematic and contact area quantification by fast spin echo MRI and correlation with cartilage health by quantitative T1ρ MRI.

    Science.gov (United States)

    Lau, Brian C; Thuillier, Daniel U; Pedoia, Valentina; Chen, Ellison Y; Zhang, Zhihong; Feeley, Brian T; Souza, Richard B

    2016-01-01

    Patellar maltracking is a leading cause of patellofemoral pain syndrome (PFPS). The aim of this study was to determine the inter- and intra-rater reliability of a semi-automated program for magnetic resonance imaging (MRI) based patellofemoral kinematics. Sixteen subjects (10 with PFPS [mean age 32.3; SD 5.2; eight females] and six controls without PFPS 19 [mean age 28.6; SD 2.8; three females]) participated in the study. One set of T2-weighted, fat-saturated fast spin-echo (FSE) MRIs were acquired from each subject in full extension and 30° of knee flexion. MRI including axial T1ρ relaxation time mapping sequences was also performed on each knee. Following image acquisitions, regions of interest for kinematic MRI, and patellar and trochlear cartilage were segmented and quantified with in-house designed spline- based MATLAB semi-automated software. Intraclass Correlations Coefficients (ICC) of calculated kinematic parameters were good to excellent, ICC > 0.8 in patellar flexion, rotation, tilt, and translation (anterior -posterior, medial -lateral, and superior -inferior), and contact area translation. Only patellar tilt in the flexed position and motion from extended to flexed state was significantly different between PFPS and control patients (p=0.002 and p=0.006, respectively). No significant correlations were identified between patellofemoral kinematics and contact area with T1ρ relaxation times. A semi-automated, spline-based kinematic MRI technique for patellofemoral kinematic and contact area quantification is highly reproducible with the potential to help better understand the role of patellofemoral maltracking in PFPS and other knee disorders. Level IV. Published by Elsevier B.V.

  20. Inter- and intra-rater reliability of patellofemoral kinematic and contact area quantification by fast spin echo MRI and correlation with cartilage health by quantitative T1ρ MRI☆

    Science.gov (United States)

    Lau, Brian C.; Thuillier, Daniel U.; Pedoia, Valentina; Chen, Ellison Y.; Zhang, Zhihong; Feeley, Brian T.; Souza, Richard B.

    2016-01-01

    Background Patellar maltracking is a leading cause of patellofemoral pain syndrome (PFPS). The aim of this study was to determine the inter- and intra-rater reliability of a semi-automated program for magnetic resonance imaging (MRI) based patellofemoral kinematics. Methods Sixteen subjects (10 with PFPS [mean age 32.3; SD 5.2; eight females] and six controls without PFPS 19 [mean age 28.6; SD 2.8; three females]) participated in the study. One set of T2-weighted, fat-saturated fast spin-echo (FSE) MRIs were acquired from each subject in full extension and 30° of knee flexion. MRI including axial T1ρ relaxation time mapping sequences was also performed on each knee. Following image acquisitions, regions of interest for kinematic MRI, and patellar and trochlear cartilage were segmented and quantified with in-house designed spline- based MATLAB semi-automated software. Results Intraclass Correlations Coefficients (ICC) of calculated kinematic parameters were good to excellent, ICC > 0.8 in patellar flexion, rotation, tilt, and translation (anterior -posterior, medial -lateral, and superior -inferior), and contact area translation. Only patellar tilt in the flexed position and motion from extended to flexed state was significantly different between PFPS and control patients (p = 0.002 and p = 0.006, respectively). No significant correlations were identified between patellofemoral kinematics and contact area with T1ρ relaxation times. Conclusions A semi-automated, spline-based kinematic MRI technique for patellofemoral kinematic and contact area quantification is highly reproducible with the potential to help better understand the role of patellofemoral maltracking in PFPS and other knee disorders. PMID:26746045

  1. Reliability engineering

    International Nuclear Information System (INIS)

    Lee, Chi Woo; Kim, Sun Jin; Lee, Seung Woo; Jeong, Sang Yeong

    1993-08-01

    This book start what is reliability? such as origin of reliability problems, definition of reliability and reliability and use of reliability. It also deals with probability and calculation of reliability, reliability function and failure rate, probability distribution of reliability, assumption of MTBF, process of probability distribution, down time, maintainability and availability, break down maintenance and preventive maintenance design of reliability, design of reliability for prediction and statistics, reliability test, reliability data and design and management of reliability.

  2. Improved mass spectrometry assay for plasma hepcidin: detection and characterization of a novel hepcidin isoform.

    Directory of Open Access Journals (Sweden)

    Coby M M Laarakkers

    Full Text Available Mass spectrometry (MS-based assays for the quantification of the iron regulatory hormone hepcidin are pivotal to discriminate between the bioactive 25-amino acid form that can effectively block the sole iron transporter ferroportin and other naturally occurring smaller isoforms without a known role in iron metabolism. Here we describe the design, validation and use of a novel stable hepcidin-25(+40 isotope as internal standard for quantification. Importantly, the relative large mass shift of 40 Da makes this isotope also suitable for easy-to-use medium resolution linear time-of-flight (TOF platforms. As expected, implementation of hepcidin-25(+40 as internal standard in our weak cation exchange (WCX TOF MS method yielded very low inter/intra run coefficients of variation. Surprisingly, however, in samples from kidney disease patients, we detected a novel peak (m/z 2673.9 with low intensity that could be identified as hepcidin-24 and had previously remained unnoticed due to peak interference with the formerly used internal standard. Using a cell-based bioassay it was shown that synthetic hepcidin-24 was, like the -22 and -20 isoforms, a significantly less potent inducer of ferroportin degradation than hepcidin-25. During prolonged storage of plasma at room temperature, we observed that a decrease in plasma hepcidin-25 was paralleled by an increase in the levels of the hepcidin-24, -22 and -20 isoforms. This provides first evidence that all determinants for the conversion of hepcidin-25 to smaller inactive isoforms are present in the circulation, which may contribute to the functional suppression of hepcidin-25, that is significantly elevated in patients with renal impairment. The present update of our hepcidin TOF MS assay together with improved insights in the source and preparation of the internal standard, and sample stability will further improve our understanding of circulating hepcidin and pave the way towards further optimization and

  3. Functional studies of sodium pump isoforms

    DEFF Research Database (Denmark)

    Clausen, Michael Jakob

    The Na+,K+-ATPase is an essential ion pump found in all animal cells. It uses the energy from ATP hydrolysis to export three Na+ and import two K+, both against their chemical gradients and for Na+ also against the electrical potential. Mammals require four Na+,K+-ATPase isoforms that each have...... unique expression profiles and specialized functional features. We use a Two Electrode Voltage Clamp setup to determine pre-steady-state and steady-state characteristics of each isoform and design chimeras to pin-point the structural elements responsible for observed differences. With this strategy we...

  4. Lipoprotein lipase isoelectric point isoforms in humans

    DEFF Research Database (Denmark)

    Badia-Villanueva, M.; Carulla, P.; Carrascal, M.

    2014-01-01

    -heparin plasma (PHP), LPL consists of a pattern of more than 8 forms of the same apparent molecular weight, but different isoelectric point (pI). In the present study we describe, for the first time, the existence of at least nine LPL pI isoforms in human PHP, with apparent pI between 6.8 and 8.6. Separation...

  5. Quantitative evaluation of alternatively spliced mRNA isoforms by label-free real-time plasmonic sensing.

    Science.gov (United States)

    Huertas, César S; Carrascosa, L G; Bonnal, S; Valcárcel, J; Lechuga, L M

    2016-04-15

    Alternative splicing of mRNA precursors enables cells to generate different protein outputs from the same gene depending on their developmental or homeostatic status. Its deregulation is strongly linked to disease onset and progression. Current methodologies for monitoring alternative splicing demand elaborate procedures and often present difficulties in discerning between closely related isoforms, e.g. due to cross-hybridization during their detection. Herein, we report a general methodology using a Surface Plasmon Resonance (SPR) biosensor for label-free monitoring of alternative splicing events in real-time, without any cDNA synthesis or PCR amplification requirements. We applied this methodology to RNA isolated from HeLa cells for the quantification of alternatively spliced isoforms of the Fas gene, involved in cancer progression through regulation of programmed cell death. We demonstrate that our methodology is isoform-specific, with virtually no cross-hybridization, achieving limits of detection (LODs) in the picoMolar (pM) range. Similar results were obtained for the detection of the BCL-X gene mRNA isoforms. The results were independently validated by RT-qPCR, with excellent concordance in the determination of isoform ratios. The simplicity and robustness of this biosensor technology can greatly facilitate the exploration of alternative splicing biomarkers in disease diagnosis and therapy. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Superposition Quantification

    Science.gov (United States)

    Chang, Li-Na; Luo, Shun-Long; Sun, Yuan

    2017-11-01

    The principle of superposition is universal and lies at the heart of quantum theory. Although ever since the inception of quantum mechanics a century ago, superposition has occupied a central and pivotal place, rigorous and systematic studies of the quantification issue have attracted significant interests only in recent years, and many related problems remain to be investigated. In this work we introduce a figure of merit which quantifies superposition from an intuitive and direct perspective, investigate its fundamental properties, connect it to some coherence measures, illustrate it through several examples, and apply it to analyze wave-particle duality. Supported by Science Challenge Project under Grant No. TZ2016002, Laboratory of Computational Physics, Institute of Applied Physics and Computational Mathematics, Beijing, Key Laboratory of Random Complex Structures and Data Science, Chinese Academy of Sciences, Grant under No. 2008DP173182

  7. WT1 isoform expression pattern in acute myeloid leukemia.

    Science.gov (United States)

    Luna, Irene; Such, Esperanza; Cervera, Jose; Barragán, Eva; Ibañez, Mariam; Gómez-Seguí, Inés; López-Pavía, María; Llop, Marta; Fuster, Oscar; Dolz, Sandra; Oltra, Silvestre; Alonso, Carmen; Vera, Belén; Lorenzo, Ignacio; Martínez-Cuadrón, David; Montesinos, Pau; Senent, M Leonor; Moscardó, Federico; Bolufer, Pascual; Sanz, Miguel A

    2013-12-01

    WT1 plays a dual role in leukemia development, probably due to an imbalance in the expression of the 4 main WT1 isoforms. We quantify their expression and evaluate them in a series of AML patients. Our data showed a predominant expression of isoform D in AML, although in a lower quantity than in normal CD34+ cells. We found a positive correlation between the total WT1 expression and A, B and C isoforms. The overexpression of WT1 in AML might be due to a relative increase in A, B and C isoforms, together with a relative decrease in isoform D expression. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Improved LC-MS/MS method for the quantification of hepcidin-25 in clinical samples.

    Science.gov (United States)

    Abbas, Ioana M; Hoffmann, Holger; Montes-Bayón, María; Weller, Michael G

    2018-06-01

    Mass spectrometry-based methods play a crucial role in the quantification of the main iron metabolism regulator hepcidin by singling out the bioactive 25-residue peptide from the other naturally occurring N-truncated isoforms (hepcidin-20, -22, -24), which seem to be inactive in iron homeostasis. However, several difficulties arise in the MS analysis of hepcidin due to the "sticky" character of the peptide and the lack of suitable standards. Here, we propose the use of amino- and fluoro-silanized autosampler vials to reduce hepcidin interaction to laboratory glassware surfaces after testing several types of vials for the preparation of stock solutions and serum samples for isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS). Furthermore, we have investigated two sample preparation strategies and two chromatographic separation conditions with the aim of developing a LC-MS/MS method for the sensitive and reliable quantification of hepcidin-25 in serum samples. A chromatographic separation based on usual acidic mobile phases was compared with a novel approach involving the separation of hepcidin-25 with solvents at high pH containing 0.1% of ammonia. Both methods were applied to clinical samples in an intra-laboratory comparison of two LC-MS/MS methods using the same hepcidin-25 calibrators with good correlation of the results. Finally, we recommend a LC-MS/MS-based quantification method with a dynamic range of 0.5-40 μg/L for the assessment of hepcidin-25 in human serum that uses TFA-based mobile phases and silanized glass vials. Graphical abstract Structure of hepcidin-25 (Protein Data Bank, PDB ID 2KEF).

  9. Reliability Engineering

    International Nuclear Information System (INIS)

    Lee, Sang Yong

    1992-07-01

    This book is about reliability engineering, which describes definition and importance of reliability, development of reliability engineering, failure rate and failure probability density function about types of it, CFR and index distribution, IFR and normal distribution and Weibull distribution, maintainability and movability, reliability test and reliability assumption in index distribution type, normal distribution type and Weibull distribution type, reliability sampling test, reliability of system, design of reliability and functionality failure analysis by FTA.

  10. Towards the production of reliable quantitative microbiological data for risk assessment: Direct quantification of Campylobacter in naturally infected chicken fecal samples using selective culture and real-time PCR

    DEFF Research Database (Denmark)

    Garcia Clavero, Ana Belén; Vigre, Håkan; Josefsen, Mathilde Hasseldam

    2015-01-01

    of Campylobacter by real-time PCR was performed using standard curves designed for two different DNA extraction methods: Easy-DNA™ Kit from Invitrogen (Easy-DNA) and NucliSENS® MiniMAG® from bioMérieux (MiniMAG). Results indicated that the estimation of the numbers of Campylobacter present in chicken fecal samples...... and for the evaluation of control strategies implemented in poultry production. The aim of this study was to compare estimates of the numbers of Campylobacter spp. in naturally infected chicken fecal samples obtained using direct quantification by selective culture and by real-time PCR. Absolute quantification....... Although there were differences in terms of estimates of Campylobacter numbers between the methods and samples, the differences between culture and real-time PCR were not statistically significant for most of the samples used in this study....

  11. Direct qPCR quantification using the Quantifiler(®) Trio DNA quantification kit.

    Science.gov (United States)

    Liu, Jason Yingjie

    2014-11-01

    The effectiveness of a direct quantification assay is essential to the adoption of the combined direct quantification/direct STR workflow. In this paper, the feasibility of using the Quantifiler(®) Trio DNA quantification kit for the direct quantification of forensic casework samples was investigated. Both low-level touch DNA samples and blood samples were collected on PE swabs and quantified directly. The increased sensitivity of the Quantifiler(®) Trio kit enabled the detection of less than 10pg of DNA in unprocessed touch samples and also minimizes the stochastic effect experienced by different targets in the same sample. The DNA quantity information obtained from a direct quantification assay using the Quantifiler(®) Trio kit can also be used to accurately estimate the optimal input DNA quantity for a direct STR amplification reaction. The correlation between the direct quantification results (Quantifiler(®) Trio kit) and the direct STR results (GlobalFiler™ PCR amplification kit(*)) for low-level touch DNA samples indicates that direct quantification using the Quantifiler(®) Trio DNA quantification kit is more reliable than the Quantifiler(®) Duo DNA quantification kit for predicting the STR results of unprocessed touch DNA samples containing less than 10pg of DNA. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  12. Specific Profile of Tau Isoforms in Argyrophylic Grain Disease

    Directory of Open Access Journals (Sweden)

    Alberto Rábano

    2013-01-01

    Full Text Available Argyrophylic grain disease (AGD is a neurodegenerative condition that has been classified among the sporadic tauopathies. Entities in this group present intracellular aggregates of hyperphosphorylated tau, giving rise to characteristic neuronal and glial inclusions. In different tauopathies, the proportion of several tau isoforms present in the aggregates shows specific patterns. AGD has been tentatively classified in the 4R group (predominance of 4R tau isoforms together with progressive supranuclear palsy and corticobasal degeneration. Pick's disease is included in the 3R group (predominance of 3R isoforms, whereas tau pathology of Alzheimer's disease represents and intermediate group (3 or 4 repeats [3R plus 4R, respectively] isoforms. In this work, we have analyzed tau present in aggregates isolated from brain samples of patients with argyrophylic grain disease. Our results indicate that the main tau isoform present in aggregates obtained from patients with AGD is a hyperphosphorylated isoform containing exons 2 and 10 but lacking exon 3.

  13. Tumorigenic properties of alternative osteopontin isoforms in mesothelioma

    Energy Technology Data Exchange (ETDEWEB)

    Ivanov, Sergey V., E-mail: Sergey.Ivanov@med.nyu.edu [Thoracic Surgery Laboratory, Cardiothoracic Surgery Department, NYU Langone Medical Center, 462 First Ave., Bellevue Hospital, Room 15N20, NY 10016 (United States); Ivanova, Alla V.; Goparaju, Chandra M.V.; Chen, Yuanbin; Beck, Amanda; Pass, Harvey I. [Thoracic Surgery Laboratory, Cardiothoracic Surgery Department, NYU Langone Medical Center, 462 First Ave., Bellevue Hospital, Room 15N20, NY 10016 (United States)

    2009-05-08

    Osteopontin (SPP1) is an inflammatory cytokine that we previously characterized as a diagnostic marker in patients with asbestos-induced malignant mesothelioma (MM). While SPP1 shows both pro- and anti-tumorigenic biological effects, little is known about the molecular basis of these activities. In this study, we demonstrate that while healthy pleura possesses all three differentially spliced SPP1 isoforms (A-C), in clinical MM specimens isoform A is markedly up-regulated and predominant. To provide a clue to possible functions of the SPP1 isoforms we next performed their functional evaluation via transient expression in MM cell lines. As a result, we report that isoforms A-C demonstrate different activities in cell proliferation, wound closure, and invasion assays. These findings suggest different functions for SPP1 isoforms and underline pro-tumorigenic properties of isoforms A and B.

  14. Oxygenation properties and isoform diversity of snake hemoglobins

    DEFF Research Database (Denmark)

    Storz, Jay F.; Natarajan, Chandrasekhar; Moriyama, Hideaki

    2015-01-01

    Available data suggest that snake hemoglobins (Hbs) are characterized by a combination of unusual structural and functional properties relative to the Hbs of other amniote vertebrates, including oxygenation-linked tetramer- dimer dissociation. However, standardized comparative data are lacking fo...... isoform of the South American rattlesnake is homologous to the minor HbD of other amniotes and, contrary to the pattern of Hb isoform differentiation in birds and turtles, exhibits a lower O2 affinity than the HbA isoform....

  15. Quantification in single photon emission computed tomography (SPECT)

    International Nuclear Information System (INIS)

    Buvat, Irene

    2005-01-01

    The objective of this lecture is to understand the possibilities and limitations of the quantitative analysis of single photon emission computed tomography (SPECT) images. It is also to identify the conditions to be fulfilled to obtain reliable quantitative measurements from images. Content: 1 - Introduction: Quantification in emission tomography - definition and challenges; quantification biasing phenomena; 2 - quantification in SPECT, problems and correction methods: Attenuation, scattering, un-stationary spatial resolution, partial volume effect, movement, tomographic reconstruction, calibration; 3 - Synthesis: actual quantification accuracy; 4 - Beyond the activity concentration measurement

  16. Real-time PCR quantification of human complement C4A and C4B genes

    Directory of Open Access Journals (Sweden)

    Fust George

    2006-01-01

    Full Text Available Abstract Background The fourth component of human complement (C4, an essential factor of the innate immunity, is represented as two isoforms (C4A and C4B in the genome. Although these genes differ only in 5 nucleotides, the encoded C4A and C4B proteins are functionally different. Based on phenotypic determination, unbalanced production of C4A and C4B is associated with several diseases, such as systemic lupus erythematosus, type 1 diabetes, several autoimmune diseases, moreover with higher morbidity and mortality of myocardial infarction and increased susceptibility for bacterial infections. Despite of this major clinical relevance, only low throughput, time and labor intensive methods have been used so far for the quantification of C4A and C4B genes. Results A novel quantitative real-time PCR (qPCR technique was developed for rapid and accurate quantification of the C4A and C4B genes applying a duplex, TaqMan based methodology. The reliable, single-step analysis provides the determination of the copy number of the C4A and C4B genes applying a wide range of DNA template concentration (0.3–300 ng genomic DNA. The developed qPCR was applied to determine C4A and C4B gene dosages in a healthy Hungarian population (N = 118. The obtained data were compared to the results of an earlier study of the same population. Moreover a set of 33 samples were analyzed by two independent methods. No significant difference was observed between the gene dosages determined by the employed techniques demonstrating the reliability of the novel qPCR methodology. A Microsoft Excel worksheet and a DOS executable are also provided for simple and automated evaluation of the measured data. Conclusion This report describes a novel real-time PCR method for single-step quantification of C4A and C4B genes. The developed technique could facilitate studies investigating disease association of different C4 isotypes.

  17. Safety and reliability assessment

    International Nuclear Information System (INIS)

    1979-01-01

    This report contains the papers delivered at the course on safety and reliability assessment held at the CSIR Conference Centre, Scientia, Pretoria. The following topics were discussed: safety standards; licensing; biological effects of radiation; what is a PWR; safety principles in the design of a nuclear reactor; radio-release analysis; quality assurance; the staffing, organisation and training for a nuclear power plant project; event trees, fault trees and probability; Automatic Protective Systems; sources of failure-rate data; interpretation of failure data; synthesis and reliability; quantification of human error in man-machine systems; dispersion of noxious substances through the atmosphere; criticality aspects of enrichment and recovery plants; and risk and hazard analysis. Extensive examples are given as well as case studies

  18. Quantification of the reliability of personnel actions from the evaluation of actual German operational experience. Final report; Quantifizierung der Zuverlaessigkeit von Personalhandlungen durch Auswertung der aktuellen deutschen Betriebserfahrung. Abschlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Preischl, W.; Fassmann, W.

    2013-07-15

    The results and their uncertainty bounds of PSA studies are considerably impacted by the assessment of human reliability. But the amount of available, generic data is not sufficient to evaluate all human actions considered in a modern PSA study adequately. Further the data are not sufficiently validated and rely as well as the proposed uncertainty bounds on expert judgement. This research project as well as the preceding project /GRS 10/ validated data recommended by the German PSA Guidelines and enlarged the amount of available data. The findings may contribute to an update of the German PSA Guidelines. In a first step of the project information about reportable events in German nuclear power plants with observed human errors (event reports, expert statements, technical documents, interviews and plant walk downs with subject matter experts from the plants) were analysed. The investigation resulted in 67 samples describing personal activities, performance conditions, the number of observed errors and the number of action performance. In a second step a new methodology was developed and applied in a pilot plant. The objective was to identify undoubtedly error free safety relevant actions, their performance conditions, and frequency as well as to prove and demonstrate that probabilistic data can be derived from that operational experience (OE). The application in the pilot plant resulted in 18 ''error free'' samples characterizing human reliability. All available samples were evaluated by use of the method of Bayes. That commonly accepted methodology was applied in order to derive probabilistic data based on samples taken from operational experience. A thorough analysis of the obtained results shows that both data sources (OE reportable events, OE with undoubtedly error free action performance) provide data with comparable quality and validity. At the end of the research project the following products are available. - Methods to select samples

  19. Software reliability

    CERN Document Server

    Bendell, A

    1986-01-01

    Software Reliability reviews some fundamental issues of software reliability as well as the techniques, models, and metrics used to predict the reliability of software. Topics covered include fault avoidance, fault removal, and fault tolerance, along with statistical methods for the objective assessment of predictive accuracy. Development cost models and life-cycle cost models are also discussed. This book is divided into eight sections and begins with a chapter on adaptive modeling used to predict software reliability, followed by a discussion on failure rate in software reliability growth mo

  20. Reliability Calculations

    DEFF Research Database (Denmark)

    Petersen, Kurt Erling

    1986-01-01

    Risk and reliability analysis is increasingly being used in evaluations of plant safety and plant reliability. The analysis can be performed either during the design process or during the operation time, with the purpose to improve the safety or the reliability. Due to plant complexity and safety...... and availability requirements, sophisticated tools, which are flexible and efficient, are needed. Such tools have been developed in the last 20 years and they have to be continuously refined to meet the growing requirements. Two different areas of application were analysed. In structural reliability probabilistic...... approaches have been introduced in some cases for the calculation of the reliability of structures or components. A new computer program has been developed based upon numerical integration in several variables. In systems reliability Monte Carlo simulation programs are used especially in analysis of very...

  1. The Role of Akt Isoforms in Colorectal Cancer

    Science.gov (United States)

    2015-09-01

    AD_________________ Award Number: W81XWH-13-1-0198 TITLE: The Role of Akt Isoforms in Colorectal Cancer PRINCIPAL INVESTIGATOR: Jatin Roper...CONTRACT NUMBER The Role of Akt Isoforms in Colorectal Cancer 5b. GRANT NUMBER W81XWH-13-1-0198 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER...substantially reduces colorectal tumorigenesis in our genetically engineered mouse model. We also successfully ablated novel downstream targets of Akt in our

  2. Characterisation of Cdkl5 transcript isoforms in rat.

    Science.gov (United States)

    Hector, Ralph D; Dando, Owen; Ritakari, Tuula E; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

    2017-03-01

    CDKL5 deficiency is a severe neurological disorder caused by mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5). The predominant human CDKL5 brain isoform is a 9.7kb transcript comprised of 18 exons with a large 6.6kb 3'-untranslated region (UTR). Mammalian models of CDKL5 disorder are currently limited to mouse, and little is known about Cdkl5 in other organisms used to model neurodevelopmental disorders, such as rat. In this study we characterise, both bioinformatically and experimentally, the rat Cdkl5 gene structure and its associated transcript isoforms. New exonic regions, splice sites and UTRs are described, confirming the presence of four distinct transcript isoforms. The predominant isoform in the brain, which we name rCdkl5_1, is orthologous to the human hCDKL5_1 and mouse mCdkl5_1 isoforms and is the most highly expressed isoform across all brain regions tested. This updated gene model of Cdkl5 in rat provides a framework for studies into its protein products and provides a reference for the development of molecular therapies for testing in rat models of CDKL5 disorder. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Isoform-selective regulation of glycogen phosphorylase by energy deprivation and phosphorylation in astrocytes

    DEFF Research Database (Denmark)

    Müller, Margit S; Pedersen, Sofie E; Walls, Anne B

    2015-01-01

    understood. In the present study, we used siRNA-mediated differential knockdown of the two isoforms of GP expressed in astrocytes, muscle isoform (GPMM), and brain isoform (GPBB), to analyze isoform-specific regulatory characteristics in a cellular setting. Subsequently, we tested the response of each...

  4. Structural diversity and evolution of the N-terminal isoform-specific region of ecdysone receptor-A and -B1 isoforms in insects

    Directory of Open Access Journals (Sweden)

    Kubo Takeo

    2010-02-01

    Full Text Available Abstract Background The ecdysone receptor (EcR regulates various cellular responses to ecdysteroids during insect development. Insects have multiple EcR isoforms with different N-terminal A/B domains that contain the isoform-specific activation function (AF-1 region. Although distinct physiologic functions of the EcR isoforms have been characterized in higher holometabolous insects, they remain unclear in basal direct-developing insects, in which only A isoform has been identified. To examine the structural basis of the EcR isoform-specific AF-1 regions, we performed a comprehensive structural comparison of the isoform-specific region of the EcR-A and -B1 isoforms in insects. Results The EcR isoforms were newly identified in 51 species of insects and non-insect arthropods, including direct-developing ametabolous and hemimetabolous insects. The comprehensive structural comparison revealed that the isoform-specific region of each EcR isoform contained evolutionally conserved microdomain structures and insect subgroup-specific structural modifications. The A isoform-specific region generally contained four conserved microdomains, including the SUMOylation motif and the nuclear localization signal, whereas the B1 isoform-specific region contained three conserved microdomains, including an acidic activator domain-like motif. In addition, the EcR-B1 isoform of holometabolous insects had a novel microdomain at the N-terminal end. Conclusions Given that the nuclear receptor AF-1 is involved in cofactor recruitment and transcriptional regulation, the microdomain structures identified in the isoform-specific A/B domains might function as signature motifs and/or as targets for cofactor proteins that play essential roles in the EcR isoform-specific AF-1 regions. Moreover, the novel microdomain in the isoform-specific region of the holometabolous insect EcR-B1 isoform suggests that the holometabolous insect EcR-B1 acquired additional transcriptional

  5. Differential Signature of the Centrosomal MARK4 Isoforms in Glioma

    Directory of Open Access Journals (Sweden)

    Ivana Magnani

    2011-01-01

    Full Text Available Background: MAP/microtubule affinity-regulating kinase 4 (MARK4 is a serine-threonine kinase expressed in two spliced isoforms, MARK4L and MARK4S, of which MARK4L is a candidate for a role in neoplastic transformation. Methods: We performed mutation analysis to identify sequence alterations possibly affecting MARK4 expression. We then investigated the MARK4L and MARK4S expression profile in 21 glioma cell lines and 36 tissues of different malignancy grades, glioblastoma-derived cancer stem cells (GBM CSCs and mouse neural stem cells (NSCs by real-time PCR, immunoblotting and immunohistochemistry. We also analyzed the sub-cellular localisation of MARK4 isoforms in glioma and normal cell lines by immunofluorescence. Results: Mutation analysis rules out sequence variations as the cause of the altered MARK4 expression in glioma. Expression profiling confirms that MARK4L is the predominant isoform, whereas MARK4S levels are significantly decreased in comparison and show an inverse correlation with tumour grade. A high MARK4L/MARK4S ratio also characterizes undifferentiated cells, such as GBM CSCs and NSCs. Accordingly, only MARK4L is expressed in brain neurogenic regions. Moreover, while both MARK4 isoforms are localised to the centrosome and midbody in glioma and normal cells, the L isoform exhibits an additional nucleolar localisation in tumour cells. Conclusions: The observed switch towards MARK4L suggests that the balance between the MARK4 isoforms is carefully guarded during neural differentiation but may be subverted in gliomagenesis. Moreover, the MARK4L nucleolar localisation in tumour cells features this MARK4 isoform as a nucleolus-associated tumour marker.

  6. Expression of various sarcomeric tropomyosin isoforms in equine striated muscles

    Directory of Open Access Journals (Sweden)

    Syamalima Dube

    2017-06-01

    Full Text Available In order to better understand the training and athletic activity of horses, we must have complete understanding of the isoform diversity of various myofibrillar protein genes like tropomyosin. Tropomyosin (TPM, a coiled-coil dimeric protein, is a component of thin filament in striated muscles. In mammals, four TPM genes (TPM1, TPM2, TPM3, and TPM4 generate a multitude of TPM isoforms via alternate splicing and/or using different promoters. Unfortunately, our knowledge of TPM isoform diversity in the horse is very limited. Hence, we undertook a comprehensive exploratory study of various TPM isoforms from horse heart and skeletal muscle. We have cloned and sequenced two sarcomeric isoforms of the TPM1 gene called TPM1α and TPM1κ, one sarcomeric isoform of the TPM2 and one of the TPM3 gene, TPM2α and TPM3α respectively. By qRT-PCR using both relative expression and copy number, we have shown that TPM1α expression compared to TPM1κ is very high in heart. On the other hand, the expression of TPM1α is higher in skeletal muscle compared to heart. Further, the expression of TPM2α and TPM3α are higher in skeletal muscle compared to heart. Using western blot analyses with CH1 monoclonal antibody we have shown the high expression levels of sarcomeric TPM proteins in cardiac and skeletal muscle. Due to the paucity of isoform specific antibodies we cannot specifically detect the expression of TPM1κ in horse striated muscle. To the best of our knowledge this is the very first report on the characterization of sarcmeric TPMs in horse striated muscle.

  7. Distinct functional interactions between actin isoforms and nonsarcomeric myosins.

    Directory of Open Access Journals (Sweden)

    Mirco Müller

    Full Text Available Despite their near sequence identity, actin isoforms cannot completely replace each other in vivo and show marked differences in their tissue-specific and subcellular localization. Little is known about isoform-specific differences in their interactions with myosin motors and other actin-binding proteins. Mammalian cytoplasmic β- and γ-actin interact with nonsarcomeric conventional myosins such as the members of the nonmuscle myosin-2 family and myosin-7A. These interactions support a wide range of cellular processes including cytokinesis, maintenance of cell polarity, cell adhesion, migration, and mechano-electrical transduction. To elucidate differences in the ability of isoactins to bind and stimulate the enzymatic activity of individual myosin isoforms, we characterized the interactions of human skeletal muscle α-actin, cytoplasmic β-actin, and cytoplasmic γ-actin with human myosin-7A and nonmuscle myosins-2A, -2B and -2C1. In the case of nonmuscle myosins-2A and -2B, the interaction with either cytoplasmic actin isoform results in 4-fold greater stimulation of myosin ATPase activity than was observed in the presence of α-skeletal muscle actin. Nonmuscle myosin-2C1 is most potently activated by β-actin and myosin-7A by γ-actin. Our results indicate that β- and γ-actin isoforms contribute to the modulation of nonmuscle myosin-2 and myosin-7A activity and thereby to the spatial and temporal regulation of cytoskeletal dynamics. FRET-based analyses show efficient copolymerization abilities for the actin isoforms in vitro. Experiments with hybrid actin filaments show that the extent of actomyosin coupling efficiency can be regulated by the isoform composition of actin filaments.

  8. FRANX. Application for analysis and quantification of the APS fire

    International Nuclear Information System (INIS)

    Snchez, A.; Osorio, F.; Ontoso, N.

    2014-01-01

    The FRANX application has been developed by EPRI within the Risk and Reliability User Group in order to facilitate the process of quantification and updating APS Fire (also covers floods and earthquakes). By applying fire scenarios are quantified in the central integrating the tasks performed during the APS fire. This paper describes the main features of the program to allow quantification of an APS Fire. (Author)

  9. Reliability Engineering

    CERN Document Server

    Lazzaroni, Massimo

    2012-01-01

    This book gives a practical guide for designers and users in Information and Communication Technology context. In particular, in the first Section, the definition of the fundamental terms according to the international standards are given. Then, some theoretical concepts and reliability models are presented in Chapters 2 and 3: the aim is to evaluate performance for components and systems and reliability growth. Chapter 4, by introducing the laboratory tests, puts in evidence the reliability concept from the experimental point of view. In ICT context, the failure rate for a given system can be

  10. Reliability training

    Science.gov (United States)

    Lalli, Vincent R. (Editor); Malec, Henry A. (Editor); Dillard, Richard B.; Wong, Kam L.; Barber, Frank J.; Barina, Frank J.

    1992-01-01

    Discussed here is failure physics, the study of how products, hardware, software, and systems fail and what can be done about it. The intent is to impart useful information, to extend the limits of production capability, and to assist in achieving low cost reliable products. A review of reliability for the years 1940 to 2000 is given. Next, a review of mathematics is given as well as a description of what elements contribute to product failures. Basic reliability theory and the disciplines that allow us to control and eliminate failures are elucidated.

  11. Reliability calculations

    International Nuclear Information System (INIS)

    Petersen, K.E.

    1986-03-01

    Risk and reliability analysis is increasingly being used in evaluations of plant safety and plant reliability. The analysis can be performed either during the design process or during the operation time, with the purpose to improve the safety or the reliability. Due to plant complexity and safety and availability requirements, sophisticated tools, which are flexible and efficient, are needed. Such tools have been developed in the last 20 years and they have to be continuously refined to meet the growing requirements. Two different areas of application were analysed. In structural reliability probabilistic approaches have been introduced in some cases for the calculation of the reliability of structures or components. A new computer program has been developed based upon numerical integration in several variables. In systems reliability Monte Carlo simulation programs are used especially in analysis of very complex systems. In order to increase the applicability of the programs variance reduction techniques can be applied to speed up the calculation process. Variance reduction techniques have been studied and procedures for implementation of importance sampling are suggested. (author)

  12. Quantification of local mobilities

    DEFF Research Database (Denmark)

    Zhang, Y. B.

    2018-01-01

    A new method for quantification of mobilities of local recrystallization boundary segments is presented. The quantification is based on microstructures characterized using electron microscopy and on determination of migration velocities and driving forces for local boundary segments. Pure aluminium...... is investigated and the results show that even for a single recrystallization boundary, different boundary segments migrate differently, and the differences can be understood based on variations in mobilities and local deformed microstructures. The present work has important implications for understanding...

  13. Systems reliability/structural reliability

    International Nuclear Information System (INIS)

    Green, A.E.

    1980-01-01

    The question of reliability technology using quantified techniques is considered for systems and structures. Systems reliability analysis has progressed to a viable and proven methodology whereas this has yet to be fully achieved for large scale structures. Structural loading variants over the half-time of the plant are considered to be more difficult to analyse than for systems, even though a relatively crude model may be a necessary starting point. Various reliability characteristics and environmental conditions are considered which enter this problem. The rare event situation is briefly mentioned together with aspects of proof testing and normal and upset loading conditions. (orig.)

  14. Molecular cloning and pharmacology of functionally distinct isoforms of the human histamine H(3) receptor

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Goodman, M W; Burstein, E S

    2002-01-01

    The pharmacology of histamine H(3) receptors suggests the presence of distinct receptor isoforms or subtypes. We herein describe multiple, functionally distinct, alternatively spliced isoforms of the human H(3) receptor. Combinatorial splicing at three different sites creates at least six distinct...... receptor isoforms, of which isoforms 1, 2, and 4, encode functional proteins. Detailed pharmacology on isoforms 1 (unspliced receptor), and 2 (which has an 80 amino acid deletion within the third intracellular loop of the protein) revealed that both isoforms displayed robust responses to a series of known...... revealed a rank order of potency at both isoforms of clobenpropit>iodophenpropit>thioperamide, and these drugs are fivefold less potent at isoform 2 than isoform 1. To further explore the pharmacology of H(3) receptor function, we screened 150 clinically relevant neuropsychiatric drugs for H(3) receptor...

  15. Proteogenomic Analysis Identifies a Novel Human SHANK3 Isoform

    Directory of Open Access Journals (Sweden)

    Fahad Benthani

    2015-05-01

    Full Text Available Mutations of the SHANK3 gene have been associated with autism spectrum disorder. Individuals harboring different SHANK3 mutations display considerable heterogeneity in their cognitive impairment, likely due to the high SHANK3 transcriptional diversity. In this study, we report a novel interaction between the Mutated in colorectal cancer (MCC protein and a newly identified SHANK3 protein isoform in human colon cancer cells and mouse brain tissue. Hence, our proteogenomic analysis identifies a new human long isoform of the key synaptic protein SHANK3 that was not predicted by the human reference genome. Taken together, our findings describe a potential new role for MCC in neurons, a new human SHANK3 long isoform and, importantly, highlight the use of proteomic data towards the re-annotation of GC-rich genomic regions.

  16. Oxygenation properties and isoform diversity of snake hemoglobins.

    Science.gov (United States)

    Storz, Jay F; Natarajan, Chandrasekhar; Moriyama, Hideaki; Hoffmann, Federico G; Wang, Tobias; Fago, Angela; Malte, Hans; Overgaard, Johannes; Weber, Roy E

    2015-11-01

    Available data suggest that snake hemoglobins (Hbs) are characterized by a combination of unusual structural and functional properties relative to the Hbs of other amniote vertebrates, including oxygenation-linked tetramer-dimer dissociation. However, standardized comparative data are lacking for snake Hbs, and the Hb isoform composition of snake red blood cells has not been systematically characterized. Here we present the results of an integrated analysis of snake Hbs and the underlying α- and β-type globin genes to characterize 1) Hb isoform composition of definitive erythrocytes, and 2) the oxygenation properties of isolated isoforms as well as composite hemolysates. We used species from three families as subjects for experimental studies of Hb function: South American rattlesnake, Crotalus durissus (Viperidae); Indian python, Python molurus (Pythonidae); and yellow-bellied sea snake, Pelamis platura (Elapidae). We analyzed allosteric properties of snake Hbs in terms of the Monod-Wyman-Changeux model and Adair four-step thermodynamic model. Hbs from each of the three species exhibited high intrinsic O2 affinities, low cooperativities, small Bohr factors in the absence of phosphates, and high sensitivities to ATP. Oxygenation properties of the snake Hbs could be explained entirely by allosteric transitions in the quaternary structure of intact tetramers, suggesting that ligation-dependent dissociation of Hb tetramers into αβ-dimers is not a universal feature of snake Hbs. Surprisingly, the major Hb isoform of the South American rattlesnake is homologous to the minor HbD of other amniotes and, contrary to the pattern of Hb isoform differentiation in birds and turtles, exhibits a lower O2 affinity than the HbA isoform. Copyright © 2015 the American Physiological Society.

  17. Identification and characterization of novel NuMA isoforms

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Jin, E-mail: petersdu2112@hotmail.com [Key Laboratory for Cell Proliferation and Regulation of the Ministry of Education, Beijing Normal University, Beijing (China); Xu, Zhe [Department of Clinical Laboratory Diagnosis, Beijing Tiantan Hospital, Capital Medical University, Beijing (China); Core Laboratory for Clinical Medical Research, Beijing Tiantan Hospital, Capital Medical University, Beijing (China); He, Dacheng [Key Laboratory for Cell Proliferation and Regulation of the Ministry of Education, Beijing Normal University, Beijing (China); Lu, Guanting, E-mail: guantlv@126.com [Beijing DnaLead Science and Technology Co., LTD, Beijing (China)

    2014-11-21

    Highlights: • Seven NuMA isoforms generated by alternative splicing were categorized into 3 groups: long, middle and short. • Both exons 15 and 16 in long NuMA were “hotspot” for alternative splicing. • Lower expression of short NuMA was observed in cancer cells compared with nonneoplastic controls. • Distinct localization pattern of short isoforms indicated different function from that of long and middle NuMA. - Abstract: The large nuclear mitotic apparatus (NuMA) has been investigated for over 30 years with functions related to the formation and maintenance of mitotic spindle poles during mitosis. However, the existence and functions of NuMA isoforms generated by alternative splicing remains unclear. In the present work, we show that at least seven NuMA isoforms (categorized into long, middle and short groups) generated by alternative splicing from a common NuMA mRNA precursor were discovered in HeLa cells and these isoforms differ mainly at the carboxyl terminus and the coiled-coil domains. Two “hotspot” exons with molecular mass of 3366-nt and 42-nt tend to be spliced during alternative splicing in long and middle groups. Furthermore, full-length coding sequences of long and middle NuMA obtained by using fusion PCR were constructed into GFP-tagged vector to illustrate their cellular localization. Long NuMA mainly localized in the nucleus with absence from nucleoli during interphase and translocated to the spindle poles in mitosis. Middle NuMA displayed the similar cell cycle-dependent distribution pattern as long NuMA. However, expression of NuMA short isoforms revealed a distinct subcellular localization. Short NuMA were present in the cytosol during the whole cycle, without colocalization with mitotic apparatus. These results have allowed us tentatively to explore a new research direction for NuMA’s various functions.

  18. Cytochrome P450 isoform selectivity in human hepatic theobromine metabolism

    Science.gov (United States)

    Gates, Simon; Miners, John O

    1999-01-01

    Aims The plasma clearance of theobromine (TB; 3,7-dimethylxanthine) is known to be induced in cigarette smokers. To determine whether TB may serve as a model substrate for cytochrome P450 (CYP) 1A2, or possibly other isoforms, studies were undertaken to identify the individual human liver microsomal CYP isoforms responsible for the conversion of TB to its primary metabolites. Methods The kinetics of formation of the primary TB metabolites 3-methylxanthine (3-MX), 7-methylxanthine (7-MX) and 3,7-dimethyluric acid (3,7-DMU) by human liver microsomes were characterized using a specific hplc procedure. Effects of CYP isoform-selective xenobiotic inhibitor/substrate probes on each pathway were determined and confirmatory studies with recombinant enzymes were performed to define the contribution of individual isoforms to 3-MX, 7-MX and 3,7-DMU formation. Results The CYP1A2 inhibitor furafylline variably inhibited (0–65%) 7-MX formation, but had no effect on other pathways. Diethyldithiocarbamate and 4-nitrophenol, probes for CYP2E1, inhibited the formation of 3-MX, 7-MX and 3,7-DMU by ≈55–60%, 35–55% and 85%, respectively. Consistent with the microsomal studies, recombinant CYP1A2 and CYP2E1 exhibited similar apparent Km values for 7-MX formation and CYP2E1 was further shown to have the capacity to convert TB to both 3-MX and 3,7-DMU. Conclusions Given the contribution of multiple isoforms to 3-MX and 7-MX formation and the negligible formation of 3,7-DMU in vivo, TB is of little value as a CYP isoform-selective substrate in humans. PMID:10215755

  19. Functions of PDE3 Isoforms in Cardiac Muscle

    Science.gov (United States)

    Movsesian, Matthew; Ahmad, Faiyaz

    2018-01-01

    Isoforms in the PDE3 family of cyclic nucleotide phosphodiesterases have important roles in cyclic nucleotide-mediated signalling in cardiac myocytes. These enzymes are targeted by inhibitors used to increase contractility in patients with heart failure, with a combination of beneficial and adverse effects on clinical outcomes. This review covers relevant aspects of the molecular biology of the isoforms that have been identified in cardiac myocytes; the roles of these enzymes in modulating cAMP-mediated signalling and the processes mediated thereby; and the potential for targeting these enzymes to improve the profile of clinical responses. PMID:29415428

  20. Modulation of neuronal differentiation by CD40 isoforms

    International Nuclear Information System (INIS)

    Hou Huayu; Obregon, Demian; Lou, Deyan; Ehrhart, Jared; Fernandez, Frank; Silver, Archie; Tan Jun

    2008-01-01

    Neuron differentiation is a complex process involving various cell-cell interactions, and multiple signaling pathways. We showed previously that CD40 is expressed and functional on mouse and human neurons. In neurons, ligation of CD40 protects against serum withdrawal-induced injury and plays a role in survival and differentiation. CD40 deficient mice display neuron dysfunction, aberrant neuron morphologic changes, and associated gross brain abnormalities. Previous studies by Tone and colleagues suggested that five isoforms of CD40 exist with two predominant isoforms expressed in humans: signal-transducible CD40 type I and a C-terminal truncated, non-signal-transducible CD40 type II. We hypothesized that differential expression of CD40 isoform type I and type II in neurons may modulate neuron differentiation. Results show that adult wild-type, and CD40 -/- deficient mice predominantly express CD40 type I and II isoforms. Whereas adult wild-type mice express mostly CD40 type I in cerebral tissues at relatively high levels, in age and gender-matched CD40 -/- mice CD40 type I expression was almost completely absent; suggesting a predominance of the non-signal-transducible CD40 type II isoform. Younger, 1 day old wild-type mice displayed less CD40 type I, and more CD40 type II, as well as, greater expression of soluble CD40 (CD40L/CD40 signal inhibitor), compared with 1 month old mice. Neuron-like N2a cells express CD40 type I and type II isoforms while in an undifferentiated state, however once induced to differentiate, CD40 type I predominates. Further, differentiated N2a cells treated with CD40 ligand express high levels of neuron specific nuclear protein (NeuN); an effect reduced by anti-CD40 type I siRNA, but not by control (non-targeting) siRNA. Altogether these data suggest that CD40 isoforms may act in a temporal fashion to modulate neuron differentiation during brain development. Thus, modulation of neuronal CD40 isoforms and CD40 signaling may represent

  1. Human reliability

    International Nuclear Information System (INIS)

    Bubb, H.

    1992-01-01

    This book resulted from the activity of Task Force 4.2 - 'Human Reliability'. This group was established on February 27th, 1986, at the plenary meeting of the Technical Reliability Committee of VDI, within the framework of the joint committee of VDI on industrial systems technology - GIS. It is composed of representatives of industry, representatives of research institutes, of technical control boards and universities, whose job it is to study how man fits into the technical side of the world of work and to optimize this interaction. In a total of 17 sessions, information from the part of ergonomy dealing with human reliability in using technical systems at work was exchanged, and different methods for its evaluation were examined and analyzed. The outcome of this work was systematized and compiled in this book. (orig.) [de

  2. Microelectronics Reliability

    Science.gov (United States)

    2017-01-17

    inverters  connected in a chain. ................................................. 5  Figure 3  Typical graph showing frequency versus square root of...developing an experimental  reliability estimating methodology that could both illuminate the  lifetime  reliability of advanced devices,  circuits and...or  FIT of the device. In other words an accurate estimate of the device  lifetime  was found and thus the  reliability  that  can  be  conveniently

  3. Fluorescent quantification of melanin.

    Science.gov (United States)

    Fernandes, Bruno; Matamá, Teresa; Guimarães, Diana; Gomes, Andreia; Cavaco-Paulo, Artur

    2016-11-01

    Melanin quantification is reportedly performed by absorption spectroscopy, commonly at 405 nm. Here, we propose the implementation of fluorescence spectroscopy for melanin assessment. In a typical in vitro assay to assess melanin production in response to an external stimulus, absorption spectroscopy clearly overvalues melanin content. This method is also incapable of distinguishing non-melanotic/amelanotic control cells from those that are actually capable of performing melanogenesis. Therefore, fluorescence spectroscopy is the best method for melanin quantification as it proved to be highly specific and accurate, detecting even small variations in the synthesis of melanin. This method can also be applied to the quantification of melanin in more complex biological matrices like zebrafish embryos and human hair. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. The landscape of isoform switches in human cancers

    DEFF Research Database (Denmark)

    Vitting-Seerup, Kristoffer; Sandelin, Albin Gustav

    2017-01-01

    highly predictive of patient survival independent of cancer types. Our data constitute an important resource for cancer researchers, available through interactive web tools. Moreover, our methods, available as an R package, enable systematic analysis of isoform switches from other RNA-seq datasets...

  5. Plectin isoforms as organizers of intermediate filament cytoarchitecture.

    Science.gov (United States)

    Wiche, Gerhard; Winter, Lilli

    2011-01-01

    Intermediate filaments (IFs) form cytoplamic and nuclear networks that provide cells with mechanical strength. Perturbation of this structural support causes cell and tissue fragility and accounts for a number of human genetic diseases. In recent years, important additional roles, nonmechanical in nature, were ascribed to IFs, including regulation of signaling pathways that control survival and growth of the cells, and vectorial processes such as protein targeting in polarized cellular settings. The cytolinker protein plectin anchors IF networks to junctional complexes, the nuclear envelope and cytoplasmic organelles and it mediates their cross talk with the actin and tubulin cytoskeleton. These functions empower plectin to wield significant influence over IF network cytoarchitecture. Moreover, the unusual diversity of plectin isoforms with different N termini and a common IF-binding (C-terminal) domain enables these isoforms to specifically associate with and thereby bridge IF networks to distinct cellular structures. Here we review the evidence for IF cytoarchitecture being controlled by specific plectin isoforms in different cell systems, including fibroblasts, endothelial cells, lens fibers, lymphocytes, myocytes, keratinocytes, neurons and astrocytes, and discuss what impact the absence of these isoforms has on IF cytoarchitecture-dependent cellular functions.

  6. Roles of the troponin isoforms during indirect flight muscle ...

    Indian Academy of Sciences (India)

    IFMs) undergo post-transcriptional and post-translational isoform changes during pupal to adult metamorphosis to meet the high energy and mechanical demands of flight. Using a newly generated Gal4 strain (UH3-Gal4) which is expressed ...

  7. Distinct Functions of Endophilin Isoforms in Synaptic Vesicle Endocytosis

    Directory of Open Access Journals (Sweden)

    Jifeng Zhang

    2015-01-01

    Full Text Available Endophilin isoforms perform distinct characteristics in their interactions with N-type Ca2+ channels and dynamin. However, precise functional differences for the endophilin isoforms on synaptic vesicle (SV endocytosis remain unknown. By coupling RNA interference and electrophysiological recording techniques in cultured rat hippocampal neurons, we investigated the functional differences of three isoforms of endophilin in SV endocytosis. The results showed that the amplitude of normalized evoked excitatory postsynaptic currents in endophilin1 knockdown neurons decreased significantly for both single train and multiple train stimulations. Similar results were found using endophilin2 knockdown neurons, whereas endophilin3 siRNA exhibited no change compared with control neurons. Endophilin1 and endophilin2 affected SV endocytosis, but the effect of endophilin1 and endophilin2 double knockdown was not different from that of either knockdown alone. This result suggested that endophilin1 and endophilin2 functioned together but not independently during SV endocytosis. Taken together, our results indicate that SV endocytosis is sustained by endophilin1 and endophilin2 isoforms, but not by endophilin3, in primary cultured hippocampal neurons.

  8. Molecular characters and expression analysis of a new isoform of ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-10-20

    Oct 20, 2008 ... isoform of the myocyte enhancer factor 2 gene from the silkworm, Bombyx mori. Qing-zhi Ling1, 2, ... BMEF2B mRNA content in the brain was measured using the combined method of quantitative RT-PCR and Southern ... specific cofactors to control gene expression in pheno- typically different muscles.

  9. Isoforms of transferrin in psoriasis patients abusing alcohol

    NARCIS (Netherlands)

    P. Hoefkens (Peter); E.M. Higgins; R.J. Ward (Roberta); H.G. van Eijk (Henk)

    1997-01-01

    textabstractThe different isoforms of transferrin have been quantified by isoelectric focusing in the sera of psoriasis patients with and without a history of abusing alcohol. In both male and female psoriasis subjects abusing alcohol, there were significant increases in the

  10. Redefining reliability

    International Nuclear Information System (INIS)

    Paulson, S.L.

    1995-01-01

    Want to buy some reliability? The question would have been unthinkable in some markets served by the natural gas business even a few years ago, but in the new gas marketplace, industrial, commercial and even some residential customers have the opportunity to choose from among an array of options about the kind of natural gas service they need--and are willing to pay for. The complexities of this brave new world of restructuring and competition have sent the industry scrambling to find ways to educate and inform its customers about the increased responsibility they will have in determining the level of gas reliability they choose. This article discusses the new options and the new responsibilities of customers, the needed for continuous education, and MidAmerican Energy Company's experiment in direct marketing of natural gas

  11. RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome

    Directory of Open Access Journals (Sweden)

    Dewey Colin N

    2011-08-01

    Full Text Available Abstract Background RNA-Seq is revolutionizing the way transcript abundances are measured. A key challenge in transcript quantification from RNA-Seq data is the handling of reads that map to multiple genes or isoforms. This issue is particularly important for quantification with de novo transcriptome assemblies in the absence of sequenced genomes, as it is difficult to determine which transcripts are isoforms of the same gene. A second significant issue is the design of RNA-Seq experiments, in terms of the number of reads, read length, and whether reads come from one or both ends of cDNA fragments. Results We present RSEM, an user-friendly software package for quantifying gene and isoform abundances from single-end or paired-end RNA-Seq data. RSEM outputs abundance estimates, 95% credibility intervals, and visualization files and can also simulate RNA-Seq data. In contrast to other existing tools, the software does not require a reference genome. Thus, in combination with a de novo transcriptome assembler, RSEM enables accurate transcript quantification for species without sequenced genomes. On simulated and real data sets, RSEM has superior or comparable performance to quantification methods that rely on a reference genome. Taking advantage of RSEM's ability to effectively use ambiguously-mapping reads, we show that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads. On the other hand, estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired-end reads, depending on the number of possible splice forms for each gene. Conclusions RSEM is an accurate and user-friendly software tool for quantifying transcript abundances from RNA-Seq data. As it does not rely on the existence of a reference genome, it is particularly useful for quantification with de novo transcriptome assemblies. In addition, RSEM has enabled valuable guidance for cost

  12. Uncertainty Quantification in Alchemical Free Energy Methods.

    Science.gov (United States)

    Bhati, Agastya P; Wan, Shunzhou; Hu, Yuan; Sherborne, Brad; Coveney, Peter V

    2018-05-02

    Alchemical free energy methods have gained much importance recently from several reports of improved ligand-protein binding affinity predictions based on their implementation using molecular dynamics simulations. A large number of variants of such methods implementing different accelerated sampling techniques and free energy estimators are available, each claimed to be better than the others in its own way. However, the key features of reproducibility and quantification of associated uncertainties in such methods have barely been discussed. Here, we apply a systematic protocol for uncertainty quantification to a number of popular alchemical free energy methods, covering both absolute and relative free energy predictions. We show that a reliable measure of error estimation is provided by ensemble simulation-an ensemble of independent MD simulations-which applies irrespective of the free energy method. The need to use ensemble methods is fundamental and holds regardless of the duration of time of the molecular dynamics simulations performed.

  13. N Termini of apPDE4 Isoforms Are Responsible for Targeting the Isoforms to Different Cellular Membranes

    Science.gov (United States)

    Jang, Deok-Jin; Park, Soo-Won; Lee, Jin-A; Lee, Changhoon; Chae, Yeon-Su; Park, Hyungju; Kim, Min-Jeong; Choi, Sun-Lim; Lee, Nuribalhae; Kim, Hyoung; Kaang, Bong-Kiun

    2010-01-01

    Phosphodiesterases (PDEs) are known to play a key role in the compartmentalization of cAMP signaling; however, the molecular mechanisms underlying intracellular localization of different PDE isoforms are not understood. In this study, we have found that each of the supershort, short, and long forms of apPDE4 showed distinct localization in the…

  14. Characterization of ß-Galactosidase Isoforms from Bacillus circulans and Their Contribution to GOS Production

    NARCIS (Netherlands)

    Warmerdam, A.; Paudel, E.; Wanqing, J.; Boom, R.M.; Janssen, A.E.M.

    2013-01-01

    A ß-galactosidase preparation from Bacillus circulans consists of four isoforms called ß-gal-A, ß-gal-B, ß-gal-C, and ß-gal-D. These isoforms differ in lactose hydrolysis and galacto-oligosaccharide (GOS) synthesis at low substrate concentrations. For this reason, using a selection of the isoforms

  15. Disease quantification in dermatology

    DEFF Research Database (Denmark)

    Greve, Tanja Maria; Kamp, Søren; Jemec, Gregor B E

    2013-01-01

    Accurate documentation of disease severity is a prerequisite for clinical research and the practice of evidence-based medicine. The quantification of skin diseases such as psoriasis currently relies heavily on clinical scores. Although these clinical scoring methods are well established and very ...

  16. One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

    International Nuclear Information System (INIS)

    Bianchi, Cristina; Torsello, Barbara; Di Stefano, Vitalba; Zipeto, Maria A.; Facchetti, Rita; Bombelli, Silvia; Perego, Roberto A.

    2013-01-01

    The non-receptor tyrosine kinase Abelson related gene (Arg/Abl2) regulates cell migration and morphogenesis by modulating the cytoskeleton. Arg promotes actin-based cell protrusions and spreading, and inhibits cell migration by attenuating stress fiber formation and contractility via activation of the RhoA inhibitor, p190RhoGAP, and by regulating focal adhesion dynamics also via CrkII phosphorylation. Eight full-length Arg isoforms with different N- and C-termini are endogenously expressed in human cells. In this paper, the eight Arg isoforms, subcloned in the pFLAG-CMV2 vector, were transfected in COS-7 cells in order to study their subcellular distribution and role in cell morphology, migration and cytoskeletal modulation. The transfected 1BSCTS Arg isoform has a nuclear distribution and phosphorylates CrkII in the nucleus, whilst the other isoforms are detected in the cytoplasm. The 1BLCTL, 1BSCTL, 1ASCTS isoforms were able to significantly decrease stress fibers, induce cell shrinkage and filopodia-like protrusions with a significant increase in p190RhoGAP phosphorylation. In contrast, 1ALCTL, 1ALCTS, 1ASCTL and 1BLCTS isoforms do not significantly decrease stress fibers and induce the formation of retraction tail-like protrusions. The 1BLCTL and 1ALCTL isoforms have different effects on cell migration and focal adhesions. All these data may open new perspectives to study the mechanisms of cell invasiveness. -Highlights: • Each of the eight Arg isoforms was transfected in COS-7 cells. • Only the 1BSCTS Arg isoform has a nuclear distribution in transfected cells. • The cytoplasmic isoforms and F-actin colocalize cortically and in cell protrusions. • Arg isoforms differently phosphorylate p190RhoGAP and CrkII. • Arg isoforms differently modulate stress fibers, cell protrusions and motility

  17. One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

    Energy Technology Data Exchange (ETDEWEB)

    Bianchi, Cristina; Torsello, Barbara; Di Stefano, Vitalba; Zipeto, Maria A.; Facchetti, Rita; Bombelli, Silvia; Perego, Roberto A., E-mail: roberto.perego@unimib.it

    2013-08-01

    The non-receptor tyrosine kinase Abelson related gene (Arg/Abl2) regulates cell migration and morphogenesis by modulating the cytoskeleton. Arg promotes actin-based cell protrusions and spreading, and inhibits cell migration by attenuating stress fiber formation and contractility via activation of the RhoA inhibitor, p190RhoGAP, and by regulating focal adhesion dynamics also via CrkII phosphorylation. Eight full-length Arg isoforms with different N- and C-termini are endogenously expressed in human cells. In this paper, the eight Arg isoforms, subcloned in the pFLAG-CMV2 vector, were transfected in COS-7 cells in order to study their subcellular distribution and role in cell morphology, migration and cytoskeletal modulation. The transfected 1BSCTS Arg isoform has a nuclear distribution and phosphorylates CrkII in the nucleus, whilst the other isoforms are detected in the cytoplasm. The 1BLCTL, 1BSCTL, 1ASCTS isoforms were able to significantly decrease stress fibers, induce cell shrinkage and filopodia-like protrusions with a significant increase in p190RhoGAP phosphorylation. In contrast, 1ALCTL, 1ALCTS, 1ASCTL and 1BLCTS isoforms do not significantly decrease stress fibers and induce the formation of retraction tail-like protrusions. The 1BLCTL and 1ALCTL isoforms have different effects on cell migration and focal adhesions. All these data may open new perspectives to study the mechanisms of cell invasiveness. -Highlights: • Each of the eight Arg isoforms was transfected in COS-7 cells. • Only the 1BSCTS Arg isoform has a nuclear distribution in transfected cells. • The cytoplasmic isoforms and F-actin colocalize cortically and in cell protrusions. • Arg isoforms differently phosphorylate p190RhoGAP and CrkII. • Arg isoforms differently modulate stress fibers, cell protrusions and motility.

  18. Human factors reliability benchmark exercise: a review

    International Nuclear Information System (INIS)

    Humphreys, P.

    1990-01-01

    The Human Factors Reliability Benchmark Exercise has addressed the issues of identification, analysis, representation and quantification of Human Error in order to identify the strengths and weaknesses of available techniques. Using a German PWR nuclear powerplant as the basis for the studies, fifteen teams undertook evaluations of a routine functional Test and Maintenance procedure plus an analysis of human actions during an operational transient. The techniques employed by the teams are discussed and reviewed on a comparative basis. The qualitative assessments performed by each team compare well, but at the quantification stage there is much less agreement. (author)

  19. Uncertainty quantification in nanomechanical measurements using the atomic force microscope

    Science.gov (United States)

    Ryan Wagner; Robert Moon; Jon Pratt; Gordon Shaw; Arvind Raman

    2011-01-01

    Quantifying uncertainty in measured properties of nanomaterials is a prerequisite for the manufacture of reliable nanoengineered materials and products. Yet, rigorous uncertainty quantification (UQ) is rarely applied for material property measurements with the atomic force microscope (AFM), a widely used instrument that can measure properties at nanometer scale...

  20. freeQuant: A Mass Spectrometry Label-Free Quantification Software Tool for Complex Proteome Analysis.

    Science.gov (United States)

    Deng, Ning; Li, Zhenye; Pan, Chao; Duan, Huilong

    2015-01-01

    Study of complex proteome brings forward higher request for the quantification method using mass spectrometry technology. In this paper, we present a mass spectrometry label-free quantification tool for complex proteomes, called freeQuant, which integrated quantification with functional analysis effectively. freeQuant consists of two well-integrated modules: label-free quantification and functional analysis with biomedical knowledge. freeQuant supports label-free quantitative analysis which makes full use of tandem mass spectrometry (MS/MS) spectral count, protein sequence length, shared peptides, and ion intensity. It adopts spectral count for quantitative analysis and builds a new method for shared peptides to accurately evaluate abundance of isoforms. For proteins with low abundance, MS/MS total ion count coupled with spectral count is included to ensure accurate protein quantification. Furthermore, freeQuant supports the large-scale functional annotations for complex proteomes. Mitochondrial proteomes from the mouse heart, the mouse liver, and the human heart were used to evaluate the usability and performance of freeQuant. The evaluation showed that the quantitative algorithms implemented in freeQuant can improve accuracy of quantification with better dynamic range.

  1. Image cytometry: nuclear and chromosomal DNA quantification.

    Science.gov (United States)

    Carvalho, Carlos Roberto; Clarindo, Wellington Ronildo; Abreu, Isabella Santiago

    2011-01-01

    Image cytometry (ICM) associates microscopy, digital image and software technologies, and has been particularly useful in spatial and densitometric cytological analyses, such as DNA ploidy and DNA content measurements. Basically, ICM integrates methodologies of optical microscopy calibration, standard density filters, digital CCD camera, and image analysis softwares for quantitative applications. Apart from all system calibration and setup, cytological protocols must provide good slide preparations for efficient and reliable ICM analysis. In this chapter, procedures for ICM applications employed in our laboratory are described. Protocols shown here for human DNA ploidy determination and quantification of nuclear and chromosomal DNA content in plants could be used as described, or adapted for other studies.

  2. Isoform expression in the multiple soluble malate dehydrogenase of Hoplias malabaricus (Erythrinidae, Characiformes

    Directory of Open Access Journals (Sweden)

    M. R. Aquino-Silva

    Full Text Available Kinetic properties and thermal stabilities of Hoplias malabaricus liver and skeletal muscle unfractionated malate dehydrogenase (MDH, EC 1.1.1.37 and its isolated isoforms were analyzed to further study the possible sMDH-A* locus duplication evolved from a recent tandem duplication. Both A (A1 and A2 and B isoforms had similar optima pH (7.5-8.0. While Hoplias A isoform could not be characterized as thermostable, B could as thermolabile. A isoforms differed from B isoform in having higher Km values for oxaloacetate. The possibly duplicated A2 isoform showed higher substrate affinity than the A1. Hoplias duplicated A isoforms may influence the direction of carbon flow between glycolisis and gluconeogenesis.

  3. Isoform expression in the multiple soluble malate dehydrogenase of Hoplias malabaricus (Erythrinidae, Characiformes

    Directory of Open Access Journals (Sweden)

    Aquino-Silva M. R.

    2003-01-01

    Full Text Available Kinetic properties and thermal stabilities of Hoplias malabaricus liver and skeletal muscle unfractionated malate dehydrogenase (MDH, EC 1.1.1.37 and its isolated isoforms were analyzed to further study the possible sMDH-A* locus duplication evolved from a recent tandem duplication. Both A (A1 and A2 and B isoforms had similar optima pH (7.5-8.0. While Hoplias A isoform could not be characterized as thermostable, B could as thermolabile. A isoforms differed from B isoform in having higher Km values for oxaloacetate. The possibly duplicated A2 isoform showed higher substrate affinity than the A1. Hoplias duplicated A isoforms may influence the direction of carbon flow between glycolisis and gluconeogenesis.

  4. Nubbin isoform antagonism governs Drosophila intestinal immune homeostasis.

    Directory of Open Access Journals (Sweden)

    Bo G Lindberg

    2018-03-01

    Full Text Available Gut immunity is regulated by intricate and dynamic mechanisms to ensure homeostasis despite a constantly changing microbial environment. Several regulatory factors have been described to participate in feedback responses to prevent aberrant immune activity. Little is, however, known about how transcriptional programs are directly tuned to efficiently adapt host gut tissues to the current microbiome. Here we show that the POU/Oct gene nubbin (nub encodes two transcription factor isoforms, Nub-PB and Nub-PD, which antagonistically regulate immune gene expression in Drosophila. Global transcriptional profiling of adult flies overexpressing Nub-PB in immunocompetent tissues revealed that this form is a strong transcriptional activator of a large set of immune genes. Further genetic analyses showed that Nub-PB is sufficient to drive expression both independently and in conjunction with nuclear factor kappa B (NF-κB, JNK and JAK/STAT pathways. Similar overexpression of Nub-PD did, conversely, repress expression of the same targets. Strikingly, isoform co-overexpression normalized immune gene transcription, suggesting antagonistic activities. RNAi-mediated knockdown of individual nub transcripts in enterocytes confirmed antagonistic regulation by the two isoforms and that both are necessary for normal immune gene transcription in the midgut. Furthermore, enterocyte-specific Nub-PB expression levels had a strong impact on gut bacterial load as well as host lifespan. Overexpression of Nub-PB enhanced bacterial clearance of ingested Erwinia carotovora carotovora 15. Nevertheless, flies quickly succumbed to the infection, suggesting a deleterious immune response. In line with this, prolonged overexpression promoted a proinflammatory signature in the gut with induction of JNK and JAK/STAT pathways, increased apoptosis and stem cell proliferation. These findings highlight a novel regulatory mechanism of host-microbe interactions mediated by antagonistic

  5. Nitric oxide synthase isoforms in spontaneous and salt hypertension

    Czech Academy of Sciences Publication Activity Database

    Hojná, Silvie; Kuneš, Jaroslav; Zicha, Josef

    2007-01-01

    Roč. 25, Suppl. 2 (2007), S 338-S 338 ISSN 0263-6352. [European Meeting on Hypertension /17./. 15.06.2007-19.06.2007, Milan] R&D Projects: GA MŠk(CZ) 1M0510 Institutional research plan: CEZ:AV0Z50110509 Keywords : nitric oxide synthase isoforms * spontaneous and salt hypertension Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery

  6. Expression of two isoforms of CD44 in human endometrium.

    Science.gov (United States)

    Behzad, F; Seif, M W; Campbell, S; Aplin, J D

    1994-10-01

    The distribution of the cell-surface adhesion glycoprotein CD44 in human endometrium was examined by immunofluorescence using six monoclonal antibodies to epitopes common to all forms of the molecule, and by reverse transcription-polymerase chain reaction (RT-PCR). Immunoreactivity was observed throughout the menstrual cycle in stroma, vessels, glandular, and luminal epithelium. Variations in staining intensity were observed, especially in the epithelial compartment. CD44 was also expressed strongly by decidualized stromal cells of first-trimester pregnancy. No systematic variation of immunoreactivity was observed with stages of the normal cycle, but a fraction (25%) of the specimens lacked reactivity in the epithelium. To determine the molecular size of the epithelial isoform, an immunoprecipitation technique was developed using surface-radioiodinated, detergent-extracted glands. This indicated the presence at the cell surface of a single dominant CD44E species with an approximate molecular mass of 130 kDa. RT-PCR was used to investigate the isoforms present in whole endometrial tissue, isolated gland fragments, and Ishikawa endometrial carcinoma cells. Complementary DNA produced from total endometrial mRNA was PCR-amplified across the splice junction between exons 5 and 15. Transcripts corresponding to the hyaluronate receptor CD44H as well as a larger isoform were identified. CD44H was absent, or very scarce, in cDNA from purified gland epithelium. In contrast, Ishikawa cells expressed this form abundantly. The glands and Ishikawa cells also expressed CD44E containing sequences encoded by exons 12, 13, and 14. These data demonstrate the presence of CD44 in human endometrium and decidua, and show that different isoforms of CD44 are associated with tissue compartments in which different functional roles can be anticipated.

  7. Apolipoprotein (A) Isoform Distribution and Plasma Lipoprotein (a ...

    African Journals Online (AJOL)

    Plasma lipoprotein (a) Concentrations and apo(a) isoforms were determined in 101 healthy Nigerian subjects (M=63), F=38; age range 17-68 years), and coronary heart disease (CHD) patients (M=19, F=17, age range 30-79 years). Median Lp(a) level was 24.4 mg/di in the CHD patients and 22.1 mg/di in the controls.

  8. Prediction of software operational reliability using testing environment factors

    International Nuclear Information System (INIS)

    Jung, Hoan Sung; Seong, Poong Hyun

    1995-01-01

    For many years, many researches have focused on the quantification of software reliability and there are many models developed to quantify software reliability. Most software reliability models estimate the reliability with the failure data collected during the test assuming that the test environments well represent the operation profile. The experiences show that the operational reliability is higher than the test reliability User's interest is on the operational reliability rather than on the test reliability, however. With the assumption that the difference in reliability results from the change of environment, testing environment factors comprising the aging factor and the coverage factor are defined in this study to predict the ultimate operational reliability with the failure data. It is by incorporating test environments applied beyond the operational profile into testing environment factors. The application results are close to the actual data

  9. RON kinase isoforms demonstrate variable cell motility in normal cells.

    Science.gov (United States)

    Greenbaum, Alissa; Rajput, Ashwani; Wan, Guanghua

    2016-09-01

    Aberrant RON (Recepteur d'Origine Nantais) tyrosine kinase activation causes the epithelial cell to evade normal growth pathways, resulting in unregulated cell proliferation, increased cell motility and decreased apoptosis. Wildtype (wt) RON has been shown to play a role in metastasis of epithelial malignancies. It presents an important potential therapeutic target for colorectal, breast, gastric and pancreatic cancer. Little is known about functional differences amongst RON isoforms RON155, RON160 and RON165. The purpose of this study was to determine the effect of various RON kinase isoforms on cell motility. Cell lines with stable expression of wtRON were generated by inserting the coding region of RON in pTagRFP (tagged red fluorescence protein plasmid). The expression constructs of RON variants (RON155, RON160 and RON165) were generated by creating a mutagenesis-based wtRON-pTag RFP plasmid and stably transfected into HEK 293 cells. The wound closure scratch assay was used to investigate the effect on cell migratory capacity of wild type RON and its variants. RON transfected cells demonstrated increased cell motility compared to HEK293 control cells. RON165 cell motility was significantly increased compared to RON160 (mean percentage of wound covered 37.37% vs. 32.40%; p = 0.03). RON tyrosine kinase isoforms have variable cell motility. This may reflect a difference in the behavior of malignant epithelial cells and their capacity for metastasis.

  10. Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

    Science.gov (United States)

    Hector, Ralph D; Dando, Owen; Landsberger, Nicoletta; Kilstrup-Nielsen, Charlotte; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

    2016-01-01

    Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5) cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR), which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

  11. Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

    Directory of Open Access Journals (Sweden)

    Ralph D Hector

    Full Text Available Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5 cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR, which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

  12. [Characterization of a malic enzyme isoform V from Mucor circinelloides].

    Science.gov (United States)

    Zhang, Yingtong; Chen, Haiqin; Song, Yuanda; Zhang, Hao; Chen, Yongquan; Chen, Wei

    2016-02-04

    We aimed at characterizing a malic enzyme isoform V from Mucor circinelloides. me1 gene encoding malic enzyme isoform V was amplified and cloned into expression vector pET28a. High-purity recombinant protein BLME1 was obtained by affinity chromatography using. Ni-NTA column and characterized subsequently. The optimum conditions were pH at 8.0 and temperature at 33 degrees C. Under optimum conditions, BLME1 activity achieved 92.8 U/mg. The K(m) for L-malate and NADP+ were 0.74960 ± 0.06120 mmol/L and 0.22070 ± 0.01810 mmol/L, the V(max) for L-malate and NADP+ were 72.820 ± 1.077 U/mg and 86.110 ± 1.665 U/mg, respectively. In addition, ions played important roles in BLME1 activity; several ions such as Mn2+, Mg2+, Co2+, Ni2+ could activate BLME1, whereas Ca2+, Cu2+ could be used as inhibitors. Additionally, the metabolic intermediates such as oxaloacetic acid and α-ketoglutaric acid inhibited the activity of BLME1, whereas succinic acid activated it. A malic enzyme isoform V from Mucor circinelloides was characterized, providing the references for further studies on this enzyme.

  13. Differential expression of syndecan isoforms during mouse incisor amelogenesis.

    Science.gov (United States)

    Muto, Taro; Miyoshi, Keiko; Munesue, Seiichi; Nakada, Hiroshi; Okayama, Minoru; Matsuo, Takashi; Noma, Takafumi

    2007-08-01

    Syndecans are transmembranous heparan sulfate proteoglycans (HSPGs) with covalently attached glycosaminoglycan side-chains located on the cell surface. The mammalian syndecan family is composed of four types of syndecans (syndecan-1 to -4). Syndecans interact with the intracellular cytoskeleton through the cytoplasmic domains of their core proteins and membrane proteins, extracellular enzymes, growth factors, and matrix components, through their heparan-sulfate chains, to regulate developmental processes.Here, as a first step to assess the possible roles of syndecan proteins in amelogenesis, we examined the expression patterns of all syndecan isoforms in continuously growing mouse incisors, in which we can overview major differentiation stages of amelogenesis at a glance. Understanding the expression domain of each syndecan isoform during specific developmental stages seems useful for investigating their physiological roles in amelogenesis. Immunohistochemical analysis of syndecan core proteins in the lower incisors from postnatal day 1 mice revealed spatially and temporally specific expression patterns, with syndecan-1 expressed in undifferentiated epithelial and mesenchymal cells, and syndecan-2, -3, and -4 in more differentiated cells. These findings suggest that each syndecan isoform functions distinctly during the amelogenesis of the incisors of mice.

  14. An Introduction To Reliability

    International Nuclear Information System (INIS)

    Park, Kyoung Su

    1993-08-01

    This book introduces reliability with definition of reliability, requirement of reliability, system of life cycle and reliability, reliability and failure rate such as summary, reliability characteristic, chance failure, failure rate which changes over time, failure mode, replacement, reliability in engineering design, reliability test over assumption of failure rate, and drawing of reliability data, prediction of system reliability, conservation of system, failure such as summary and failure relay and analysis of system safety.

  15. Platelet amyloid precursor protein isoform expression in Alzheimer's disease: evidence for peripheral marker.

    Science.gov (United States)

    Vignini, A; Sartini, D; Morganti, S; Nanetti, L; Luzzi, S; Provinciali, L; Mazzanti, L; Emanuelli, M

    2011-01-01

    Alzheimer's disease (AD) is a chronic neurodegenerative disorder characterized by a progressive cognitive and memory decline. Among peripheral markers of AD, great interest has been focused on the amyloid precursor protein (APP). In this regard, platelets represent an important peripheral source of APP since it has been demonstrated that the three major isoforms, that are constituted of 770, 751 and 695 aa residues, are inserted in the membrane of resting platelets. APP 751 and APP 770 contain a Kunitz-type serine protease inhibitor domain (APP KPI) and APP 695 lacks this domain. To address this issue, we first examined the platelet APP isoform mRNAs prospectively as biomarker for the diagnosis of AD by means of real-time quantitative PCR, and then evaluated the correlation between APP mRNA expression levels and cognitive impairment of enrolled subjects. Differential gene expression measurements in the AD patient group (n=18) revealed a significant up-regulation of APP TOT (1.52-fold), APP KPI (1.32-fold), APP 770 (1.33-fold) and APP 751 (1.26-fold) compared to controls (n=22). Moreover, a statistically significant positive correlation was found between APP mRNA levels (TOT, KPI, 770 and 751) and cognitive impairment. Since AD definitive diagnosis still relies on pathological evaluation at autopsy, the present results are consistent with the hypothesis that platelet APP could be considered a potential reliable peripheral marker for studying AD and could contribute to define a signature for the presence of AD pathology.

  16. Prediction of safety critical software operational reliability from test reliability using testing environment factors

    International Nuclear Information System (INIS)

    Jung, Hoan Sung; Seong, Poong Hyun

    1999-01-01

    It has been a critical issue to predict the safety critical software reliability in nuclear engineering area. For many years, many researches have focused on the quantification of software reliability and there have been many models developed to quantify software reliability. Most software reliability models estimate the reliability with the failure data collected during the test assuming that the test environments well represent the operation profile. User's interest is however on the operational reliability rather than on the test reliability. The experiences show that the operational reliability is higher than the test reliability. With the assumption that the difference in reliability results from the change of environment, from testing to operation, testing environment factors comprising the aging factor and the coverage factor are developed in this paper and used to predict the ultimate operational reliability with the failure data in testing phase. It is by incorporating test environments applied beyond the operational profile into testing environment factors. The application results show that the proposed method can estimate the operational reliability accurately. (Author). 14 refs., 1 tab., 1 fig

  17. Reliability Based Management of Marine Fouling

    DEFF Research Database (Denmark)

    Faber, Michael Havbro; Hansen, Peter Friis

    1999-01-01

    The present paper describes the results of a recent study on the application of methods from structural reliability to optimise management of marine fouling on jacket type structures.In particular the study addresses effects on the structural response by assessment and quantification of uncertain......The present paper describes the results of a recent study on the application of methods from structural reliability to optimise management of marine fouling on jacket type structures.In particular the study addresses effects on the structural response by assessment and quantification...... of uncertainties of a set of parameters. These are the seasonal variation of marine fouling parameters, the wave loading (taking into account the seasonal variation in sea-state statistics), and the effects of spatial variations and seasonal effects of marine fouling parameters. Comparison of design values...

  18. Characterisation of Cdkl5transcript isoforms in rat

    OpenAIRE

    Hector, Ralph D.; Dando, Owen; Ritakari, Tuula E.; Kind, Peter C.; Bailey, Mark E.S.; Cobb, Stuart R.

    2017-01-01

    CDKL5 deficiency is a severe neurological disorder caused by mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5). The predominant human CDKL5 brain isoform is a 9.7kb transcript comprised of 18 exons with a large 6.6kb 3'-untranslated region (UTR). Mammalian models of CDKL5 disorder are currently limited to mouse, and little is known about Cdkl5 in other organisms used to model neurodevelopmental disorders, such as rat. In this study we characterise, both bioinformatically a...

  19. Overexpression of EMMPRIN Isoform 2 Is Associated with Head and Neck Cancer Metastasis

    OpenAIRE

    Huang, Zhiquan; Tan, Ning; Guo, Weijie; Wang, Lili; Li, Haigang; Zhang, Tianyu; Liu, Xiaojia; Xu, Qin; Li, Jinsong; Guo, Zhongmin

    2014-01-01

    Extracellular matrix metalloproteinase inducer (EMMPRIN), a plasma membrane protein of the immunoglobulin (Ig) superfamily, has been reported to promote cancer cell invasion and metastasis in several human malignancies. However, the roles of the different EMMPRIN isoforms and their associated mechanisms in head and neck cancer progression remain unknown. Using quantitative real-time PCR, we found that EMMPRIN isoform 2 (EMMPRIN-2) was the only isoform that was overexpressed in both head and n...

  20. Identification of signals that facilitate isoform specific nucleolar localization of myosin IC

    Energy Technology Data Exchange (ETDEWEB)

    Schwab, Ryan S.; Ihnatovych, Ivanna; Yunus, Sharifah Z.S.A.; Domaradzki, Tera [Department of Physiology and Biophysics, University at Buffalo—State University of New York, Buffalo, NY (United States); Hofmann, Wilma A., E-mail: whofmann@buffalo.edu [Department of Physiology and Biophysics, University at Buffalo—State University of New York, Buffalo, NY (United States)

    2013-05-01

    Myosin IC is a single headed member of the myosin superfamily that localizes to the cytoplasm and the nucleus, where it is involved in transcription by RNA polymerases I and II, intranuclear transport, and nuclear export. In mammalian cells, three isoforms of myosin IC are expressed that differ only in the addition of short isoform-specific N-terminal peptides. Despite the high sequence homology, the isoforms show differences in cellular distribution, in localization to nuclear substructures, and in their interaction with nuclear proteins through yet unknown mechanisms. In this study, we used EGFP-fusion constructs that express truncated or mutated versions of myosin IC isoforms to detect regions that are involved in isoform-specific localization. We identified two nucleolar localization signals (NoLS). One NoLS is located in the myosin IC isoform B specific N-terminal peptide, the second NoLS is located upstream of the neck region within the head domain. We demonstrate that both NoLS are functional and necessary for nucleolar localization of specifically myosin IC isoform B. Our data provide a first mechanistic explanation for the observed functional differences between the myosin IC isoforms and are an important step toward our understanding of the underlying mechanisms that regulate the various and distinct functions of myosin IC isoforms. - Highlights: ► Two NoLS have been identified in the myosin IC isoform B sequence. ► Both NoLS are necessary for myosin IC isoform B specific nucleolar localization. ► First mechanistic explanation of functional differences between the isoforms.

  1. A highly sensitive method for quantification of iohexol

    DEFF Research Database (Denmark)

    Schulz, A.; Boeringer, F.; Swifka, J.

    2014-01-01

    -chromatography-electrospray-massspectrometry (LC-ESI-MS) approach using the multiple reaction monitoring mode for iohexol quantification. In order to test whether a significantly decreased amount of iohexol is sufficient for reliable quantification, a LC-ESI-MS approach was assessed. We analyzed the kinetics of iohexol in rats after application...... of different amounts of iohexol (15 mg to 150 1.tg per rat). Blood sampling was conducted at four time points, at 15, 30, 60, and 90 min, after iohexol injection. The analyte (iohexol) and the internal standard (iotha(amic acid) were separated from serum proteins using a centrifugal filtration device...... with a cut-off of 3 kDa. The chromatographic separation was achieved on an analytical Zorbax SB C18 column. The detection and quantification were performed on a high capacity trap mass spectrometer using positive ion ESI in the multiple reaction monitoring (MRM) mode. Furthermore, using real-time polymerase...

  2. Frontiers of reliability

    CERN Document Server

    Basu, Asit P; Basu, Sujit K

    1998-01-01

    This volume presents recent results in reliability theory by leading experts in the world. It will prove valuable for researchers, and users of reliability theory. It consists of refereed invited papers on a broad spectrum of topics in reliability. The subjects covered include Bayesian reliability, Bayesian reliability modeling, confounding in a series system, DF tests, Edgeworth approximation to reliability, estimation under random censoring, fault tree reduction for reliability, inference about changes in hazard rates, information theory and reliability, mixture experiment, mixture of Weibul

  3. NHS-A isoform of the NHS gene is a novel interactor of ZO-1.

    Science.gov (United States)

    Sharma, Shiwani; Koh, Katrina S Y; Collin, Caitlin; Dave, Alpana; McMellon, Amy; Sugiyama, Yuki; McAvoy, John W; Voss, Anne K; Gécz, Jozef; Craig, Jamie E

    2009-08-15

    Mutations in the NHS (Nance-Horan Syndrome) gene lead to severe congenital cataracts, dental defects and sometimes mental retardation. NHS encodes two protein isoforms, NHS-A and -1A that display cell-type dependent differential expression and localization. Here we demonstrate that of these two isoforms, the NHS-A isoform associates with the cell membrane in the presence of intercellular contacts and it immunoprecipitates with the tight junction protein ZO-1 in MDCK (Madin Darby Canine Kidney) epithelial cells and in neonatal rat lens. The NHS-1A isoform however is a cytoplasmic protein. Both Nhs isoforms are expressed during mouse development. Immunolabelling of developing mouse with the anti-NHS antibody that detects both isoforms revealed the protein in the developing head including the eye and brain. It was primarily expressed in epithelium including neural epithelium and certain vascular endothelium but only weakly expressed in mesenchymal cells. In the epithelium and vascular endothelium the protein associated with the cell membrane and co-localized with ZO-1, which indirectly indicates expression of the Nhs-A isoform in these structures. Membrane localization of the protein in the lens vesicle similarly supports Nhs-A expression. In conclusion, the NHS-A isoform of NHS is a novel interactor of ZO-1 and may have a role at tight junctions. This isoform is important in mammalian development especially of the organs in the head.

  4. Prostaglandin D Synthase Isoforms from Cerebrospinal Fluid Vary with Brain Pathology

    Directory of Open Access Journals (Sweden)

    Michael G. Harrington

    2006-01-01

    Full Text Available Glutathione independent prostaglandin D synthase (Swissprot P41222, PTGDS has been identified in human cerebrospinal fluid and some changes in PTGDS in relation to disease have been reported. However, little is known of the extent that PTGDS isoforms fluctuate across a large range of congenital and acquired diseases. The purpose of this study was to examine changes in PTGDS isoforms in such a population. Spinal fluid from 22 healthy study participants (normal controls with no classifiable neurological or psychiatric diagnosis was obtained and PTGDS isoforms were identified by specific immunostaining and mass spectrometry after denaturing 2D gel electrophoresis. The PTGDS isoforms in controls consisted of five charge isoforms that were always present and a small number of occasional, low abundance isoforms. A qualitative survey of 98 different people with a wide range of congenital and acquired diseases revealed striking changes. Loss of the control isoforms occurred in congenital malformations of the nervous system. Gain of additional isoforms occurred in some degenerative, most demyelinating and vasculitic diseases, as well as in Creutzfeldt-Jakob disease. A retrospective analysis of published data that quantified relative amounts of PTGDS in multiple sclerosis, schizophrenia and Parkinson’s disease compared to controls revealed significant dysregulation. It is concluded that qualitative and quantitative fluctuations of cerebrospinal fluid PTGDS isoforms reflect both major and subtle brain pathophysiology.

  5. Comparison of transferrin isoform analysis by capillary electrophoresis and HPLC for screening congenital disorders of glycosylation.

    Science.gov (United States)

    Dave, Mihika B; Dherai, Alpa J; Udani, Vrajesh P; Hegde, Anaita U; Desai, Neelu A; Ashavaid, Tester F

    2018-01-01

    Transferrin, a major glycoprotein has different isoforms depending on the number of sialic acid residues present on its oligosaccharide chain. Genetic variants of transferrin as well as the primary (CDG) & secondary glycosylation defects lead to an altered transferrin pattern. Isoform analysis methods are based on charge/mass variations. We aimed to compare the performance of commercially available capillary electrophoresis CDT kit for diagnosing congenital disorders of glycosylation with our in-house optimized HPLC method for transferrin isoform analysis. The isoform pattern of 30 healthy controls & 50 CDG-suspected patients was determined by CE using a Carbohydrate-Deficient Transferrin kit. The results were compared with in-house HPLC-based assay for transferrin isoforms. Transferrin isoform pattern for healthy individuals showed a predominant tetrasialo transferrin fraction followed by pentasialo, trisialo, and disialotransferrin. Two of 50 CDG-suspected patients showed the presence of asialylated isoforms. The results were comparable with isoform pattern obtained by HPLC. The commercial controls showed a <20% CV for each isoform. Bland Altman plot showed the difference plot to be within +1.96 with no systemic bias in the test results by HPLC & CE. The CE method is rapid, reproducible and comparable with HPLC and can be used for screening Glycosylation defects. © 2017 Wiley Periodicals, Inc.

  6. Bisphenol A differentially activates protein kinase C isoforms in murine placental tissue

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Wenjuan; Huang, Hui; Wang, Yanfei [Biochemistry Programme, School of Life Sciences, Faculty of Science, The Chinese University of Hong Kong, Shatin, N.T. (Hong Kong); Wong, Tsz Yan [Food and Nutritional Sciences Programme, School of Life Sciences, Faculty of Science, The Chinese University of Hong Kong, Shatin, N.T. (Hong Kong); Wang, C.C. [Department of Obstetrics and Gynecology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, N.T. (Hong Kong); Leung, Lai K., E-mail: laikleung@cuhk.edu.hk [Biochemistry Programme, School of Life Sciences, Faculty of Science, The Chinese University of Hong Kong, Shatin, N.T. (Hong Kong); Food and Nutritional Sciences Programme, School of Life Sciences, Faculty of Science, The Chinese University of Hong Kong, Shatin, N.T. (Hong Kong)

    2013-06-01

    Bisphenol A is utilized to make polycarbonate plastics and is an environmental pollutant. Recent research has indicated that it is an endocrine disruptor and may interfere with reproductive processes. Our lab has previously shown that bisphenol A could regulate corticotrophin releasing hormone and aromatase in cultured placental cells. In the present study, the effect of bisphenol A on these two genes in the placenta was investigated in mice. Pregnant ICR mice were gavaged with bisphenol A at 2, 20 and 200 mg/kg body weight/day from E13 to E16 and were euthanized at E17. Compared to the control mice, increased plasma estrogen and corticotrophin releasing hormone were observed in bisphenol A-treated mice. Messenger RNA quantification indicated that placental crh but not cyp19 was induced in mice treated with bisphenol A. Tracking the related signaling pathway, we found that protein kinase C ζ/λ and δ were activated in the placentas of bisphenol A-treated mice. As the gene promoter of crh contains CRE and the half site of ERE, either phospho-PKC or estrogen could stimulate the gene transactivation. These results indicate that bisphenol A might increase plasma concentrations of estradiol, testosterone, corticotrophin releasing hormone and placental phospho-PKC ζ/λ and δ in mice. Ultimately, the incidence of premature birth in these mice could increase. - Highlights: • The pollutant bisphenol A differentially activated PKC isoforms in the placenta. • CRE-binding activity in the nuclear protein of placenta was increased. • Bisphenol A induces CRH mRNA expression in mice.

  7. Assessment of orthologous splicing isoforms in human and mouse orthologous genes

    Directory of Open Access Journals (Sweden)

    Horner David S

    2010-10-01

    Full Text Available Abstract Background Recent discoveries have highlighted the fact that alternative splicing and alternative transcripts are the rule, rather than the exception, in metazoan genes. Since multiple transcript and protein variants expressed by the same gene are, by definition, structurally distinct and need not to be functionally equivalent, the concept of gene orthology should be extended to the transcript level in order to describe evolutionary relationships between structurally similar transcript variants. In other words, the identification of true orthology relationships between gene products now should progress beyond primary sequence and "splicing orthology", consisting in ancestrally shared exon-intron structures, is required to define orthologous isoforms at transcript level. Results As a starting step in this direction, in this work we performed a large scale human- mouse gene comparison with a twofold goal: first, to assess if and to which extent traditional gene annotations such as RefSeq capture genuine splicing orthology; second, to provide a more detailed annotation and quantification of true human-mouse orthologous transcripts defined as transcripts of orthologous genes exhibiting the same splicing patterns. Conclusions We observed an identical exon/intron structure for 32% of human and mouse orthologous genes. This figure increases to 87% using less stringent criteria for gene structure similarity, thus implying that for about 13% of the human RefSeq annotated genes (and about 25% of the corresponding transcripts we could not identify any mouse transcript showing sufficient similarity to be confidently assigned as a splicing ortholog. Our data suggest that current gene and transcript data may still be rather incomplete - with several splicing variants still unknown. The observation that alternative splicing produces large numbers of alternative transcripts and proteins, some of them conserved across species and others truly species

  8. Bisphenol A differentially activates protein kinase C isoforms in murine placental tissue

    International Nuclear Information System (INIS)

    Tan, Wenjuan; Huang, Hui; Wang, Yanfei; Wong, Tsz Yan; Wang, C.C.; Leung, Lai K.

    2013-01-01

    Bisphenol A is utilized to make polycarbonate plastics and is an environmental pollutant. Recent research has indicated that it is an endocrine disruptor and may interfere with reproductive processes. Our lab has previously shown that bisphenol A could regulate corticotrophin releasing hormone and aromatase in cultured placental cells. In the present study, the effect of bisphenol A on these two genes in the placenta was investigated in mice. Pregnant ICR mice were gavaged with bisphenol A at 2, 20 and 200 mg/kg body weight/day from E13 to E16 and were euthanized at E17. Compared to the control mice, increased plasma estrogen and corticotrophin releasing hormone were observed in bisphenol A-treated mice. Messenger RNA quantification indicated that placental crh but not cyp19 was induced in mice treated with bisphenol A. Tracking the related signaling pathway, we found that protein kinase C ζ/λ and δ were activated in the placentas of bisphenol A-treated mice. As the gene promoter of crh contains CRE and the half site of ERE, either phospho-PKC or estrogen could stimulate the gene transactivation. These results indicate that bisphenol A might increase plasma concentrations of estradiol, testosterone, corticotrophin releasing hormone and placental phospho-PKC ζ/λ and δ in mice. Ultimately, the incidence of premature birth in these mice could increase. - Highlights: • The pollutant bisphenol A differentially activated PKC isoforms in the placenta. • CRE-binding activity in the nuclear protein of placenta was increased. • Bisphenol A induces CRH mRNA expression in mice

  9. A Review of Metallothionein Isoforms and their Role in Pathophysiology

    Directory of Open Access Journals (Sweden)

    Senthil kumar M

    2011-05-01

    Full Text Available Abstract The Metallothionein (MT is a protein which has several interesting biological effects and has been demonstrated increase focus on the role of MT in various biological systems in the past three decades. The studies on the role of MT were limited with few areas like apoptosis and antioxidants in selected organs even fifty years after its discovery. Now acknowledge the exploration of various isoforms of MT such as MT-I, MT-II, MT-III and MT-IV and other isoforms in various biological systems. Strong evidence exists that MT modulates complex diseases and the immune system in the body but the primary function of MT still remains unknown. This review's main objective is to explore the capability to specifically manipulate MT levels in cells and in animals to provide answers regarding how MT could impact those complex disease scenarios. The experimental result mentioned in this review related among MT, zinc, cadmium, diabetic, heart disease, bone retardation, neuro toxicity, kidney dysfunction, cancer, and brain suggest novel method for exploration and contribute significantly to the growing scientist to research further in this field.

  10. Entropy-based model for miRNA isoform analysis.

    Directory of Open Access Journals (Sweden)

    Shengqin Wang

    Full Text Available MiRNAs have been widely studied due to their important post-transcriptional regulatory roles in gene expression. Many reports have demonstrated the evidence of miRNA isoform products (isomiRs in high-throughput small RNA sequencing data. However, the biological function involved in these molecules is still not well investigated. Here, we developed a Shannon entropy-based model to estimate isomiR expression profiles of high-throughput small RNA sequencing data extracted from miRBase webserver. By using the Kolmogorov-Smirnov statistical test (KS test, we demonstrated that the 5p and 3p miRNAs present more variants than the single arm miRNAs. We also found that the isomiR variant, except the 3' isomiR variant, is strongly correlated with Minimum Free Energy (MFE of pre-miRNA, suggesting the intrinsic feature of pre-miRNA should be one of the important factors for the miRNA regulation. The functional enrichment analysis showed that the miRNAs with high variation, particularly the 5' end variation, are enriched in a set of critical functions, supporting these molecules should not be randomly produced. Our results provide a probabilistic framework for miRNA isoforms analysis, and give functional insights into pre-miRNA processing.

  11. Human factors reliability benchmark exercise

    International Nuclear Information System (INIS)

    Poucet, A.

    1989-08-01

    The Joint Research Centre of the European Commission has organised a Human Factors Reliability Benchmark Exercise (HF-RBE) with the aim of assessing the state of the art in human reliability modelling and assessment. Fifteen teams from eleven countries, representing industry, utilities, licensing organisations and research institutes, participated in the HF-RBE. The HF-RBE was organised around two study cases: (1) analysis of routine functional Test and Maintenance (TPM) procedures: with the aim of assessing the probability of test induced failures, the probability of failures to remain unrevealed and the potential to initiate transients because of errors performed in the test; (2) analysis of human actions during an operational transient: with the aim of assessing the probability that the operators will correctly diagnose the malfunctions and take proper corrective action. This report summarises the contributions received from the participants and analyses these contributions on a comparative basis. The aim of this analysis was to compare the procedures, modelling techniques and quantification methods used, to obtain insight in the causes and magnitude of the variability observed in the results, to try to identify preferred human reliability assessment approaches and to get an understanding of the current state of the art in the field identifying the limitations that are still inherent to the different approaches

  12. Expression of phosphoinositide-specific phospholipase C isoforms in native endothelial cells.

    Science.gov (United States)

    Béziau, Delphine M; Toussaint, Fanny; Blanchette, Alexandre; Dayeh, Nour R; Charbel, Chimène; Tardif, Jean-Claude; Dupuis, Jocelyn; Ledoux, Jonathan

    2015-01-01

    Phospholipase C (PLC) comprises a superfamily of enzymes that play a key role in a wide array of intracellular signalling pathways, including protein kinase C and intracellular calcium. Thirteen different mammalian PLC isoforms have been identified and classified into 6 families (PLC-β, γ, δ, ε, ζ and η) based on their biochemical properties. Although the expression of PLC isoforms is tissue-specific, concomitant expression of different PLC has been reported, suggesting that PLC family is involved in multiple cellular functions. Despite their critical role, the PLC isoforms expressed in native endothelial cells (ECs) remains undetermined. A conventional PCR approach was initially used to elucidate the mRNA expression pattern of PLC isoforms in 3 distinct murine vascular beds: mesenteric (MA), pulmonary (PA) and middle cerebral arteries (MCA). mRNA encoding for most PLC isoforms was detected in MA, MCA and PA with the exception of η2 and β2 (only expressed in PA), δ4 (only expressed in MCA), η1 (expressed in all but MA) and ζ (not detected in any vascular beds tested). The endothelial-specific PLC expression was then sought in freshly isolated ECs. Interestingly, the PLC expression profile appears to differ across the investigated arterial beds. While mRNA for 8 of the 13 PLC isoforms was detected in ECs from MA, two additional PLC isoforms were detected in ECs from PA and MCA. Co-expression of multiple PLC isoforms in ECs suggests an elaborate network of signalling pathways: PLC isoforms may contribute to the complexity or diversity of signalling by their selective localization in cellular microdomains. However in situ immunofluorescence revealed a homogeneous distribution for all PLC isoforms probed (β3, γ2 and δ1) in intact endothelium. Although PLC isoforms play a crucial role in endothelial signal transduction, subcellular localization alone does not appear to be sufficient to determine the role of PLC in the signalling microdomains found in the

  13. Identification and characterization of novel smoothelin isoforms in vascular smooth muscle.

    Science.gov (United States)

    Krämer, J; Quensel, C; Meding, J; Cardoso, M C; Leonhardt, H

    2001-01-01

    Smoothelin is a cytoskeletal protein specifically expressed in differentiated smooth muscle cells and has been shown to colocalize with smooth muscle alpha actin. In addition to the small smoothelin isoform of 59 kD, we recently identified a large smoothelin isoform of 117 kD. The aim of this study was to identify and characterize novel smoothelin isoforms. The genomic structure and sequence of the smoothelin gene were determined by genomic PCR, RT-PCR and DNA sequencing. Comparison of the cDNA and genomic sequences shows that the small smoothelin isoform is generated by transcription initiation 10 kb downstream of the start site of the large isoform. In addition to the known smoothelin cDNA (c1 isoform) we identified two novel cDNA variants (c2 and c3 isoform) that are generated by alternative splicing within a region, which shows similarity to the spectrin family of F-actin cross-linking proteins. Visceral organs express the c1 form, while the c2 form prevails in well-vascularized tissue as analyzed by RT-PCR. We then generated specific antibodies against the major smoothelin isoforms and could show by Western blotting and immunohistochemistry that the large isoform is specifically expressed in vascular smooth muscle cells, while the small isoform is abundant in visceral smooth muscle. These results strongly suggest that the smoothelin gene contains a vascular and a visceral smooth muscle promoter. The cell-type-specific expression of smoothelin isoforms that are associated with actin filaments may play a role in the modulation of the contractile properties of different smooth muscle cell types. Copyright 2001 S. Karger AG, Basel

  14. Expression of phosphoinositide-specific phospholipase C isoforms in native endothelial cells.

    Directory of Open Access Journals (Sweden)

    Delphine M Béziau

    Full Text Available Phospholipase C (PLC comprises a superfamily of enzymes that play a key role in a wide array of intracellular signalling pathways, including protein kinase C and intracellular calcium. Thirteen different mammalian PLC isoforms have been identified and classified into 6 families (PLC-β, γ, δ, ε, ζ and η based on their biochemical properties. Although the expression of PLC isoforms is tissue-specific, concomitant expression of different PLC has been reported, suggesting that PLC family is involved in multiple cellular functions. Despite their critical role, the PLC isoforms expressed in native endothelial cells (ECs remains undetermined. A conventional PCR approach was initially used to elucidate the mRNA expression pattern of PLC isoforms in 3 distinct murine vascular beds: mesenteric (MA, pulmonary (PA and middle cerebral arteries (MCA. mRNA encoding for most PLC isoforms was detected in MA, MCA and PA with the exception of η2 and β2 (only expressed in PA, δ4 (only expressed in MCA, η1 (expressed in all but MA and ζ (not detected in any vascular beds tested. The endothelial-specific PLC expression was then sought in freshly isolated ECs. Interestingly, the PLC expression profile appears to differ across the investigated arterial beds. While mRNA for 8 of the 13 PLC isoforms was detected in ECs from MA, two additional PLC isoforms were detected in ECs from PA and MCA. Co-expression of multiple PLC isoforms in ECs suggests an elaborate network of signalling pathways: PLC isoforms may contribute to the complexity or diversity of signalling by their selective localization in cellular microdomains. However in situ immunofluorescence revealed a homogeneous distribution for all PLC isoforms probed (β3, γ2 and δ1 in intact endothelium. Although PLC isoforms play a crucial role in endothelial signal transduction, subcellular localization alone does not appear to be sufficient to determine the role of PLC in the signalling microdomains found

  15. System Reliability Engineering

    International Nuclear Information System (INIS)

    Lim, Tae Jin

    2005-02-01

    This book tells of reliability engineering, which includes quality and reliability, reliability data, importance of reliability engineering, reliability and measure, the poisson process like goodness of fit test and the poisson arrival model, reliability estimation like exponential distribution, reliability of systems, availability, preventive maintenance such as replacement policies, minimal repair policy, shock models, spares, group maintenance and periodic inspection, analysis of common cause failure, and analysis model of repair effect.

  16. Detection and quantification of beef and pork materials in meat products by duplex droplet digital PCR

    OpenAIRE

    Cai, Yicun; He, Yuping; Lv, Rong; Chen, Hongchao; Wang, Qiang; Pan, Liangwen

    2017-01-01

    Meat products often consist of meat from multiple animal species, and inaccurate food product adulteration and mislabeling can negatively affect consumers. Therefore, a cost-effective and reliable method for identification and quantification of animal species in meat products is required. In this study, we developed a duplex droplet digital PCR (dddPCR) detection and quantification system to simultaneously identify and quantify the source of meat in samples containing a mixture of beef (Bos t...

  17. Molecular Pharmacology of VEGF-A Isoforms: Binding and Signalling at VEGFR2.

    Science.gov (United States)

    Peach, Chloe J; Mignone, Viviane W; Arruda, Maria Augusta; Alcobia, Diana C; Hill, Stephen J; Kilpatrick, Laura E; Woolard, Jeanette

    2018-04-23

    Vascular endothelial growth factor-A (VEGF-A) is a key mediator of angiogenesis, signalling via the class IV tyrosine kinase receptor family of VEGF Receptors (VEGFRs). Although VEGF-A ligands bind to both VEGFR1 and VEGFR2, they primarily signal via VEGFR2 leading to endothelial cell proliferation, survival, migration and vascular permeability. Distinct VEGF-A isoforms result from alternative splicing of the Vegfa gene at exon 8, resulting in VEGF xxx a or VEGF xxx b isoforms. Alternative splicing events at exons 5⁻7, in addition to recently identified posttranslational read-through events, produce VEGF-A isoforms that differ in their bioavailability and interaction with the co-receptor Neuropilin-1. This review explores the molecular pharmacology of VEGF-A isoforms at VEGFR2 in respect to ligand binding and downstream signalling. To understand how VEGF-A isoforms have distinct signalling despite similar affinities for VEGFR2, this review re-evaluates the typical classification of these isoforms relative to the prototypical, “pro-angiogenic” VEGF 165 a. We also examine the molecular mechanisms underpinning the regulation of VEGF-A isoform signalling and the importance of interactions with other membrane and extracellular matrix proteins. As approved therapeutics targeting the VEGF-A/VEGFR signalling axis largely lack long-term efficacy, understanding these isoform-specific mechanisms could aid future drug discovery efforts targeting VEGF receptor pharmacology.

  18. Comprehensive analysis of tropomyosin isoforms in skeletal muscles by top-down proteomics.

    Science.gov (United States)

    Jin, Yutong; Peng, Ying; Lin, Ziqing; Chen, Yi-Chen; Wei, Liming; Hacker, Timothy A; Larsson, Lars; Ge, Ying

    2016-04-01

    Mammalian skeletal muscles are heterogeneous in nature and are capable of performing various functions. Tropomyosin (Tpm) is a major component of the thin filament in skeletal muscles and plays an important role in controlling muscle contraction and relaxation. Tpm is known to consist of multiple isoforms resulting from different encoding genes and alternative splicing, along with post-translational modifications. However, a systematic characterization of Tpm isoforms in skeletal muscles is still lacking. Therefore, we employed top-down mass spectrometry (MS) to identify and characterize Tpm isoforms present in different skeletal muscles from multiple species, including swine, rat, and human. Our study revealed that Tpm1.1 and Tpm2.2 are the two major Tpm isoforms in swine and rat skeletal muscles, whereas Tpm1.1, Tpm2.2, and Tpm3.12 are present in human skeletal muscles. Tandem MS was utilized to identify the sequences of the major Tpm isoforms. Furthermore, quantitative analysis revealed muscle-type specific differences in the abundance of un-modified and modified Tpm isoforms in rat and human skeletal muscles. This study represents the first systematic investigation of Tpm isoforms in skeletal muscles, which not only demonstrates the capabilities of top-down MS for the comprehensive characterization of skeletal myofilament proteins but also provides the basis for further studies on these Tpm isoforms in muscle-related diseases.

  19. Quantification of competitive value of documents

    Directory of Open Access Journals (Sweden)

    Pavel Šimek

    2009-01-01

    Full Text Available The majority of Internet users use the global network to search for different information using fulltext search engines such as Google, Yahoo!, or Seznam. The web presentation operators are trying, with the help of different optimization techniques, to get to the top places in the results of fulltext search engines. Right there is a great importance of Search Engine Optimization and Search Engine Marketing, because normal users usually try links only on the first few pages of the fulltext search engines results on certain keywords and in catalogs they use primarily hierarchically higher placed links in each category. Key to success is the application of optimization methods which deal with the issue of keywords, structure and quality of content, domain names, individual sites and quantity and reliability of backward links. The process is demanding, long-lasting and without a guaranteed outcome. A website operator without advanced analytical tools do not identify the contribution of individual documents from which the entire web site consists. If the web presentation operators want to have an overview of their documents and web site in global, it is appropriate to quantify these positions in a specific way, depending on specific key words. For this purpose serves the quantification of competitive value of documents, which consequently sets global competitive value of a web site. Quantification of competitive values is performed on a specific full-text search engine. For each full-text search engine can be and often are, different results. According to published reports of ClickZ agency or Market Share is according to the number of searches by English-speaking users most widely used Google search engine, which has a market share of more than 80%. The whole procedure of quantification of competitive values is common, however, the initial step which is the analysis of keywords depends on a choice of the fulltext search engine.

  20. Stereo-particle image velocimetry uncertainty quantification

    International Nuclear Information System (INIS)

    Bhattacharya, Sayantan; Vlachos, Pavlos P; Charonko, John J

    2017-01-01

    Particle image velocimetry (PIV) measurements are subject to multiple elemental error sources and thus estimating overall measurement uncertainty is challenging. Recent advances have led to a posteriori uncertainty estimation methods for planar two-component PIV. However, no complete methodology exists for uncertainty quantification in stereo PIV. In the current work, a comprehensive framework is presented to quantify the uncertainty stemming from stereo registration error and combine it with the underlying planar velocity uncertainties. The disparity in particle locations of the dewarped images is used to estimate the positional uncertainty of the world coordinate system, which is then propagated to the uncertainty in the calibration mapping function coefficients. Next, the calibration uncertainty is combined with the planar uncertainty fields of the individual cameras through an uncertainty propagation equation and uncertainty estimates are obtained for all three velocity components. The methodology was tested with synthetic stereo PIV data for different light sheet thicknesses, with and without registration error, and also validated with an experimental vortex ring case from 2014 PIV challenge. Thorough sensitivity analysis was performed to assess the relative impact of the various parameters to the overall uncertainty. The results suggest that in absence of any disparity, the stereo PIV uncertainty prediction method is more sensitive to the planar uncertainty estimates than to the angle uncertainty, although the latter is not negligible for non-zero disparity. Overall the presented uncertainty quantification framework showed excellent agreement between the error and uncertainty RMS values for both the synthetic and the experimental data and demonstrated reliable uncertainty prediction coverage. This stereo PIV uncertainty quantification framework provides the first comprehensive treatment on the subject and potentially lays foundations applicable to volumetric

  1. RAGE receptor and its soluble isoforms in diabetes mellitus complications

    Directory of Open Access Journals (Sweden)

    Mauren Isfer Anghebem Oliveira

    2013-04-01

    Full Text Available Chronic hyperglycemia, which is present in all types of diabetes, increases the formation of advanced glycation end-products (AGEs. The interaction of AGEs with receptor of advanced glycation end-products (RAGE initiates a cascade of pro-inflammatory and pro-coagulant processes that result in oxidative stress, stimulating the formation and accumulation of more AGE molecules. This cyclic process, denominated metabolic memory, may explain the persistency of diabetic vascular complications in patients with satisfactory glycemic control. The RAGE found in several cell membranes is also present in soluble isoforms (esRAGE and cRAGE, which are generated by alternative deoxyribonucleic acid splicing or by proteolytic cleavage. This review focuses on new research into these mediators as potential biomarkers for vascular complications in diabetes.

  2. Metallothionein Isoform Expression in Benign and Malignant Thyroid Lesions.

    Science.gov (United States)

    Wojtczak, Beata; Pula, Bartosz; Gomulkiewicz, Agnieszka; Olbromski, Mateusz; Podhorska-Okolow, Marzena; Domoslawski, Paweł; Bolanowski, Marek; Daroszewski, Jacek; Dziegiel, Piotr

    2017-09-01

    Metallothioneins (MTs) are involved in numerous cell processes such as binding and transport of zinc and copper ions, differentiation, proliferation and apoptosis, therefore contributing to carcinogenesis. Scarce data exist on their expression in benign and malignant lesions of the thyroid. mRNA expression of functional isoforms of MT genes (MT1A, MT1B, MT1E, MT1F, MT1G, MT1H, MT1X, MT2A, MT4) was studied in 17 nodular goiters (NG), 12 follicular adenomas (FA) and 26 papillary thyroid carcinomas (PTC). One-way ANOVA revealed significant differences in mRNA expression levels of MT1A (pbenign and malignant lesions. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  3. Improved quantification of farnesene during microbial production from Saccharomyces cerevisiae in two-liquid-phase fermentations

    DEFF Research Database (Denmark)

    Tippmann, Stefan; Nielsen, Jens; Khoomrung, Sakda

    2016-01-01

    Organic solvents are widely used in microbial fermentations to reduce gas stripping effects and capture hydrophobic or toxic compounds. Reliable quantification of biochemical products in these overlays is highly challenging and practically difficult. Here, we present a significant improvement...... carryover could be minimized. Direct quantification of farnesene in dodecane was achieved by GC-FID whereas GC-MS demonstrated to be an excellent technique for identification of known and unknown metabolites. The GC-FID is a suitable technique for direct quantification of farnesene in complex matrices...

  4. Troponin T isoform expression is modulated during Atlantic Halibut metamorphosis

    Directory of Open Access Journals (Sweden)

    Llewellyn Lynda

    2007-06-01

    Full Text Available Abstract Background Flatfish metamorphosis is a thyroid hormone (TH driven process which leads to a dramatic change from a symmetrical larva to an asymmetrical juvenile. The effect of THs on muscle and in particular muscle sarcomer protein genes is largely unexplored in fish. The change in Troponin T (TnT, a pivotal protein in the assembly of skeletal muscles sarcomeres and a modulator of calcium driven muscle contraction, during flatfish metamophosis is studied. Results In the present study five cDNAs for halibut TnT genes were cloned; three were splice variants arising from a single fast TnT (fTnT gene; a fourth encoded a novel teleost specific fTnT-like cDNA (AfTnT expressed exclusively in slow muscle and the fifth encoded the teleost specific sTnT2. THs modified the expression of halibut fTnT isoforms which changed from predominantly basic to acidic isoforms during natural and T4 induced metamorphosis. In contrast, expression of red muscle specific genes, AfTnT and sTnT2, did not change during natural metamorphosis or after T4 treatment. Prior to and after metamorphosis no change in the dorso-ventral symmetry or temporal-spatial expression pattern of TnT genes and muscle fibre organization occurred in halibut musculature. Conclusion Muscle organisation in halibut remains symmetrical even after metamorphosis suggesting TH driven changes are associated with molecular adaptations. We hypothesize that species specific differences in TnT gene expression in teleosts underlies different larval muscle developmental programs which better adapts them to the specific ecological constraints.

  5. Benchmark of systematic human action reliability procedure

    International Nuclear Information System (INIS)

    Spurgin, A.J.; Hannaman, G.W.; Moieni, P.

    1986-01-01

    Probabilistic risk assessment (PRA) methodology has emerged as one of the most promising tools for assessing the impact of human interactions on plant safety and understanding the importance of the man/machine interface. Human interactions were considered to be one of the key elements in the quantification of accident sequences in a PRA. The approach to quantification of human interactions in past PRAs has not been very systematic. The Electric Power Research Institute sponsored the development of SHARP to aid analysts in developing a systematic approach for the evaluation and quantification of human interactions in a PRA. The SHARP process has been extensively peer reviewed and has been adopted by the Institute of Electrical and Electronics Engineers as the basis of a draft guide for the industry. By carrying out a benchmark process, in which SHARP is an essential ingredient, however, it appears possible to assess the strengths and weaknesses of SHARP to aid human reliability analysts in carrying out human reliability analysis as part of a PRA

  6. Verb aspect, alternations and quantification

    Directory of Open Access Journals (Sweden)

    Svetla Koeva

    2015-11-01

    Full Text Available Verb aspect, alternations and quantification In this paper we are briefly discuss the nature of Bulgarian verb aspect and argue that the verb aspect pairs are different lexical units with different (although related meaning, different argument structure (reflecting categories, explicitness and referential status of arguments and different sets of semantic and syntactic alternations. The verb prefixes resulting in perfective verbs derivation in some cases can be interpreted as lexical quantifiers as well. Thus the Bulgarian verb aspect is related (in different way both with the potential for the generation of alternations and with the prefixal lexical quantification. It is shown that the scope of the lexical quantification by means of verbal prefixes is the quantified verb phrase and the scope remains constant in all derived alternations. The paper concerns the basic issues of these complex problems, while the detailed description of the conditions satisfying particular alternation or particular lexical quantification are subject of a more detailed study.

  7. Uncertainty quantification for environmental models

    Science.gov (United States)

    Hill, Mary C.; Lu, Dan; Kavetski, Dmitri; Clark, Martyn P.; Ye, Ming

    2012-01-01

    ]. There are also bootstrapping and cross-validation approaches.Sometimes analyses are conducted using surrogate models [12]. The availability of so many options can be confusing. Categorizing methods based on fundamental questions assists in communicating the essential results of uncertainty analyses to stakeholders. Such questions can focus on model adequacy (e.g., How well does the model reproduce observed system characteristics and dynamics?) and sensitivity analysis (e.g., What parameters can be estimated with available data? What observations are important to parameters and predictions? What parameters are important to predictions?), as well as on the uncertainty quantification (e.g., How accurate and precise are the predictions?). The methods can also be classified by the number of model runs required: few (10s to 1000s) or many (10,000s to 1,000,000s). Of the methods listed above, the most computationally frugal are generally those based on local derivatives; MCMC methods tend to be among the most computationally demanding. Surrogate models (emulators)do not necessarily produce computational frugality because many runs of the full model are generally needed to create a meaningful surrogate model. With this categorization, we can, in general, address all the fundamental questions mentioned above using either computationally frugal or demanding methods. Model development and analysis can thus be conducted consistently using either computation-ally frugal or demanding methods; alternatively, different fundamental questions can be addressed using methods that require different levels of effort. Based on this perspective, we pose the question: Can computationally frugal methods be useful companions to computationally demanding meth-ods? The reliability of computationally frugal methods generally depends on the model being reasonably linear, which usually means smooth nonlin-earities and the assumption of Gaussian errors; both tend to be more valid with more linear

  8. Quantification of renal function

    International Nuclear Information System (INIS)

    Mubarak, Amani Hayder

    1999-06-01

    The evaluation of glomerular filtration rate (GFR) with Tc99m-DTPA using single injection with multiple blood sample method (plasma clearance), is a standard and reliable method but the procedure is complicated and may not suitable for routine clinical use. Alternatively, estimation of GFR by using Tc99m-DTPA and gamma camera computer system is very simple, dose not require sampling of blood or urine and provide individual kidney value of GFR (integral, uptake index methods)

  9. Quantification of informed opinion

    International Nuclear Information System (INIS)

    Rasmuson, D.M.

    1985-01-01

    The objective of this session, Quantification of Informed Opinion, is to provide the statistician with a better understanding of this important area. The NRC uses informed opinion, sometimes called engineering judgment or subjective judgment, in many areas. Sometimes informed opinion is the only source of information that exists, especially in phenomenological areas, such as steam explosions, where experiments are costly and phenomena are very difficult to measure. There are many degrees of informed opinion. These vary from the weatherman who makes predictions concerning relatively high probability events with a large data base to the phenomenological expert who must use his intuition tempered with basic knowledge and little or no measured data to predict the behavior of events with a low probability of occurrence. The first paper in this session provides the reader with an overview of the subject area. The second paper provides some aspects that must be considered in the collection of informed opinion to improve the quality of the information. The final paper contains an example of the use of informed opinion in the area of seismic hazard characterization. These papers should be useful to researchers and statisticians who need to collect and use informed opinion in their work

  10. Quantification In Neurology

    Directory of Open Access Journals (Sweden)

    Netravati M

    2005-01-01

    Full Text Available There is a distinct shift of emphasis in clinical neurology in the last few decades. A few years ago, it was just sufficient for a clinician to precisely record history, document signs, establish diagnosis and write prescription. In the present context, there has been a significant intrusion of scientific culture in clinical practice. Several criteria have been proposed, refined and redefined to ascertain accurate diagnosis for many neurological disorders. Introduction of the concept of impairment, disability, handicap and quality of life has added new dimension to the measurement of health and disease and neurological disorders are no exception. "Best guess" treatment modalities are no more accepted and evidence based medicine has become an integral component of medical care. Traditional treatments need validation and new therapies require vigorous trials. Thus, proper quantification in neurology has become essential, both in practice and research methodology in neurology. While this aspect is widely acknowledged, there is a limited access to a comprehensive document pertaining to measurements in neurology. This following description is a critical appraisal of various measurements and also provides certain commonly used rating scales/scores in neurological practice.

  11. AMSAA Reliability Growth Guide

    National Research Council Canada - National Science Library

    Broemm, William

    2000-01-01

    ... has developed reliability growth methodology for all phases of the process, from planning to tracking to projection. The report presents this methodology and associated reliability growth concepts.

  12. The fair value of operational reliability

    International Nuclear Information System (INIS)

    Patino-Echeverri, Dalia; Morel, Benoit

    2005-01-01

    Information about the uncertainties that surround the operation of the power system can be used to enlighten the debate of how much reliability should be pursued and how resources should be allocated to pursue it. In this paper we present a method to determine the value of having flexible generators to react to load fluctuations. This value can be seen as the value of hedging against the uncertainty on the load due to the volatility of the demand and the possibility of congestion. Because having this flexibility can be related to a financial option, we will use an extension of options theory and in particular the risk-neutral valuation method, to find a risk neutral quantification of its value. We illustrate our point valuing the flexibility that leads to ''operational reliability'' in the PJM market. Our formula for that value is what we call ''the fair value'' of operational reliability. (Author)

  13. Glycosylation differences contribute to distinct catalytic properties among bone alkaline phosphatase isoforms.

    Science.gov (United States)

    Halling Linder, Cecilia; Narisawa, Sonoko; Millán, José Luis; Magnusson, Per

    2009-11-01

    Three circulating human bone alkaline phosphatase (BALP) isoforms (B1, B2, and B/I) can be distinguished in healthy individuals and a fourth isoform (B1x) has been discovered in patients with chronic kidney disease and in bone tissue. The present study was designed to correlate differing glycosylation patterns of each BALP isoform with their catalytic activity towards presumptive physiological substrates and to compare those properties with two recombinant isoforms of the tissue-nonspecific ALP (TNALP) isozyme, i.e., TNALP-flag, used extensively for mutation analysis of hypophosphatasia mutations and sALP-FcD(10), a chimeric enzyme recently used as therapeutic drug in a mouse model of infantile hypophosphatasia. The BALP isoforms were prepared from human osteosarcoma (SaOS-2) cells and the kinetic properties were evaluated using the synthetic substrate p-nitrophenylphosphate (pNPP) at pH 7.4 and 9.8, and the three suggested endogenous physiological substrates, i.e., inorganic pyrophosphate (PP(i)), pyridoxal 5'-phosphate (PLP), and phosphoethanolamine (PEA) at pH 7.4. Qualitative glycosylation differences were also assessed by lectin binding and precipitation. The k(cat)/K(M) was higher for B2 for all the investigated substrates. The catalytic activity towards PEA was essentially undetectable. The kinetic activity for TNALP-flag and sALP-FcD(10) was similar to the activity of the human BALP isoforms. The BALP isoforms differed in their lectin binding properties and dose-dependent lectin precipitation, which also demonstrated differences between native and denatured BALP isoforms. The observed differences in lectin specificity were attributed to N-linked carbohydrates. In conclusion, we demonstrate significantly different catalytic properties among the BALP isoforms due to structural differences in posttranslational glycosylation. Our data also suggests that PEA is not an endogenous substrate for the BALP isoforms or for the recombinant TNALP isoforms. The TNALP

  14. Human factors considerations for reliability and safety

    International Nuclear Information System (INIS)

    Carnino, A.

    1985-01-01

    Human factors in many industries have become an important issue, since the last few years. They should be considered during the whole life time of a plant: design, fabrication and construction, licensing, operation. Improvements have been performed in the field of man-machine interface such as procedures, control room lay-out, operator aids, training. In order to meet the needs of reliability and probabilistic risk studies, quantification of human errors has been developed but needs still improvements in the field of cognitive behaviour, diagnosis and representation errors. Data banks to support these quantifications are still in a development stage. This applies to nuclear power plants and several examples are given to illustrate the above ideas. In conclusion, human factors field is in a very quickly evolving process but the tendency is still to adapt the man to the machines whilst the reverse would be desirable

  15. A reliability simulation language for reliability analysis

    International Nuclear Information System (INIS)

    Deans, N.D.; Miller, A.J.; Mann, D.P.

    1986-01-01

    The results of work being undertaken to develop a Reliability Description Language (RDL) which will enable reliability analysts to describe complex reliability problems in a simple, clear and unambiguous way are described. Component and system features can be stated in a formal manner and subsequently used, along with control statements to form a structured program. The program can be compiled and executed on a general-purpose computer or special-purpose simulator. (DG)

  16. Inulin isoforms differ by repeated additions of one crystal unit cell

    Science.gov (United States)

    Cooper, Peter D.; Barclay, Thomas G.; Ginic-Markovic, Milena; Gerson, Andrea R.; Petrovsky, Nikolai

    2014-01-01

    Inulin isoforms, especially delta inulin, are important biologically as immune activators and clinically as vaccine adjuvants. In exploring action mechanisms, we previously found regular increments in thermal properties of the seven-member inulin isoform series that suggested regular additions of some energetic structural unit. Because the previous isolates carried additional longer chains that masked defining ranges, these were contrasted with new isoform isolates comprising only inulin chain lengths defining that isoform. The new series began with 19 fructose units per chain (alpha-1 inulin), increasing regularly by 6 fructose units per isoform. Thus the ‘energetic unit’ equates to 6 fructose residues per chain. All isoforms showed indistinguishable X-ray diffraction patterns that were also identical with known inulin crystals. We conclude that an ‘energetic unit’ equates to one helix turn of 6 fructose units per chain as found in one unit cell of the inulin crystal. Each isoform chain comprised progressively more helix turns plus one additional fructose and glucose residues per chain. PMID:24528745

  17. VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis

    Directory of Open Access Journals (Sweden)

    Gareth W. Fearnley

    2016-05-01

    Full Text Available Vascular endothelial growth factor A (VEGF-A binding to the receptor tyrosine kinase VEGFR2 triggers multiple signal transduction pathways, which regulate endothelial cell responses that control vascular development. Multiple isoforms of VEGF-A can elicit differential signal transduction and endothelial responses. However, it is unclear how such cellular responses are controlled by isoform-specific VEGF-A–VEGFR2 complexes. Increasingly, there is the realization that the membrane trafficking of receptor–ligand complexes influences signal transduction and protein turnover. By building on these concepts, our study shows for the first time that three different VEGF-A isoforms (VEGF-A165, VEGF-A121 and VEGF-A145 promote distinct patterns of VEGFR2 endocytosis for delivery into early endosomes. This differential VEGFR2 endocytosis and trafficking is linked to VEGF-A isoform-specific signal transduction events. Disruption of clathrin-dependent endocytosis blocked VEGF-A isoform-specific VEGFR2 activation, signal transduction and caused substantial depletion in membrane-bound VEGFR1 and VEGFR2 levels. Furthermore, such VEGF-A isoforms promoted differential patterns of VEGFR2 ubiquitylation, proteolysis and terminal degradation. Our study now provides novel insights into how different VEGF-A isoforms can bind the same receptor tyrosine kinase and elicit diverse cellular outcomes.

  18. VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis.

    Science.gov (United States)

    Fearnley, Gareth W; Smith, Gina A; Abdul-Zani, Izma; Yuldasheva, Nadira; Mughal, Nadeem A; Homer-Vanniasinkam, Shervanthi; Kearney, Mark T; Zachary, Ian C; Tomlinson, Darren C; Harrison, Michael A; Wheatcroft, Stephen B; Ponnambalam, Sreenivasan

    2016-05-15

    Vascular endothelial growth factor A (VEGF-A) binding to the receptor tyrosine kinase VEGFR2 triggers multiple signal transduction pathways, which regulate endothelial cell responses that control vascular development. Multiple isoforms of VEGF-A can elicit differential signal transduction and endothelial responses. However, it is unclear how such cellular responses are controlled by isoform-specific VEGF-A-VEGFR2 complexes. Increasingly, there is the realization that the membrane trafficking of receptor-ligand complexes influences signal transduction and protein turnover. By building on these concepts, our study shows for the first time that three different VEGF-A isoforms (VEGF-A165, VEGF-A121 and VEGF-A145) promote distinct patterns of VEGFR2 endocytosis for delivery into early endosomes. This differential VEGFR2 endocytosis and trafficking is linked to VEGF-A isoform-specific signal transduction events. Disruption of clathrin-dependent endocytosis blocked VEGF-A isoform-specific VEGFR2 activation, signal transduction and caused substantial depletion in membrane-bound VEGFR1 and VEGFR2 levels. Furthermore, such VEGF-A isoforms promoted differential patterns of VEGFR2 ubiquitylation, proteolysis and terminal degradation. Our study now provides novel insights into how different VEGF-A isoforms can bind the same receptor tyrosine kinase and elicit diverse cellular outcomes. © 2016. Published by The Company of Biologists Ltd.

  19. Regulation of cardiac remodeling by cardiac Na/K-ATPase isoforms

    Directory of Open Access Journals (Sweden)

    Lijun Catherine Liu

    2016-09-01

    Full Text Available Cardiac remodeling occurs after cardiac pressure/volume overload or myocardial injury during the development of heart failure and is a determinant of heart failure. Preventing or reversing remodeling is a goal of heart failure therapy. Human cardiomyocyte Na+/K+-ATPase has multiple α isoforms (1-3. The expression of the α subunit of the Na+/K+-ATPase is often altered in hypertrophic and failing hearts. The mechanisms are unclear. There are limited data from human cardiomyocytes. Abundant evidences from rodents show that Na+/K+-ATPase regulates cardiac contractility, cell signaling, hypertrophy and fibrosis. The α1 isoform of the Na+/K+-ATPase is the ubiquitous isoform and possesses both pumping and signaling functions. The α2 isoform of the Na+/K+-ATPase regulates intracellular Ca2+ signaling, contractility and pathological hypertrophy. The α3 isoform of the Na+/K+-ATPase may also be a target for cardiac hypertrophy. Restoration of cardiac Na+/K+-ATPase expression may be an effective approach for prevention of cardiac remodeling. In this article, we will overview: (1 the distribution and function of isoform specific Na+/K+-ATPase in the cardiomyocytes. (2 the role of cardiac Na+/K+-ATPase in the regulation of cell signaling, contractility, cardiac hypertrophy and fibrosis in vitro and in vivo. Selective targeting of cardiac Na+/K+-ATPase isoform may offer a new target for the prevention of cardiac remodeling.

  20. High Molecular Weight Isoforms of Growth Hormone In Cells of the Immune System

    Science.gov (United States)

    Weigent, Douglas A.

    2013-01-01

    A substantial body of research exists to support the idea that cells of the immune system produce growth hormone (GH). However, the structure and mechanism of action of lymphocyte-derived GH continues to remain largely unknown. Here we present the results of Western analysis of whole cell extracts showing that different molecular weight isoforms of GH of approximately 100 kDa, 65 kDa, and 48 kDa can be detected in primary mouse cells of the immune system and in the mouse EL4 cell line. The identity of the 65 kDa and 48 kDa isoforms of GH were confirmed by mass spectrometry. The various isoforms were detected in both enriched T and B spleen cell populations. The large molecular weight isoform appears to reside primarily in the cytoplasm whereas the lower molecular weight 65 kDa and 48 kDa isoforms were detected primarily in the nucleus. These results also suggest that GH isoforms are induced by oxidative stress. In EL4 cells overexpressing GH, the expression of luciferase controlled by a promoter containing the antioxidant response element is increased almost three-fold above control. The data suggest that the induction of isoforms of the GH molecule in cells of the immune system may be an important mechanism of adaptation and/or protection of lymphoid cells under conditions of oxidative stress. PMID:21741628

  1. Isoform-selective regulation of glycogen phosphorylase by energy deprivation and phosphorylation in astrocytes.

    Science.gov (United States)

    Müller, Margit S; Pedersen, Sofie E; Walls, Anne B; Waagepetersen, Helle S; Bak, Lasse K

    2015-01-01

    Glycogen phosphorylase (GP) is activated to degrade glycogen in response to different stimuli, to support both the astrocyte's own metabolic demand and the metabolic needs of neurons. The regulatory mechanism allowing such a glycogenolytic response to distinct triggers remains incompletely understood. In the present study, we used siRNA-mediated differential knockdown of the two isoforms of GP expressed in astrocytes, muscle isoform (GPMM), and brain isoform (GPBB), to analyze isoform-specific regulatory characteristics in a cellular setting. Subsequently, we tested the response of each isoform to phosphorylation, triggered by incubation with norepinephrine (NE), and to AMP, increased by glucose deprivation in cells in which expression of one GP isoform had been silenced. Successful knockdown was demonstrated on the protein level by Western blot, and on a functional level by determination of glycogen content showing an increase in glycogen levels following knockdown of either GPMM or GPBB. NE triggered glycogenolysis within 15 min in control cells and after GPBB knockdown. However, astrocytes in which expression of GPMM had been silenced showed a delay in response to NE, with glycogen levels significantly reduced only after 60 min. In contrast, allosteric activation of GP by AMP, induced by glucose deprivation, seemed to mainly affect GPBB, as only knockdown of GPBB, but not of GPMM, delayed the glycogenolytic response to glucose deprivation. Our results indicate that the two GP isoforms expressed in astrocytes respond to different physiological triggers, therefore conferring distinct metabolic functions of brain glycogen. © 2014 Wiley Periodicals, Inc.

  2. Myosin isoform fiber type and fiber size in the tail of the Virginia opossum (Didelphis virginiana).

    Science.gov (United States)

    Hazimihalis, P J; Gorvet, M A; Butcher, M T

    2013-01-01

    Muscle fiber type is a well studied property in limb muscles, however, much less is understood about myosin heavy chain (MHC) isoform expression in caudal muscles of mammalian tails. Didelphid marsupials are an interesting lineage in this context as all species have prehensile tails, but show a range of tail-function depending on either their arboreal or terrestrial locomotor habits. Differences in prehensility suggest that MHC isoform fiber types may also be different, in that terrestrial opossums may have a large distribution of oxidative fibers for object carrying tasks instead of faster, glycolytic fiber types expected in mammals with long tails. To test this hypothesis, MHC isoform fiber type and their regional distribution (proximal/transitional/distal) were determined in the tail of the Virginia opossum (Didelphis virginiana). Fiber types were determined by a combination of myosin-ATPase histochemistry, immunohistochemistry, and SDS-PAGE. Results indicate a predominance of the fast MHC-2A and -2X isoforms in each region of the tail. The presence of two fast isoforms, in addition to the slow MHC-1 isoform, was confirmed by SDS-PAGE analysis. The overall MHC isoform fiber type distribution for the tail was: 25% MHC-1, 71% MHC-2A/X hybrid, and 4% MHC-1/2A hybrid. Oxidative MHC-2A/X isoform fibers were found to be relatively large in cross-section compared to slow, oxidative MHC-1 and MHC-1/2A hybrid fibers. A large percentage of fast MHC-2A/X hybrids fibers may be suggestive of an evolutionary transition in MHC isoform distribution (fast-to-slow fiber type) in the tail musculature of an opossum with primarily a terrestrial locomotor habit and adaptive tail-function. Copyright © 2012 Wiley Periodicals, Inc.

  3. Detection of VEGF-A(xxx)b isoforms in human tissues.

    Science.gov (United States)

    Bates, David O; Mavrou, Athina; Qiu, Yan; Carter, James G; Hamdollah-Zadeh, Maryam; Barratt, Shaney; Gammons, Melissa V; Millar, Ann B; Salmon, Andrew H J; Oltean, Sebastian; Harper, Steven J

    2013-01-01

    Vascular Endothelial Growth Factor-A (VEGF-A) can be generated as multiple isoforms by alternative splicing. Two families of isoforms have been described in humans, pro-angiogenic isoforms typified by VEGF-A165a, and anti-angiogenic isoforms typified by VEGF-A165b. The practical determination of expression levels of alternative isoforms of the same gene may be complicated by experimental protocols that favour one isoform over another, and the use of specific positive and negative controls is essential for the interpretation of findings on expression of the isoforms. Here we address some of the difficulties in experimental design when investigating alternative splicing of VEGF isoforms, and discuss the use of appropriate control paradigms. We demonstrate why use of specific control experiments can prevent assumptions that VEGF-A165b is not present, when in fact it is. We reiterate, and confirm previously published experimental design protocols that demonstrate the importance of using positive controls. These include using known target sequences to show that the experimental conditions are suitable for PCR amplification of VEGF-A165b mRNA for both q-PCR and RT-PCR and to ensure that mispriming does not occur. We also provide evidence that demonstrates that detection of VEGF-A165b protein in mice needs to be tightly controlled to prevent detection of mouse IgG by a secondary antibody. We also show that human VEGF165b protein can be immunoprecipitated from cultured human cells and that immunoprecipitating VEGF-A results in protein that is detected by VEGF-A165b antibody. These findings support the conclusion that more information on the biology of VEGF-A165b isoforms is required, and confirm the importance of the experimental design in such investigations, including the use of specific positive and negative controls.

  4. The polysaccharide inulin is characterized by an extensive series of periodic isoforms with varying biological actions

    Science.gov (United States)

    Cooper, Peter D; Barclay, Thomas G; Ginic-Markovic, Milena; Petrovsky, Nikolai

    2013-01-01

    In studying the molecular basis for the potent immune activity of previously described gamma and delta inulin particles and to assist in production of inulin adjuvants under Good Manufacturing Practice, we identified five new inulin isoforms, bringing the total to seven plus the amorphous form. These isoforms comprise the step-wise inulin developmental series amorphous → alpha-1 (AI-1) → alpha-2 (AI-2) → gamma (GI) → delta (DI) → zeta (ZI) → epsilon (EI) → omega (OI) in which each higher isoform can be made either by precipitating dissolved inulin or by direct conversion from its precursor, both cases using regularly increasing temperatures. At higher temperatures, the shorter inulin polymer chains are released from the particle and so the key difference between isoforms is that each higher isoform comprises longer polymer chains than its precursor. An increasing trend of degree of polymerization is confirmed by end-group analysis using 1H nuclear magnetic resonance spectroscopy. Inulin isoforms were characterized by the critical temperatures of abrupt phase-shifts (solubilizations or precipitations) in water suspensions. Such (aqueous) “melting” or “freezing” points are diagnostic and occur in strikingly periodic steps reflecting quantal increases in noncovalent bonding strength and increments in average polymer lengths. The (dry) melting points as measured by modulated differential scanning calorimetry similarly increase in regular steps. We conclude that the isoforms differ in repeated increments of a precisely repeating structural element. Each isoform has a different spectrum of biological activities and we show the higher inulin isoforms to be more potent alternative complement pathway activators. PMID:23853206

  5. Learning-dependent gene expression of CREB1 isoforms in the molluscan brain

    Directory of Open Access Journals (Sweden)

    Hisayo Sadamoto

    2010-05-01

    Full Text Available Cyclic AMP-responsive element binding protein1 (CREB1 has multiple functions in gene regulation. Various studies have reported that CREB1-dependent gene induction is necessary for memory formation and long-lasting behavioral changes in both vertebrates and invertebrates. In the present study, we characterized Lymnaea CREB1 (LymCREB1 mRNA isoforms of spliced variants in the central nervous system (CNS of the pond snail Lymnaea stagnalis. Among these spliced variants, the three isoforms that code a whole LymCREB1 protein are considered to be the activators for gene regulation. The other four isoforms, which code truncated LymCREB1 proteins with no kinase inducible domain, are the repressors. For a better understanding of the possible roles of different LymCREB1 isoforms, the expression level of these isoform mRNAs was investigated by a real-time quantitative RT-PCR method. Further, we examined the changes in gene expression for all the isoforms in the CNS after conditioned taste aversion (CTA learning or backward conditioning as a control. The results showed that CTA learning increased LymCREB1 gene expression, but it did not change the activator/repressor ratio. Our findings showed that the repressor isoforms, as well as the activator ones, are expressed in large amounts in the CNS, and the gene expression of CREB1 isoforms appeared to be specific for the given stimulus. This was the first quantitative analysis of the expression patterns of CREB1 isoforms at the mRNA level and their association with learning behavior.

  6. Progesterone receptor isoform A may regulate the effects of neoadjuvant aglepristone in canine mammary carcinoma

    DEFF Research Database (Denmark)

    Guil-Luna, Silvia; Stenvang, Jan; Brünner, Nils

    2014-01-01

    RNA expression of progesterone receptor isoforms A and B in mammary carcinomas in dogs treated with 20 mg/Kg of aglepristone (n¿=¿22) or vehicle (n¿=¿5) twice before surgery.ResultsFormalin-fixed, paraffin-embedded tissue samples taken before and after treatment were used to analyse total progesterone receptor......-receptor positive and isoform-A positive tumours in aglepristone-treated dogs.ConclusionsThese findings suggest that the antiproliferative effects of aglepristone in canine mammary carcinomas are mediated by progesterone receptor isoform A....

  7. A taxonomy for human reliability analysis

    International Nuclear Information System (INIS)

    Beattie, J.D.; Iwasa-Madge, K.M.

    1984-01-01

    A human interaction taxonomy (classification scheme) was developed to facilitate human reliability analysis in a probabilistic safety evaluation of a nuclear power plant, being performed at Ontario Hydro. A human interaction occurs, by definition, when operators or maintainers manipulate, or respond to indication from, a plant component or system. The taxonomy aids the fault tree analyst by acting as a heuristic device. It helps define the range and type of human errors to be identified in the construction of fault trees, while keeping the identification by different analysts consistent. It decreases the workload associated with preliminary quantification of the large number of identified interactions by including a category called 'simple interactions'. Fault tree analysts quantify these according to a procedure developed by a team of human reliability specialists. The interactions which do not fit into this category are called 'complex' and are quantified by the human reliability team. The taxonomy is currently being used in fault tree construction in a probabilistic safety evaluation. As far as can be determined at this early stage, the potential benefits of consistency and completeness in identifying human interactions and streamlining the initial quantification are being realized

  8. Effects of humic acid on DNA quantification with Quantifiler® Human DNA Quantification kit and short tandem repeat amplification efficiency.

    Science.gov (United States)

    Seo, Seung Bum; Lee, Hye Young; Zhang, Ai Hua; Kim, Hye Yeon; Shin, Dong Hoon; Lee, Soong Deok

    2012-11-01

    Correct DNA quantification is an essential part to obtain reliable STR typing results. Forensic DNA analysts often use commercial kits for DNA quantification; among them, real-time-based DNA quantification kits are most frequently used. Incorrect DNA quantification due to the presence of PCR inhibitors may affect experiment results. In this study, we examined the alteration degree of DNA quantification results estimated in DNA samples containing a PCR inhibitor by using a Quantifiler® Human DNA Quantification kit. For experiments, we prepared approximately 0.25 ng/μl DNA samples containing various concentrations of humic acid (HA). The quantification results were 0.194-0.303 ng/μl at 0-1.6 ng/μl HA (final concentration in the Quantifiler reaction) and 0.003-0.168 ng/μl at 2.4-4.0 ng/μl HA. Most DNA quantity was undetermined when HA concentration was higher than 4.8 ng/μl HA. The C (T) values of an internal PCR control (IPC) were 28.0-31.0, 36.5-37.1, and undetermined at 0-1.6, 2.4, and 3.2 ng/μl HA. These results indicate that underestimated DNA quantification results may be obtained in the DNA sample with high C (T) values of IPC. Thus, researchers should carefully interpret the DNA quantification results. We additionally examined the effects of HA on the STR amplification by using an Identifiler® kit and a MiniFiler™ kit. Based on the results of this study, it is thought that a better understanding of various effects of HA would help researchers recognize and manipulate samples containing HA.

  9. Critical points of DNA quantification by real-time PCR – effects of DNA extraction method and sample matrix on quantification of genetically modified organisms

    Directory of Open Access Journals (Sweden)

    Žel Jana

    2006-08-01

    Full Text Available Abstract Background Real-time PCR is the technique of choice for nucleic acid quantification. In the field of detection of genetically modified organisms (GMOs quantification of biotech products may be required to fulfil legislative requirements. However, successful quantification depends crucially on the quality of the sample DNA analyzed. Methods for GMO detection are generally validated on certified reference materials that are in the form of powdered grain material, while detection in routine laboratories must be performed on a wide variety of sample matrixes. Due to food processing, the DNA in sample matrixes can be present in low amounts and also degraded. In addition, molecules of plant origin or from other sources that affect PCR amplification of samples will influence the reliability of the quantification. Further, the wide variety of sample matrixes presents a challenge for detection laboratories. The extraction method must ensure high yield and quality of the DNA obtained and must be carefully selected, since even components of DNA extraction solutions can influence PCR reactions. GMO quantification is based on a standard curve, therefore similarity of PCR efficiency for the sample and standard reference material is a prerequisite for exact quantification. Little information on the performance of real-time PCR on samples of different matrixes is available. Results Five commonly used DNA extraction techniques were compared and their suitability for quantitative analysis was assessed. The effect of sample matrix on nucleic acid quantification was assessed by comparing 4 maize and 4 soybean matrixes. In addition 205 maize and soybean samples from routine analysis were analyzed for PCR efficiency to assess variability of PCR performance within each sample matrix. Together with the amount of DNA needed for reliable quantification, PCR efficiency is the crucial parameter determining the reliability of quantitative results, therefore it was

  10. Critical points of DNA quantification by real-time PCR--effects of DNA extraction method and sample matrix on quantification of genetically modified organisms.

    Science.gov (United States)

    Cankar, Katarina; Stebih, Dejan; Dreo, Tanja; Zel, Jana; Gruden, Kristina

    2006-08-14

    Real-time PCR is the technique of choice for nucleic acid quantification. In the field of detection of genetically modified organisms (GMOs) quantification of biotech products may be required to fulfil legislative requirements. However, successful quantification depends crucially on the quality of the sample DNA analyzed. Methods for GMO detection are generally validated on certified reference materials that are in the form of powdered grain material, while detection in routine laboratories must be performed on a wide variety of sample matrixes. Due to food processing, the DNA in sample matrixes can be present in low amounts and also degraded. In addition, molecules of plant origin or from other sources that affect PCR amplification of samples will influence the reliability of the quantification. Further, the wide variety of sample matrixes presents a challenge for detection laboratories. The extraction method must ensure high yield and quality of the DNA obtained and must be carefully selected, since even components of DNA extraction solutions can influence PCR reactions. GMO quantification is based on a standard curve, therefore similarity of PCR efficiency for the sample and standard reference material is a prerequisite for exact quantification. Little information on the performance of real-time PCR on samples of different matrixes is available. Five commonly used DNA extraction techniques were compared and their suitability for quantitative analysis was assessed. The effect of sample matrix on nucleic acid quantification was assessed by comparing 4 maize and 4 soybean matrixes. In addition 205 maize and soybean samples from routine analysis were analyzed for PCR efficiency to assess variability of PCR performance within each sample matrix. Together with the amount of DNA needed for reliable quantification, PCR efficiency is the crucial parameter determining the reliability of quantitative results, therefore it was chosen as the primary criterion by which to

  11. Critical points of DNA quantification by real-time PCR – effects of DNA extraction method and sample matrix on quantification of genetically modified organisms

    Science.gov (United States)

    Cankar, Katarina; Štebih, Dejan; Dreo, Tanja; Žel, Jana; Gruden, Kristina

    2006-01-01

    Background Real-time PCR is the technique of choice for nucleic acid quantification. In the field of detection of genetically modified organisms (GMOs) quantification of biotech products may be required to fulfil legislative requirements. However, successful quantification depends crucially on the quality of the sample DNA analyzed. Methods for GMO detection are generally validated on certified reference materials that are in the form of powdered grain material, while detection in routine laboratories must be performed on a wide variety of sample matrixes. Due to food processing, the DNA in sample matrixes can be present in low amounts and also degraded. In addition, molecules of plant origin or from other sources that affect PCR amplification of samples will influence the reliability of the quantification. Further, the wide variety of sample matrixes presents a challenge for detection laboratories. The extraction method must ensure high yield and quality of the DNA obtained and must be carefully selected, since even components of DNA extraction solutions can influence PCR reactions. GMO quantification is based on a standard curve, therefore similarity of PCR efficiency for the sample and standard reference material is a prerequisite for exact quantification. Little information on the performance of real-time PCR on samples of different matrixes is available. Results Five commonly used DNA extraction techniques were compared and their suitability for quantitative analysis was assessed. The effect of sample matrix on nucleic acid quantification was assessed by comparing 4 maize and 4 soybean matrixes. In addition 205 maize and soybean samples from routine analysis were analyzed for PCR efficiency to assess variability of PCR performance within each sample matrix. Together with the amount of DNA needed for reliable quantification, PCR efficiency is the crucial parameter determining the reliability of quantitative results, therefore it was chosen as the primary

  12. Arogenate Dehydratase Isoforms Differentially Regulate Anthocyanin Biosynthesis in Arabidopsis thaliana.

    Science.gov (United States)

    Chen, Qingbo; Man, Cong; Li, Danning; Tan, Huijuan; Xie, Ye; Huang, Jirong

    2016-12-05

    Anthocyanins, a group of L-phenylalanine (Phe)-derived flavonoids, have been demonstrated to play important roles in plant stress resistance and interactions between plants and insects. Although the anthocyanin biosynthetic pathway and its regulatory mechanisms have been extensively studied, it remains unclear whether the level of Phe supply affects anthocyanin biosynthesis. Here, we investigated the roles of arogenate dehydratases (ADTs), the key enzymes that catalyze the conversion of arogenate into Phe, in sucrose-induced anthocyanin biosynthesis in Arabidopsis. Genetic analysis showed that all six ADT isoforms function redundantly in anthocyanin biosynthesis but have differential contributions. ADT2 contributes the most to anthocyanin accumulation, followed by ADT1 and ADT3, and ADT4-ADT6. We found that anthocyanin content is positively correlated with the levels of Phe and sucrose-induced ADT transcripts in seedlings. Consistently, addition of Phe to the medium could dramatically increase anthocyanin content in the wild-type plants and rescue the phenotype of the adt1 adt3 double mutant regarding the anthocyanin accumulation. Moreover, transgenic plants overexpressing ADT4, which appears to be less sensitive to Phe than overexpression of ADT2, hyperaccumulate Phe and produce elevated level of anthocyanins. Taken together, our results suggest that the level of Phe is an important regulatory factor for sustaining anthocyanin biosynthesis. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  13. Differential susceptibility of RAE-1 isoforms to mouse cytomegalovirus.

    Science.gov (United States)

    Arapovic, Jurica; Lenac, Tihana; Antulov, Ronald; Polic, Bojan; Ruzsics, Zsolt; Carayannopoulos, Leonidas N; Koszinowski, Ulrich H; Krmpotic, Astrid; Jonjic, Stipan

    2009-08-01

    The NKG2D receptor is one of the most potent activating natural killer cell receptors involved in antiviral responses. The mouse NKG2D ligands MULT-1, RAE-1, and H60 are regulated by murine cytomegalovirus (MCMV) proteins m145, m152, and m155, respectively. In addition, the m138 protein interferes with the expression of both MULT-1 and H60. We show here that one of five RAE-1 isoforms, RAE-1delta, is resistant to downregulation by MCMV and that this escape has functional importance in vivo. Although m152 retained newly synthesized RAE-1delta and RAE-1gamma in the endoplasmic reticulum, no viral regulator was able to affect the mature RAE-1delta form which remains expressed on the surfaces of infected cells. This differential susceptibility to downregulation by MCMV is not a consequence of faster maturation of RAE-1delta compared to RAE-1gamma but rather an intrinsic property of the mature surface-resident protein. This difference can be attributed to the absence of a PLWY motif from RAE-1delta. Altogether, these findings provide evidence for a novel mechanism of host escape from viral immunoevasion of NKG2D-dependent control.

  14. Results of the event sequence reliability benchmark exercise

    International Nuclear Information System (INIS)

    Silvestri, E.

    1990-01-01

    The Event Sequence Reliability Benchmark Exercise is the fourth of a series of benchmark exercises on reliability and risk assessment, with specific reference to nuclear power plant applications, and is the logical continuation of the previous benchmark exercises on System Analysis Common Cause Failure and Human Factors. The reference plant is the Nuclear Power Plant at Grohnde Federal Republic of Germany a 1300 MW PWR plant of KWU design. The specific objective of the Exercise is to model, to quantify and to analyze such event sequences initiated by the occurrence of a loss of offsite power that involve the steam generator feed. The general aim is to develop a segment of a risk assessment, which ought to include all the specific aspects and models of quantification, such as common canal failure, Human Factors and System Analysis, developed in the previous reliability benchmark exercises, with the addition of the specific topics of dependences between homologous components belonging to different systems featuring in a given event sequence and of uncertainty quantification, to end up with an overall assessment of: - the state of the art in risk assessment and the relative influences of quantification problems in a general risk assessment framework. The Exercise has been carried out in two phases, both requiring modelling and quantification, with the second phase adopting more restrictive rules and fixing certain common data, as emerged necessary from the first phase. Fourteen teams have participated in the Exercise mostly from EEC countries, with one from Sweden and one from the USA. (author)

  15. Reliability data banks

    International Nuclear Information System (INIS)

    Cannon, A.G.; Bendell, A.

    1991-01-01

    Following an introductory chapter on Reliability, what is it, why it is needed, how it is achieved and measured, the principles of reliability data bases and analysis methodologies are the subject of the next two chapters. Achievements due to the development of data banks are mentioned for different industries in the next chapter, FACTS, a comprehensive information system for industrial safety and reliability data collection in process plants are covered next. CREDO, the Central Reliability Data Organization is described in the next chapter and is indexed separately, as is the chapter on DANTE, the fabrication reliability Data analysis system. Reliability data banks at Electricite de France and IAEA's experience in compiling a generic component reliability data base are also separately indexed. The European reliability data system, ERDS, and the development of a large data bank come next. The last three chapters look at 'Reliability data banks, - friend foe or a waste of time'? and future developments. (UK)

  16. Suncor maintenance and reliability

    Energy Technology Data Exchange (ETDEWEB)

    Little, S. [Suncor Energy, Calgary, AB (Canada)

    2006-07-01

    Fleet maintenance and reliability at Suncor Energy was discussed in this presentation, with reference to Suncor Energy's primary and support equipment fleets. This paper also discussed Suncor Energy's maintenance and reliability standard involving people, processes and technology. An organizational maturity chart that graphed organizational learning against organizational performance was illustrated. The presentation also reviewed the maintenance and reliability framework; maintenance reliability model; the process overview of the maintenance and reliability standard; a process flow chart of maintenance strategies and programs; and an asset reliability improvement process flow chart. An example of an improvement initiative was included, with reference to a shovel reliability review; a dipper trip reliability investigation; bucket related failures by type and frequency; root cause analysis of the reliability process; and additional actions taken. Last, the presentation provided a graph of the results of the improvement initiative and presented the key lessons learned. tabs., figs.

  17. Receptor-isoform-selective insulin analogues give tissue-preferential effects

    DEFF Research Database (Denmark)

    Vienberg, Sara Gry; Bouman, Stephan D; Sørensen, Heidi

    2011-01-01

    The relative expression patterns of the two IR (insulin receptor) isoforms, +/- exon 11 (IR-B/IR-A respectively), are tissue-dependent. Therefore we have developed insulin analogues with different binding affinities for the two isoforms to test whether tissue-preferential biological effects can...... be attained. In rats and mice, IR-B is the most prominent isoform in the liver (> 95%) and fat (> 90%), whereas in muscles IR-A is the dominant isoform (> 95%). As a consequence, the insulin analogue INS-A, which has a higher relative affinity for human IR-A, had a higher relative potency [compared with HI...... (human insulin)] for glycogen synthesis in rat muscle strips (26%) than for glycogen accumulation in rat hepatocytes (5%) and for lipogenesis in rat adipocytes (4%). In contrast, the INS-B analogue, which has an increased affinity for human IR-B, had higher relative potencies (compared with HI...

  18. The Structure and Function of the Na,K-ATPase Isoforms in Health and Disease

    Directory of Open Access Journals (Sweden)

    Michael V. Clausen

    2017-06-01

    Full Text Available The sodium and potassium gradients across the plasma membrane are used by animal cells for numerous processes, and the range of demands requires that the responsible ion pump, the Na,K-ATPase, can be fine-tuned to the different cellular needs. Therefore, several isoforms are expressed of each of the three subunits that make a Na,K-ATPase, the alpha, beta and FXYD subunits. This review summarizes the various roles and expression patterns of the Na,K-ATPase subunit isoforms and maps the sequence variations to compare the differences structurally. Mutations in the Na,K-ATPase genes encoding alpha subunit isoforms have severe physiological consequences, causing very distinct, often neurological diseases. The differences in the pathophysiological effects of mutations further underline how the kinetic parameters, regulation and proteomic interactions of the Na,K-ATPase isoforms are optimized for the individual cellular needs.

  19. The Structure and Function of the Na,K-ATPase Isoforms in Health and Disease.

    Science.gov (United States)

    Clausen, Michael V; Hilbers, Florian; Poulsen, Hanne

    2017-01-01

    The sodium and potassium gradients across the plasma membrane are used by animal cells for numerous processes, and the range of demands requires that the responsible ion pump, the Na,K-ATPase, can be fine-tuned to the different cellular needs. Therefore, several isoforms are expressed of each of the three subunits that make a Na,K-ATPase, the alpha, beta and FXYD subunits. This review summarizes the various roles and expression patterns of the Na,K-ATPase subunit isoforms and maps the sequence variations to compare the differences structurally. Mutations in the Na,K-ATPase genes encoding alpha subunit isoforms have severe physiological consequences, causing very distinct, often neurological diseases. The differences in the pathophysiological effects of mutations further underline how the kinetic parameters, regulation and proteomic interactions of the Na,K-ATPase isoforms are optimized for the individual cellular needs.

  20. An abnormally glycosylated isoform of erythropoietin in hemangioblastoma is associated with polycythemia.

    Science.gov (United States)

    Delanghe, Sigurd E; Dierick, Jan; Maenhout, Thomas M; Zabeau, Lennart; Tavernier, Jan; Claes, Kathleen; Bleyen, Joris; Delanghe, Joris R

    2015-01-01

    Hemangioblastomas express erythropoietin and the patients often present with polycythemia. Serum erythropoietin was measured using a commercial immunoassay, a functional erythropoietin assay and iso-electric focusing. Despite the polycythemia, serum erythropoietin remained low, while a functional erythropoietin-assay showed a 4-5 higher activity in serum compared to the immunoassay. Iso-electric focusing of serum erythropoietin indicated overrepresentation of highly sialylated erythropoietin isoforms produced by the tumor. As a result, altered affinity of the monoclonal antibody used in the immunoassay for the hypersialylated isoforms was suggested. Analysis of erythropoietin isoforms may be helpful in distinguishing the ectopic erythropoietin isoforms from normally glycosylated erythropoietin. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Altered β-Amyloid Precursor Protein Isoforms in Mexican Alzheimer’s Disease Patients

    Directory of Open Access Journals (Sweden)

    V. J. Sánchez-González

    2006-01-01

    Full Text Available Objective: To determine the β-amyloid precursor protein (βAPP isoforms ratio as a risk factor for Alzheimer’s Disease and to assess its relationship with demographic and genetic variables of the disease.

  2. Enhanced protein electrophoresis technique for separating human skeletal muscle myosin heavy chain isoforms

    Science.gov (United States)

    Bamman, M. M.; Clarke, M. S.; Talmadge, R. J.; Feeback, D. L.

    1999-01-01

    Talmadge and Roy (J. Appl. Physiol. 1993, 75, 2337-2340) previously established a sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE) protocol for separating all four rat skeletal muscle myosin heavy chain (MHC) isoforms (MHC I, IIa, IIx, IIb); however, when applied to human muscle, the type II MHC isoforms (Ila, IIx) are not clearly distinguished. In this brief paper we describe a modification of the SDS-PAGE protocol which yields distinct and consistent separation of all three adult human MHC isoforms (MHC I, IIa, IIx) in a minigel system. MHC specificity of each band was confirmed by Western blot using three monoclonal IgG antibodies (mAbs) immunoreactive against MHCI (mAb MHCs, Novacastra Laboratories), MHCI+IIa (mAb BF-35), and MHCIIa+IIx (mAb SC-71). Results provide a valuable SDS-PAGE minigel technique for separating MHC isoforms in human muscle without the difficult task of casting gradient gels.

  3. Development of isoform-specific sensors of polypeptide GalNAc-transferase activity

    DEFF Research Database (Denmark)

    Song, Lina; Bachert, Collin; Schjoldager, Katrine T

    2014-01-01

    sequence influenced their activity and required modification, which we carried out based on previous in vitro work. Significantly, the modified T2 and T3 sensors were activated only in cells lacking their corresponding isozymes. Thus, we have developed T2- and T3-specific sensors that will be valuable......Humans express up to 20 isoforms of GalNAc-transferase (herein T1-T20) that localize to the Golgi apparatus and initiate O-glycosylation. Regulation of this enzyme family affects a vast array of proteins transiting the secretory pathway and diseases arise upon misregulation of specific isoforms....... Surprisingly, molecular probes to monitor GalNAc-transferase activity are lacking and there exist no effective global or isoform-specific inhibitors. Here we describe the development of T2- and T3-isoform specific fluorescence sensors that traffic in the secretory pathway. Each sensor yielded little signal...

  4. The Accelerator Reliability Forum

    CERN Document Server

    Lüdeke, Andreas; Giachino, R

    2014-01-01

    A high reliability is a very important goal for most particle accelerators. The biennial Accelerator Reliability Workshop covers topics related to the design and operation of particle accelerators with a high reliability. In order to optimize the over-all reliability of an accelerator one needs to gather information on the reliability of many different subsystems. While a biennial workshop can serve as a platform for the exchange of such information, the authors aimed to provide a further channel to allow for a more timely communication: the Particle Accelerator Reliability Forum [1]. This contribution will describe the forum and advertise it’s usage in the community.

  5. Enhanced expression of two discrete isoforms of matrix metalloproteinase-2 in experimental and human diabetic nephropathy.

    Directory of Open Access Journals (Sweden)

    Sang Soo Kim

    Full Text Available We recently reported on the enhanced expression of two isoforms of matrix metalloproteinase-2 (MMP-2 in human renal transplantation delayed graft function. These consist of the conventional secreted, full length MMP-2 isoform (FL-MMP-2 and a novel intracellular N-Terminal Truncated isoform (NTT-MMP-2 generated by oxidative stress-mediated activation of an alternate promoter in the MMP-2 first intron. Here we evaluated the effect of hyperglycemia and diabetes mellitus on the in vitro and in vivo expression of the two MMP-2 isoforms.We quantified the abundance of the FL-MMP-2 and NTT-MMP-2 transcripts by qPCR in HK2 cells cultured in high glucose or 4-hydroxy-2-hexenal (HHE and tested the effects of the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC. The streptozotocin (STZ murine model of Type I diabetes mellitus and renal biopsies of human diabetic nephropathy were used in this study.Both isoforms of MMP-2 in HK2 cells were upregulated by culture in high glucose or with HHE. PDTC treatment did not suppress high glucose-mediated FL-MMP-2 expression but potently inhibited NTT-MMP-2 expression. With STZ-treated mice, renal cortical expression of both isoforms was increased (FL-MMP-2, 1.8-fold; NTT-MMP-2, greater than 7-fold. Isoform-specific immunohistochemical staining revealed low, but detectable levels of the FL-MMP-2 isoform in controls, while NTT-MMP-2 was not detected. While there was a modest increase in tubular epithelial cell staining for FL-MMP-2 in STZ-treated mice, NTT-MMP-2 was intensely expressed in a basolateral pattern. FL-MMP-2 and NTT-MMP-2 isoform expression as quantified by qPCR were both significantly elevated in renal biopsies of human diabetic nephropathy (12-fold and 3-fold, respectively.The expression of both isoforms of MMP-2 was enhanced in an experimental model of diabetic nephropathy and in human diabetic nephropathy. Selective MMP-2 isoform inhibition could offer a novel approach for the treatment of diabetic renal

  6. Each individual isoform of the dopamine D2 receptor protects from lactotroph hyperplasia.

    Science.gov (United States)

    Radl, Daniela; De Mei, Claudia; Chen, Eric; Lee, Hyuna; Borrelli, Emiliana

    2013-06-01

    Dopamine acting through D2 receptors (D2Rs) controls lactotroph proliferation and prolactin (PRL) levels. Ablation of this receptor in mice results in lactotroph hyperplasia and prolactinomas in aged females. Alternative splicing of the Drd2 gene generates 2 independent isoforms, a long (D2L) and a short (D2S) isoform, which are present in all D2R-expressing cells. Here, we addressed the role of D2L and D2S on lactotroph physiology through the generation and analysis of D2S-null mice and their comparison with D2L-null animals. These mice represent a valuable tool with which to investigate dopamine-dependent isoform-specific signaling in the pituitary gland. We sought to assess the existence of a more prominent role of D2L or D2S in controlling PRL expression and lactotroph hyperplasia. Importantly, we found that D2L and D2S are specifically linked to independent transduction pathways in the pituitary. D2L-mediated signaling inhibits the AKT/protein kinase B kinase activity whereas D2S, in contrast, is required for the activation of the ERK 1/2 pathway. Under normal conditions, presence of only 1 of the 2 D2R isoforms in vivo prevents hyperprolactinemia, formation of lactotroph's hyperplasia, and tumorigenesis that is observed when both isoforms are deleted as in D2R-/- mice. However, the protective function of the single D2R isoforms is overridden when single isoform-knockout mice are challenged by chronic estrogen treatments as they show increased PRL production and lactotroph hyperplasia. Our study indicates that signaling from each of the D2R isoforms is sufficient to maintain lactotroph homeostasis in physiologic conditions; however, signaling from both is necessary in conditions simulating pathologic states.

  7. Genomic organization and the tissue distribution of alternatively spliced isoforms of the mouse Spatial gene

    Directory of Open Access Journals (Sweden)

    Mattei Marie-Geneviève

    2004-07-01

    Full Text Available Abstract Background The stromal component of the thymic microenvironment is critical for T lymphocyte generation. Thymocyte differentiation involves a cascade of coordinated stromal genes controlling thymocyte survival, lineage commitment and selection. The "Stromal Protein Associated with Thymii And Lymph-node" (Spatial gene encodes a putative transcription factor which may be involved in T-cell development. In the testis, the Spatial gene is also expressed by round spermatids during spermatogenesis. Results The Spatial gene maps to the B3-B4 region of murine chromosome 10 corresponding to the human syntenic region 10q22.1. The mouse Spatial genomic DNA is organised into 10 exons and is alternatively spliced to generate two short isoforms (Spatial-α and -γ and two other long isoforms (Spatial-δ and -ε comprising 5 additional exons on the 3' site. Here, we report the cloning of a new short isoform, Spatial-β, which differs from other isoforms by an additional alternative exon of 69 bases. This new exon encodes an interesting proline-rich signature that could confer to the 34 kDa Spatial-β protein a particular function. By quantitative TaqMan RT-PCR, we have shown that the short isoforms are highly expressed in the thymus while the long isoforms are highly expressed in the testis. We further examined the inter-species conservation of Spatial between several mammals and identified that the protein which is rich in proline and positive amino acids, is highly conserved. Conclusions The Spatial gene generates at least five alternative spliced variants: three short isoforms (Spatial-α, -β and -γ highly expressed in the thymus and two long isoforms (Spatial-δ and -ε highly expressed in the testis. These alternative spliced variants could have a tissue specific function.

  8. Electrophoretic Mobility of Cardiac Myosin Heavy Chain Isoforms Revisited: Application of MALDI TOF/TOF Analysis

    Czech Academy of Sciences Publication Activity Database

    Arnoštová, P.; Jedelsky, P. L.; Soukup, Tomáš; Žurmanová, J.

    2011-01-01

    Roč. 2011, - (2011), e634253 ISSN 1110-7243 R&D Projects: GA AV ČR IAAX01110901; GA ČR(CZ) GA304/08/0256 Institutional research plan: CEZ:AV0Z50110509 Keywords : cardiac MyHC isoforms * MyHC isoform mobility * effect of thyroid hormones * mass spectrometry * SDS-PAGE and western blot Subject RIV: ED - Physiology Impact factor: 2.436, year: 2011

  9. Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk

    DEFF Research Database (Denmark)

    Jacobsen, Stine; Niewold, T.A.; Kornalijnslijper, E.

    2005-01-01

    The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically pruduced SAA isoforms in plasma and milk fra cows with mastitis.......The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically pruduced SAA isoforms in plasma and milk fra cows with mastitis....

  10. Differences in sialic acid residues among bone alkaline phosphatase isoforms: a physical, biochemical, and immunological characterization.

    Science.gov (United States)

    Magnusson, P; Farley, J R

    2002-12-01

    High-performance liquid chromatography (HPLC) separates three human bone alkaline phosphatase (BALP) isoforms in serum; two major BALP isoforms, B1 and B2, and a minor fraction, B/I, which is composed on average of 70% bone and 30% intestinal ALP. The current studies were intended to identify an in vitro source of the BALP isoforms for physical, biochemical, and immunological characterizations. The three BALP isoforms were identified in extracts of human osteosarcoma (SaOS-2) cells, by HPLC, after separation by anion-exchange chromatography. All three BALP isoforms were similar with respect to freeze-thaw stability, solubility, heat inactivation, and inhibition by L-phenylalanine, L-homoarginine, and levamisole. The isoforms were also kinetically similar (i.e., maximal velocity and KM at pH 8.8 and pH 10.0). The isoforms differed, however, with respect to sensitivity to precipitation with wheat germ agglutinin (WGA), P acid residues was estimated to be 29 and 45, for each B1 and B2 homodimer, respectively. Apparent discrepancies between these estimates of molecular weight and estimates based on gel filtration chromatography were attributed to nonspecific interactions between carbohydrate residues and the gel filtration beads. All three BALP isoforms showed similar dose-dependent linearity in the commercial Alkphase-B and Tandem-MP Ostase immunoassays, r = 0.944 and r = 0.985, respectively (P acid residues compared with B/I, which mainly explains the apparent differences in molecular weight. Future investigations will focus on the clinical and functional significance of the revealed differences in sialic acid residues.

  11. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution.

    Directory of Open Access Journals (Sweden)

    Niels Jacob Aachmann-Andersen

    Full Text Available The membrane-assisted isoform immunoassay (MAIIA quantitates erythropoietin (EPO isoforms as percentages of migrated isoforms (PMI. We evaluated the effect of recombinant human EPO (rhEPO on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13; high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13; or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3 % (mean (SD. High-dose Epoetin beta decreased PMI on days 4 and 11 to 31.0 (4.2% (p<0.00001 and 45.2 (7.3% (p<0.00001. Low-dose Epoetin beta decreased PMI on days 4 and 11 to 46.0 (12.8% (p<0.00001 and 46.1 (10.4% (p<0.00001. In both rhEPO groups, PMI on day 25 was still decreased (high-dose Epoetin beta: 72.9 (19.4% (p=0.029; low-dose Epoetin beta: 73.1 (17.8% (p=0.039. In conclusion, Epoetin beta leaves a footprint in the plasma-EPO isoform pattern. MAIIA can detect changes in EPO isoform distribution up til at least three weeks after administration of Epoetin beta even though the total EPO concentration has returned to normal.

  12. Differential expression of mRNAs for protein kinase inhibitor isoforms in mouse brain.

    OpenAIRE

    Seasholtz, A F; Gamm, D M; Ballestero, R P; Scarpetta, M A; Uhler, M D

    1995-01-01

    Many neurotransmitters are known to regulate neuronal cell function by means of activation of cAMP-dependent protein kinase (PKA) and phosphorylation of neuronal substrate proteins, including transcription factors and ion channels. Here, we have characterized the gene expression of two isoforms of a protein kinase inhibitor (PKI) specific for PKA in mouse brain by RNase protection and in situ hybridization histochemistry. The studies demonstrate that the PKI alpha isoform is abundant in many ...

  13. Myosin heavy-chain isoforms in the flight and leg muscles of hummingbirds and zebra finches.

    Science.gov (United States)

    Velten, Brandy P; Welch, Kenneth C

    2014-06-01

    Myosin heavy chain (MHC) isoform complement is intimately related to a muscle's contractile properties, yet relatively little is known about avian MHC isoforms or how they may vary with fiber type and/or the contractile properties of a muscle. The rapid shortening of muscles necessary to power flight at the high wingbeat frequencies of ruby-throated hummingbirds and zebra finches (25-60 Hz), along with the varied morphology and use of the hummingbird hindlimb, provides a unique opportunity to understand how contractile and morphological properties of avian muscle may be reflected in MHC expression. Isoforms of the hummingbird and zebra finch flight and hindlimb muscles were electrophoretically separated and compared with those of other avian species representing different contractile properties and fiber types. The flight muscles of the study species operate at drastically different contraction rates and are composed of different histochemically defined fiber types, yet each exhibited the same, single MHC isoform corresponding to the chicken adult fast isoform. Thus, despite quantitative differences in the contractile demands of flight muscles across species, this isoform appears necessary for meeting the performance demands of avian powered flight. Variation in flight muscle contractile performance across species may be due to differences in the structural composition of this conserved isoform and/or variation within other mechanically linked proteins. The leg muscles were more varied in their MHC isoform composition across both muscles and species. The disparity in hindlimb MHC expression between hummingbirds and the other species highlights previously observed differences in fiber type composition and thrust production during take-off. Copyright © 2014 the American Physiological Society.

  14. Robust stratification of breast cancer subtypes using differential patterns of transcript isoform expression.

    Directory of Open Access Journals (Sweden)

    Thomas P Stricker

    2017-03-01

    Full Text Available Breast cancer, the second leading cause of cancer death of women worldwide, is a heterogenous disease with multiple different subtypes. These subtypes carry important implications for prognosis and therapy. Interestingly, it is known that these different subtypes not only have different biological behaviors, but also have distinct gene expression profiles. However, it has not been rigorously explored whether particular transcriptional isoforms are also differentially expressed among breast cancer subtypes, or whether transcript isoforms from the same sets of genes can be used to differentiate subtypes. To address these questions, we analyzed the patterns of transcript isoform expression using a small set of RNA-sequencing data for eleven Estrogen Receptor positive (ER+ subtype and fourteen triple negative (TN subtype tumors. We identified specific sets of isoforms that distinguish these tumor subtypes with higher fidelity than standard mRNA expression profiles. We found that alternate promoter usage, alternative splicing, and alternate 3'UTR usage are differentially regulated in breast cancer subtypes. Profiling of isoform expression in a second, independent cohort of 68 tumors confirmed that expression of splice isoforms differentiates breast cancer subtypes. Furthermore, analysis of RNAseq data from 594 cases from the TCGA cohort confirmed the ability of isoform usage to distinguish breast cancer subtypes. Also using our expression data, we identified several RNA processing factors that were differentially expressed between tumor subtypes and/or regulated by estrogen receptor, including YBX1, YBX2, MAGOH, MAGOHB, and PCBP2. RNAi knock-down of these RNA processing factors in MCF7 cells altered isoform expression. These results indicate that global dysregulation of splicing in breast cancer occurs in a subtype-specific and reproducible manner and is driven by specific differentially expressed RNA processing factors.

  15. Prospective comparison of liver stiffness measurements between two point wave elastography methods: Virtual ouch quantification and elastography point quantification

    Energy Technology Data Exchange (ETDEWEB)

    Yoo, Hyun Suk; Lee, Jeong Min; Yoon, Jeong Hee; Lee, Dong Ho; Chang, Won; Han, Joon Koo [Seoul National University Hospital, Seoul (Korea, Republic of)

    2016-09-15

    To prospectively compare technical success rate and reliable measurements of virtual touch quantification (VTQ) elastography and elastography point quantification (ElastPQ), and to correlate liver stiffness (LS) measurements obtained by the two elastography techniques. Our study included 85 patients, 80 of whom were previously diagnosed with chronic liver disease. The technical success rate and reliable measurements of the two kinds of point shear wave elastography (pSWE) techniques were compared by χ{sup 2} analysis. LS values measured using the two techniques were compared and correlated via Wilcoxon signed-rank test, Spearman correlation coefficient, and 95% Bland-Altman limit of agreement. The intraobserver reproducibility of ElastPQ was determined by 95% Bland-Altman limit of agreement and intraclass correlation coefficient (ICC). The two pSWE techniques showed similar technical success rate (98.8% for VTQ vs. 95.3% for ElastPQ, p = 0.823) and reliable LS measurements (95.3% for VTQ vs. 90.6% for ElastPQ, p = 0.509). The mean LS measurements obtained by VTQ (1.71 ± 0.47 m/s) and ElastPQ (1.66 ± 0.41 m/s) were not significantly different (p = 0.209). The LS measurements obtained by the two techniques showed strong correlation (r = 0.820); in addition, the 95% limit of agreement of the two methods was 27.5% of the mean. Finally, the ICC of repeat ElastPQ measurements was 0.991. Virtual touch quantification and ElastPQ showed similar technical success rate and reliable measurements, with strongly correlated LS measurements. However, the two methods are not interchangeable due to the large limit of agreement.

  16. Quantification of uranyl in presence of citric acid

    International Nuclear Information System (INIS)

    Garcia G, N.; Barrera D, C.E.; Ordonez R, E.

    2007-01-01

    To determine the influence that has the organic matter of the soil on the uranyl sorption on some solids is necessary to have a detection technique and quantification of uranyl that it is reliable and sufficiently quick in the obtaining of results. For that in this work, it intends to carry out the uranyl quantification in presence of citric acid modifying the Fluorescence induced by UV-Vis radiation technique. Since the uranyl ion is very sensitive to the medium that contains it, (speciation, pH, ionic forces, etc.) it was necessary to develop an analysis technique that stands out the fluorescence of uranyl ion avoiding the out one that produce the organic acids. (Author)

  17. C/EBPβ Isoforms Expression in the Rat Brain during the Estrous Cycle

    Directory of Open Access Journals (Sweden)

    Valeria Hansberg-Pastor

    2015-01-01

    Full Text Available The CCAAT/enhancer-binding protein beta (C/EBPβ is a transcription factor expressed in different areas of the brain that regulates the expression of several genes involved in cell differentiation and proliferation. This protein has three isoforms (LAP1, LAP2, and LIP with different transcription activation potential. The role of female sex hormones in the expression pattern of C/EBPβ isoforms in the rat brain has not yet been described. In this study we demonstrate by western blot that the expression of the three C/EBPβ isoforms changes in different brain areas during the estrous cycle. In the cerebellum, LAP2 content diminished on diestrus and proestrus and LIP content diminished on proestrus and estrus days. In the prefrontal cortex, LIP content was higher on proestrus and estrus days. In the hippocampus, LAP isoforms presented a switch on diestrus day, since LAP1 content was the highest while that of LAP2 was the lowest. The LAP2 isoform was the most abundant one in all the three brain areas. The LAP/LIP ratio changed throughout the cycle and was tissue specific. These results suggest that C/EBPβ isoforms expression changes in a tissue-specific manner in the rat brain due to the changes in sex steroid hormone levels presented during the estrous cycle.

  18. Proliferation marker pKi-67 occurs in different isoforms with various cellular effects.

    Science.gov (United States)

    Schmidt, Mirko H H; Broll, Rainer; Bruch, Hans-Peter; Finniss, Susan; Bögler, Oliver; Duchrow, Michael

    2004-04-15

    The Ki-67 antigen, pKi-67, is a commonly used proliferation marker in research and pathology. It has been recognized that the protein exists in two different splice variants that differ in one exon. In the current work, we present three new splice variants of human pKi-67 consisting of two naturally occurring isoforms and one atypical version. Additionally, data is presented indicating that alternative splicing of the pKi-67 N-terminus is common in tumor cell lines. Analyzing 93 tissues mainly consisting of brain tumor specimens, we found evidence that long and short isoform can be expressed independently of each other. Induction of mitosis in human peripheral blood mononuclear cells revealed that short pKi-67 appears earlier in the cell cycle than the long isoform and reaches its expression maximum when transcription of the latter sets in. Finally, transfection of mammalian culture cells with exon 7 (specific for the long pKi-67 isoform and not present in the short isoform) in a tetracycline regulated expression system decreased the rate of cell proliferation without affecting the cell cycle. In summary, we present evidence that the pKi-67 N-terminus is differentially spliced resulting in at least five different isoforms with different functions. Copyright 2004 Wiley-Liss, Inc.

  19. Developmental changes in circulating IL-8/CXCL8 isoforms in neonates.

    Science.gov (United States)

    Maheshwari, Akhil; Voitenok, Nikolai N; Akalovich, Svetlana; Shaik, Sadiq S; Randolph, David A; Sims, Brian; Patel, Rakesh P; Killingsworth, Cheryl R; Fallon, Michael B; Ohls, Robin K

    2009-04-01

    Interleukin-8 (IL-8/CXCL8) is widely expressed in fetal tissues although inflammatory changes are not seen. Circulating IL-8 is comprised of an endothelial-derived [ala-IL-8](77) isoform and another, more potent [ser-IL-8](72) secreted by most other cells; [ala-IL-8](77) can be converted into [ser-IL-8](72) by proteolytic removal of an N-terminal pentapeptide from [ala-IL-8](77). In this study, we show [ala-IL-8](77) is the predominant circulating isoform of IL-8 in premature neonates but not in term neonates/adults, who have [ser-IL-8](72) as the major isoform. This isoform switch from the less potent [ala-IL-8](77) to [ser-IL-8](72) correlates with a maturational increase in the neutrophil chemotactic potency of plasma IL-8. The emergence of [ser-IL-8](72) as the major isoform is likely due to increased plasma [ala-IL-8](77)-convertase activity and/or changes in the cellular sources of IL-8. Developmental changes in IL-8 isoforms may serve to minimize its inflammatory effects in the fetus and also provide a mechanism to restore its full activity after birth.

  20. Characterization of 14-3-3 isoforms expressed in the Echinococcus granulosus pathogenic larval stage.

    Science.gov (United States)

    Teichmann, Aline; Vargas, Daiani M; Monteiro, Karina M; Meneghetti, Bruna V; Dutra, Cristine S; Paredes, Rodolfo; Galanti, Norbel; Zaha, Arnaldo; Ferreira, Henrique B

    2015-04-03

    The 14-3-3 protein family of eukaryotic regulators was studied in Echinococcus granulosus, the causative agent of cystic hydatid disease. These proteins mediate important cellular processes in eukaryotes and are expected to play important roles in parasite biology. Six isoforms of E. granulosus 14-3-3 genes and proteins (Eg14-3-3.1-6) were analyzed, and their phylogenetic relationships were established with bona fide 14-3-3 orthologous proteins from eukaryotic species. Eg14-3-3 isoforms with previous evidence of expression (Eg14-3-3.1-4) in E. granulosus pathogenic larval stage (metacestode) were cloned, and recombinant proteins were used for functional studies. These protein isoforms were detected in different components of E. granulosus metacestode, including interface components with the host. The roles that are played by Eg14-3-3 proteins in parasite biology were inferred from the repertoires of interacting proteins with each isoform, as assessed by gel overlay, cross-linking, and affinity chromatography assays. A total of 95 Eg14-3-3 protein ligands were identified by mass spectrometry. Eg14-3-3 isoforms have shared partners (44 proteins), indicating some overlapping functions; however, they also bind exclusive partners (51 proteins), suggesting Eg14-3-3 functional specialization. These ligand repertoires indicate the involvement of Eg14-3-3 proteins in multiple biochemical pathways in the E. granulosus metacestode and note some degree of isoform specialization.

  1. Molecular modeling study on tunnel behavior in different histone deacetylase isoforms.

    Directory of Open Access Journals (Sweden)

    Sundarapandian Thangapandian

    Full Text Available Histone deacetylases (HDACs have emerged as effective therapeutic targets in the treatment of various diseases including cancers as these enzymes directly involved in the epigenetic regulation of genes. However the development of isoform-selective HDAC inhibitors has been a challenge till date since all HDAC enzymes possess conserved tunnel-like active site. In this study, using molecular dynamics simulation we have analyzed the behavior of tunnels present in HDAC8, 10, and 11 enzymes of class I, II, and IV, respectively. We have identified the equivalent tunnel forming amino acids in these three isoforms and found that they are very much conserved with subtle differences to be utilized in selective inhibitor development. One amino acid, methionine of HDAC8, among six tunnel forming residues is different in isoforms of other classes (glutamic acid (E in HDAC10 and leucine (L in HDAC 11 based on which mutations were introduced in HDAC11, the less studied HDAC isoform, to observe the effects of this change. The HDAC8-like (L268M mutation in the tunnel forming residues has almost maintained the deep and narrow tunnel as present in HDAC8 whereas HDAC10-like (L268E mutation has changed the tunnel wider and shallow as observed in HDAC10. These results explained the importance of the single change in the tunnel formation in different isoforms. The observations from this study can be utilized in the development of isoform-selective HDAC inhibitors.

  2. Glutamic acid decarboxylase isoform distribution in transgenic mouse septum: an anti-GFP immunofluorescence study.

    Science.gov (United States)

    Verimli, Ural; Sehirli, Umit S

    2016-09-01

    The septum is a basal forebrain region located between the lateral ventricles in rodents. It consists of lateral and medial divisions. Medial septal projections regulate hippocampal theta rhythm whereas lateral septal projections are involved in processes such as affective functions, memory formation, and behavioral responses. Gamma-aminobutyric acidergic neurons of the septal region possess the 65 and 67 isoforms of the enzyme glutamic acid decarboxylase. Although data on the glutamic acid decarboxylase isoform distribution in the septal region generally appears to indicate glutamic acid decarboxylase 67 dominance, different studies have given inconsistent results in this regard. The aim of this study was therefore to obtain information on the distributions of both of these glutamic acid decarboxylase isoforms in the septal region in transgenic mice. Two animal groups of glutamic acid decarboxylase-green fluorescent protein knock-in transgenic mice were utilized in the experiment. Brain sections from the region were taken for anti-green fluorescent protein immunohistochemistry in order to obtain estimated quantitative data on the number of gamma-aminobutyric acidergic neurons. Following the immunohistochemical procedures, the mean numbers of labeled cells in the lateral and medial septal nuclei were obtained for the two isoform groups. Statistical analysis yielded significant results which indicated that the 65 isoform of glutamic acid decarboxylase predominates in both lateral and medial septal nuclei (unpaired two-tailed t-test p glutamic acid decarboxylase isoform 65 in the septal region in glutamic acid decarboxylase-green fluorescent protein transgenic mice.

  3. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution

    DEFF Research Database (Denmark)

    Aachmann-Andersen, Niels Jacob; Just Christensen, Søren; Lisbjerg, Kristian

    2014-01-01

    The membrane-assisted isoform immunoassay (MAIIA) quantitates erythropoietin (EPO) isoforms as percentages of migrated isoforms (PMI). We evaluated the effect of recombinant human EPO (rhEPO) on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross......-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13); high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13); or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N......-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3) % (mean (SD)). High-dose Epoetin beta decreased PMI...

  4. MMP-2 Isoforms in Aortic Tissue and Serum of Patients with Ascending Aortic Aneurysms and Aortic Root Aneurysms

    Science.gov (United States)

    Tscheuschler, Anke; Meffert, Philipp; Beyersdorf, Friedhelm; Heilmann, Claudia; Kocher, Nadja; Uffelmann, Xenia; Discher, Philipp; Siepe, Matthias; Kari, Fabian A.

    2016-01-01

    Objective The need for biological markers of aortic wall stress and risk of rupture or dissection of ascending aortic aneurysms is obvious. To date, wall stress cannot be related to a certain biological marker. We analyzed aortic tissue and serum for the presence of different MMP-2 isoforms to find a connection between serum and tissue MMP-2 and to evaluate the potential of different MMP-2 isoforms as markers of high wall stress. Methods Serum and aortic tissue from n = 24 patients and serum from n = 19 healthy controls was analyzed by ELISA and gelatin zymography. 24 patients had ascending aortic aneurysms, 10 of them also had aortic root aneurysms. Three patients had normally functioning valves, 12 had regurgitation alone, eight had regurgitation and stenosis and one had only stenosis. Patients had bicuspid and tricuspid aortic valves (9/15). Serum samples were taken preoperatively, and the aortic wall specimen collected during surgical aortic repair. Results Pro-MMP-2 was identified in all serum and tissue samples. Pro-MMP-2 was detected in all tissue and serum samples from patients with ascending aortic/aortic root aneurysms, irrespective of valve morphology or other clinical parameters and in serum from healthy controls. We also identified active MMP-2 in all tissue samples from patients with ascending aortic/aortic root aneurysms. None of the analyzed serum samples revealed signals relatable to active MMP-2. No correlation between aortic tissue total MMP-2 or tissue pro-MMP-2 or tissue active MMP-2 and serum MMP-2 was found and tissue MMP-2/pro-MMP-2/active MMP-2 did not correlate with aortic diameter. This evidence shows that pro-MMP-2 is the predominant MMP-2 species in serum of patients and healthy individuals and in aneurysmatic aortic tissue, irrespective of aortic valve configuration. Active MMP-2 species are either not released into systemic circulation or not detectable in serum. There is no reliable connection between aortic tissue—and serum MMP-2

  5. Human Reliability Program Overview

    Energy Technology Data Exchange (ETDEWEB)

    Bodin, Michael

    2012-09-25

    This presentation covers the high points of the Human Reliability Program, including certification/decertification, critical positions, due process, organizational structure, program components, personnel security, an overview of the US DOE reliability program, retirees and academia, and security program integration.

  6. Power electronics reliability analysis.

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Mark A.; Atcitty, Stanley

    2009-12-01

    This report provides the DOE and industry with a general process for analyzing power electronics reliability. The analysis can help with understanding the main causes of failures, downtime, and cost and how to reduce them. One approach is to collect field maintenance data and use it directly to calculate reliability metrics related to each cause. Another approach is to model the functional structure of the equipment using a fault tree to derive system reliability from component reliability. Analysis of a fictitious device demonstrates the latter process. Optimization can use the resulting baseline model to decide how to improve reliability and/or lower costs. It is recommended that both electric utilities and equipment manufacturers make provisions to collect and share data in order to lay the groundwork for improving reliability into the future. Reliability analysis helps guide reliability improvements in hardware and software technology including condition monitoring and prognostics and health management.

  7. Reliability of software

    International Nuclear Information System (INIS)

    Kopetz, H.

    1980-01-01

    Common factors and differences in the reliability of hardware and software; reliability increase by means of methods of software redundancy. Maintenance of software for long term operating behavior. (HP) [de

  8. Quantification of heterogeneity observed in medical images

    International Nuclear Information System (INIS)

    Brooks, Frank J; Grigsby, Perry W

    2013-01-01

    There has been much recent interest in the quantification of visually evident heterogeneity within functional grayscale medical images, such as those obtained via magnetic resonance or positron emission tomography. In the case of images of cancerous tumors, variations in grayscale intensity imply variations in crucial tumor biology. Despite these considerable clinical implications, there is as yet no standardized method for measuring the heterogeneity observed via these imaging modalities. In this work, we motivate and derive a statistical measure of image heterogeneity. This statistic measures the distance-dependent average deviation from the smoothest intensity gradation feasible. We show how this statistic may be used to automatically rank images of in vivo human tumors in order of increasing heterogeneity. We test this method against the current practice of ranking images via expert visual inspection. We find that this statistic provides a means of heterogeneity quantification beyond that given by other statistics traditionally used for the same purpose. We demonstrate the effect of tumor shape upon our ranking method and find the method applicable to a wide variety of clinically relevant tumor images. We find that the automated heterogeneity rankings agree very closely with those performed visually by experts. These results indicate that our automated method may be used reliably to rank, in order of increasing heterogeneity, tumor images whether or not object shape is considered to contribute to that heterogeneity. Automated heterogeneity ranking yields objective results which are more consistent than visual rankings. Reducing variability in image interpretation will enable more researchers to better study potential clinical implications of observed tumor heterogeneity

  9. Quantification of heterogeneity observed in medical images.

    Science.gov (United States)

    Brooks, Frank J; Grigsby, Perry W

    2013-03-02

    There has been much recent interest in the quantification of visually evident heterogeneity within functional grayscale medical images, such as those obtained via magnetic resonance or positron emission tomography. In the case of images of cancerous tumors, variations in grayscale intensity imply variations in crucial tumor biology. Despite these considerable clinical implications, there is as yet no standardized method for measuring the heterogeneity observed via these imaging modalities. In this work, we motivate and derive a statistical measure of image heterogeneity. This statistic measures the distance-dependent average deviation from the smoothest intensity gradation feasible. We show how this statistic may be used to automatically rank images of in vivo human tumors in order of increasing heterogeneity. We test this method against the current practice of ranking images via expert visual inspection. We find that this statistic provides a means of heterogeneity quantification beyond that given by other statistics traditionally used for the same purpose. We demonstrate the effect of tumor shape upon our ranking method and find the method applicable to a wide variety of clinically relevant tumor images. We find that the automated heterogeneity rankings agree very closely with those performed visually by experts. These results indicate that our automated method may be used reliably to rank, in order of increasing heterogeneity, tumor images whether or not object shape is considered to contribute to that heterogeneity. Automated heterogeneity ranking yields objective results which are more consistent than visual rankings. Reducing variability in image interpretation will enable more researchers to better study potential clinical implications of observed tumor heterogeneity.

  10. Reliability of lifeline networks under seismic hazard

    International Nuclear Information System (INIS)

    Selcuk, A. Sevtap; Yuecemen, M. Semih

    1999-01-01

    Lifelines, such as pipelines, transportation, communication and power transmission systems, are networks which extend spatially over large geographical regions. The quantification of the reliability (survival probability) of a lifeline under seismic threat requires attention, as the proper functioning of these systems during or after a destructive earthquake is vital. In this study, a lifeline is idealized as an equivalent network with the capacity of its elements being random and spatially correlated and a comprehensive probabilistic model for the assessment of the reliability of lifelines under earthquake loads is developed. The seismic hazard that the network is exposed to is described by a probability distribution derived by using the past earthquake occurrence data. The seismic hazard analysis is based on the 'classical' seismic hazard analysis model with some modifications. An efficient algorithm developed by Yoo and Deo (Yoo YB, Deo N. A comparison of algorithms for terminal pair reliability. IEEE Transactions on Reliability 1988; 37: 210-215) is utilized for the evaluation of the network reliability. This algorithm eliminates the CPU time and memory capacity problems for large networks. A comprehensive computer program, called LIFEPACK is coded in Fortran language in order to carry out the numerical computations. Two detailed case studies are presented to show the implementation of the proposed model

  11. Reliable Design Versus Trust

    Science.gov (United States)

    Berg, Melanie; LaBel, Kenneth A.

    2016-01-01

    This presentation focuses on reliability and trust for the users portion of the FPGA design flow. It is assumed that the manufacturer prior to hand-off to the user tests FPGA internal components. The objective is to present the challenges of creating reliable and trusted designs. The following will be addressed: What makes a design vulnerable to functional flaws (reliability) or attackers (trust)? What are the challenges for verifying a reliable design versus a trusted design?

  12. Pocket Handbook on Reliability

    Science.gov (United States)

    1975-09-01

    exponencial distributions Weibull distribution, -xtimating reliability, confidence intervals, relia- bility growth, 0. P- curves, Bayesian analysis. 20 A S...introduction for those not familiar with reliability and a good refresher for those who are currently working in the area. LEWIS NERI, CHIEF...includes one or both of the following objectives: a) prediction of the current system reliability, b) projection on the system reliability for someI future

  13. Principles of Bridge Reliability

    DEFF Research Database (Denmark)

    Thoft-Christensen, Palle; Nowak, Andrzej S.

    The paper gives a brief introduction to the basic principles of structural reliability theory and its application to bridge engineering. Fundamental concepts like failure probability and reliability index are introduced. Ultimate as well as serviceability limit states for bridges are formulated......, and as an example the reliability profile and a sensitivity analyses for a corroded reinforced concrete bridge is shown....

  14. Reliability in engineering '87

    International Nuclear Information System (INIS)

    Tuma, M.

    1987-01-01

    The participants heard 51 papers dealing with the reliability of engineering products. Two of the papers were incorporated in INIS, namely ''Reliability comparison of two designs of low pressure regeneration of the 1000 MW unit at the Temelin nuclear power plant'' and ''Use of probability analysis of reliability in designing nuclear power facilities.''(J.B.)

  15. P120-catenin isoforms 1A and 3A differently affect invasion and proliferation of lung cancer cells

    International Nuclear Information System (INIS)

    Liu Yang; Dong Qianze; Zhao Yue; Dong Xinjun; Miao Yuan; Dai Shundong; Yang Zhiqiang; Zhang Di; Wang Yan; Li Qingchang; Zhao Chen; Wang Enhua

    2009-01-01

    Different isoforms of p120-catenin (p120ctn), a member of the Armadillo gene family, are variably expressed in different tissues as a result of alternative splicing and the use of multiple translation initiation codons. When expressed in cancer cells, these isoforms may confer different properties with respect to cell adhesion and invasion. We have previously reported that the p120ctn isoforms 1 and 3 were the most highly expressed isoforms in normal lung tissues, and their expression level was reduced in lung tumor cells. To precisely define their biological roles, we transfected p120ctn isoforms 1A and 3A into the lung cancer cell lines A549 and NCI-H460. Enhanced expression of p120ctn isoform 1A not only upregulated E-cadherin and β-catenin, but also downregulated the Rac1 activity, and as a result, inhibited the ability of cells to invade. In contrast, overexpression of p120ctn isoform 3A led to the inactivation of Cdc42 and the activation of RhoA, and had a smaller influence on invasion. However, we found that isoform 3A had a greater ability than isoform 1A in both inhibiting the cell cycle and reducing tumor cell proliferation. The present study revealed that p120ctn isoforms 1A and 3A differently regulated the adhesive, proliferative, and invasive properties of lung cancer cells through distinct mechanisms

  16. Prediction of software operational reliability using testing environment factor

    International Nuclear Information System (INIS)

    Jung, Hoan Sung

    1995-02-01

    Software reliability is especially important to customers these days. The need to quantify software reliability of safety-critical systems has been received very special attention and the reliability is rated as one of software's most important attributes. Since the software is an intellectual product of human activity and since it is logically complex, the failures are inevitable. No standard models have been established to prove the correctness and to estimate the reliability of software systems by analysis and/or testing. For many years, many researches have focused on the quantification of software reliability and there are many models developed to quantify software reliability. Most software reliability models estimate the reliability with the failure data collected during the test assuming that the test environments well represent the operation profile. User's interest is on the operational reliability rather than on the test reliability, however. The experiences show that the operational reliability is higher than the test reliability. With the assumption that the difference in reliability results from the change of environment, testing environment factor comprising the aging factor and the coverage factor are defined in this work to predict the ultimate operational reliability with the failure data. It is by incorporating test environments applied beyond the operational profile into testing environment factor Test reliability can also be estimated with this approach without any model change. The application results are close to the actual data. The approach used in this thesis is expected to be applicable to ultra high reliable software systems that are used in nuclear power plants, airplanes, and other safety-critical applications

  17. Expression of 14-3-3 protein isoforms in mouse oocytes, eggs and ovarian follicular development

    Directory of Open Access Journals (Sweden)

    De Santanu

    2012-01-01

    Full Text Available Abstract Background The 14-3-3 (YWHA proteins are a highly conserved, ubiquitously expressed family of proteins. Seven mammalian isoforms of 14-3-3 are known (β, γ, ε, ζ, η, τ and, σ. These proteins associate with many intracellular proteins involved in a variety of cellular processes including regulation of the cell cycle, metabolism and protein trafficking. We are particularly interested in the role of 14-3-3 in meiosis in mammalian eggs and the role 14-3-3 proteins may play in ovarian function. Therefore, we examined the expression of 14-3-3 proteins in mouse oocyte and egg extracts by Western blotting after polyacrylamide gel electrophoresis, viewed fixed cells by indirect immunofluorescence, and examined mouse ovarian cells by immunohistochemical staining to study the expression of the different 14-3-3 isoforms. Results We have determined that all of the mammalian 14-3-3 isoforms are expressed in mouse eggs and ovarian follicular cells including oocytes. Immunofluorescence confocal microscopy of isolated oocytes and eggs confirmed the presence of all of the isoforms with characteristic differences in some of their intracellular localizations. For example, some isoforms (β, ε, γ, and ζ are expressed more prominently in peripheral cytoplasm compared to the germinal vesicles in oocytes, but are uniformly dispersed within eggs. On the other hand, 14-3-3η is diffusely dispersed in the oocyte, but attains a uniform punctate distribution in the egg with marked accumulation in the region of the meiotic spindle apparatus. Immunohistochemical staining detected all isoforms within ovarian follicles, with some similarities as well as notable differences in relative amounts, localizations and patterns of expression in multiple cell types at various stages of follicular development. Conclusions We found that mouse oocytes, eggs and follicular cells within the ovary express all seven isoforms of the 14-3-3 protein. Examination of the

  18. DETECTION AND QUANTIFICATION OF COW FECAL POLLUTION WITH REAL-TIME PCR

    Science.gov (United States)

    Assessment of health risk and fecal bacteria loads associated with cow fecal pollution requires a reliable host-specific genetic marker and a rapid quantification method. We report the development of quantitative PCR assays for enumeration of two recently described cow-specific g...

  19. Reliable computer systems.

    Science.gov (United States)

    Wear, L L; Pinkert, J R

    1993-11-01

    In this article, we looked at some decisions that apply to the design of reliable computer systems. We began with a discussion of several terms such as testability, then described some systems that call for highly reliable hardware and software. The article concluded with a discussion of methods that can be used to achieve higher reliability in computer systems. Reliability and fault tolerance in computers probably will continue to grow in importance. As more and more systems are computerized, people will want assurances about the reliability of these systems, and their ability to work properly even when sub-systems fail.

  20. Human factor reliability program

    International Nuclear Information System (INIS)

    Knoblochova, L.

    2017-01-01

    The human factor's reliability program was at Slovenske elektrarne, a.s. (SE) nuclear power plants. introduced as one of the components Initiatives of Excellent Performance in 2011. The initiative's goal was to increase the reliability of both people and facilities, in response to 3 major areas of improvement - Need for improvement of the results, Troubleshooting support, Supporting the achievement of the company's goals. The human agent's reliability program is in practice included: - Tools to prevent human error; - Managerial observation and coaching; - Human factor analysis; -Quick information about the event with a human agent; -Human reliability timeline and performance indicators; - Basic, periodic and extraordinary training in human factor reliability(authors)

  1. Differential CARM1 Isoform Expression in Subcellular Compartments and among Malignant and Benign Breast Tumors.

    Directory of Open Access Journals (Sweden)

    David Shlensky

    Full Text Available Coactivator-associated arginine methyltransferase 1 (CARM1 is a coactivator for ERα and cancer-relevant transcription factors, and can methylate diverse cellular targets including histones. CARM1 is expressed in one of two alternative splice isoforms, full-length CARM1 (CARM1FL and truncated CARM1 (CARM1ΔE15. CARM1FL and CARM1ΔE15 function differently in transcriptional regulation, protein methylation, and mediation of pre-mRNA splicing in cellular models.To investigate the functional roles and the prognosis potential of CARM1 alternative spliced isoforms in breast cancer, we used recently developed antibodies to detect differential CARM1 isoform expression in subcellular compartments and among malignant and benign breast tumors.Immunofluorescence in MDA-MB-231 and BG-1 cell lines demonstrated that CARM1ΔE15 is the dominant isoform expressed in the cytoplasm, and CARM1FL is more nuclear localized. CARM1ΔE15 was found to be more sensitive to Hsp90 inhibition than CARM1FL, indicating that the truncated isoform may be the oncogenic form. Clinical cancer samples did not have significantly higher expression of CARM1FL or CARM1ΔE15 than benign breast samples at the level of mRNA or histology. Furthermore neither CARM1FL nor CARM1ΔE15 expression correlated with breast cancer molecular subtypes, tumor size, or lymph node involvement.The analysis presented here lends new insights into the possible oncogenic role of CARM1ΔE15. This study also demonstrates no obvious association of CARM1 isoform expression and clinical correlates in breast cancer. Recent studies, however, have shown that CARM1 expression correlates with poor prognosis, indicating a need for further studies of both CARM1 isoforms in a large cohort of breast cancer specimens.

  2. Drosophila TRPA1 isoforms detect UV light via photochemical production of H2O2

    Science.gov (United States)

    Guntur, Ananya R.; Gu, Pengyu; Takle, Kendra; Chen, Jingyi; Xiang, Yang; Yang, Chung-Hui

    2015-01-01

    The transient receptor potential A1 (TRPA1) channel is an evolutionarily conserved detector of temperature and irritant chemicals. Here, we show that two specific isoforms of TRPA1 in Drosophila are H2O2 sensitive and that they can detect strong UV light via sensing light-induced production of H2O2. We found that ectopic expression of these H2O2-sensitive Drosophila TRPA1 (dTRPA1) isoforms conferred UV sensitivity to light-insensitive HEK293 cells and Drosophila neurons, whereas expressing the H2O2-insensitive isoform did not. Curiously, when expressed in one specific group of motor neurons in adult flies, the H2O2-sensitive dTRPA1 isoforms were as competent as the blue light-gated channelrhodopsin-2 in triggering motor output in response to light. We found that the corpus cardiacum (CC) cells, a group of neuroendocrine cells that produce the adipokinetic hormone (AKH) in the larval ring gland endogenously express these H2O2-sensitive dTRPA1 isoforms and that they are UV sensitive. Sensitivity of CC cells required dTRPA1 and H2O2 production but not conventional phototransduction molecules. Our results suggest that specific isoforms of dTRPA1 can sense UV light via photochemical production of H2O2. We speculate that UV sensitivity conferred by these isoforms in CC cells may allow young larvae to activate stress response—a function of CC cells—when they encounter strong UV, an aversive stimulus for young larvae. PMID:26443856

  3. Gene duplication and the evolution of hemoglobin isoform differentiation in birds.

    Science.gov (United States)

    Grispo, Michael T; Natarajan, Chandrasekhar; Projecto-Garcia, Joana; Moriyama, Hideaki; Weber, Roy E; Storz, Jay F

    2012-11-02

    The majority of bird species co-express two functionally distinct hemoglobin (Hb) isoforms in definitive erythrocytes as follows: HbA (the major adult Hb isoform, with α-chain subunits encoded by the α(A)-globin gene) and HbD (the minor adult Hb isoform, with α-chain subunits encoded by the α(D)-globin gene). The α(D)-globin gene originated via tandem duplication of an embryonic α-like globin gene in the stem lineage of tetrapod vertebrates, which suggests the possibility that functional differentiation between the HbA and HbD isoforms may be attributable to a retained ancestral character state in HbD that harkens back to a primordial, embryonic function. To investigate this possibility, we conducted a combined analysis of protein biochemistry and sequence evolution to characterize the structural and functional basis of Hb isoform differentiation in birds. Functional experiments involving purified HbA and HbD isoforms from 11 different bird species revealed that HbD is characterized by a consistently higher O(2) affinity in the presence of allosteric effectors such as organic phosphates and Cl(-) ions. In the case of both HbA and HbD, analyses of oxygenation properties under the two-state Monod-Wyman-Changeux allosteric model revealed that the pH dependence of Hb-O(2) affinity stems primarily from changes in the O(2) association constant of deoxy (T-state)-Hb. Ancestral sequence reconstructions revealed that the amino acid substitutions that distinguish the adult-expressed Hb isoforms are not attributable to the retention of an ancestral (pre-duplication) character state in the α(D)-globin gene that is shared with the embryonic α-like globin gene.

  4. Gene Duplication and the Evolution of Hemoglobin Isoform Differentiation in Birds*

    Science.gov (United States)

    Grispo, Michael T.; Natarajan, Chandrasekhar; Projecto-Garcia, Joana; Moriyama, Hideaki; Weber, Roy E.; Storz, Jay F.

    2012-01-01

    The majority of bird species co-express two functionally distinct hemoglobin (Hb) isoforms in definitive erythrocytes as follows: HbA (the major adult Hb isoform, with α-chain subunits encoded by the αA-globin gene) and HbD (the minor adult Hb isoform, with α-chain subunits encoded by the αD-globin gene). The αD-globin gene originated via tandem duplication of an embryonic α-like globin gene in the stem lineage of tetrapod vertebrates, which suggests the possibility that functional differentiation between the HbA and HbD isoforms may be attributable to a retained ancestral character state in HbD that harkens back to a primordial, embryonic function. To investigate this possibility, we conducted a combined analysis of protein biochemistry and sequence evolution to characterize the structural and functional basis of Hb isoform differentiation in birds. Functional experiments involving purified HbA and HbD isoforms from 11 different bird species revealed that HbD is characterized by a consistently higher O2 affinity in the presence of allosteric effectors such as organic phosphates and Cl− ions. In the case of both HbA and HbD, analyses of oxygenation properties under the two-state Monod-Wyman-Changeux allosteric model revealed that the pH dependence of Hb-O2 affinity stems primarily from changes in the O2 association constant of deoxy (T-state)-Hb. Ancestral sequence reconstructions revealed that the amino acid substitutions that distinguish the adult-expressed Hb isoforms are not attributable to the retention of an ancestral (pre-duplication) character state in the αD-globin gene that is shared with the embryonic α-like globin gene. PMID:22962007

  5. Development and characterization of human monoclonal antibodies that neutralize multiple TGFβ isoforms.

    Science.gov (United States)

    Bedinger, Daniel; Lao, Llewelyn; Khan, Shireen; Lee, Steve; Takeuchi, Toshihiko; Mirza, Amer M

    2016-01-01

    Transforming growth factor (TGF)β levels are elevated in, and drive the progression of, numerous disease states such as advanced metastatic cancer and systemic and ocular fibrosis. There are 3 main isoforms, TGFβ1, 2, and 3. As multiple TGFβ isoforms are involved in disease processes, maximal therapeutic efficacy may require neutralization of 2 or more of the TGFβ isoforms. Fully human antibody phage display libraries were used to discover a number of antibodies that bind and neutralize various combinations of TGFβ1, 2 or 3. The primary panning did not yield any uniformly potent pan-isoform neutralizing antibodies; therefore, an antibody that displayed potent TGFβ 1, 2 inhibition, but more modest affinity versus TGFβ3, was affinity matured by shuffling with a light chain sub-library and further screening. This process yielded a high affinity pan-isoform neutralizing clone. Antibodies were analyzed and compared by binding affinity, as well as receptor and epitope competition by surface plasmon resonance methods. The antibodies were also shown to neutralize TGFβ effects in vitro in 3 assays: 1) interleukin (IL)-4 induced HT-2 cell proliferation; 2) TGFβ-mediated IL-11 release by A549 cells; and 3) decreasing SMAD2 phosphorylation in Detroit 562 cells. The antibodies' potency in these in vitro assays correlated well with their isoform-specific affinities. Furthermore, the ability of the affinity-matured clone to decrease tumor burden in a Detroit 562 xenograft study was superior to that of the parent clone. This affinity-matured antibody acts as a very potent inhibitor of all 3 main isoforms of TGFβ and may have utility for therapeutic intervention in human disease.

  6. Quantification of biofilm in microtiter plates: overview of testing conditions and practical recommendations for assessment of biofilm production by staphylococci.

    Science.gov (United States)

    Stepanović, Srdjan; Vuković, Dragana; Hola, Veronika; Di Bonaventura, Giovanni; Djukić, Slobodanka; Cirković, Ivana; Ruzicka, Filip

    2007-08-01

    The details of all steps involved in the quantification of biofilm formation in microtiter plates are described. The presented protocol incorporates information on assessment of biofilm production by staphylococci, gained both by direct experience as well as by analysis of methods for assaying biofilm production. The obtained results should simplify quantification of biofilm formation in microtiter plates, and make it more reliable and comparable among different laboratories.

  7. NEW MODEL FOR QUANTIFICATION OF ICT DEPENDABLE ORGANIZATIONS RESILIENCE

    Directory of Open Access Journals (Sweden)

    Zora Arsovski

    2011-03-01

    Full Text Available Business environment today demands high reliable organizations in every segment to be competitive on the global market. Beside that, ICT sector is becoming irreplaceable in many fields of business, from the communication to the complex systems for process control and production. To fulfill those requirements and to develop further, many organizations worldwide are implementing business paradigm called - organizations resilience. Although resilience is well known term in many science fields, it is not well studied due to its complex nature. This paper is dealing with developing the new model for assessment and quantification of ICT dependable organizations resilience.

  8. Uncertainty Quantification and Statistical Engineering for Hypersonic Entry Applications

    Science.gov (United States)

    Cozmuta, Ioana

    2011-01-01

    NASA has invested significant resources in developing and validating a mathematical construct for TPS margin management: a) Tailorable for low/high reliability missions; b) Tailorable for ablative/reusable TPS; c) Uncertainty Quantification and Statistical Engineering are valuable tools not exploited enough; and d) Need to define strategies combining both Theoretical Tools and Experimental Methods. The main reason for this lecture is to give a flavor of where UQ and SE could contribute and hope that the broader community will work with us to improve in these areas.

  9. Uncertainty quantification in ion–solid interaction simulations

    Energy Technology Data Exchange (ETDEWEB)

    Preuss, R., E-mail: preuss@ipp.mpg.de; Toussaint, U. von

    2017-02-15

    Within the framework of Bayesian uncertainty quantification we propose a non-intrusive reduced-order spectral approach (polynomial chaos expansion) to the simulation of ion–solid interactions. The method not only reduces the number of function evaluations but provides simultaneously a quantitative measure for which combinations of inputs have the most important impact on the result. It is applied to SDTRIM-simulations (Möller et al., 1988) with several uncertain and Gaussian distributed input parameters (i.e. angle, projectile energy, surface binding energy, target composition) and the results are compared to full-grid based approaches and sampling based methods with respect to reliability, efficiency and scalability.

  10. DNA imaging and quantification using chemi-luminescent probes

    International Nuclear Information System (INIS)

    Dorner, G.; Redjdal, N.; Laniece, P.; Siebert, R.; Tricoire, H.; Valentin, L.

    1999-01-01

    During this interdisciplinary study we have developed an ultra sensitive and reliable imaging system of DNA labelled by chemiluminescence. Based on a liquid nitrogen cooled CCD, the system achieves sensitivities down to 10 fg/mm 2 labelled DNA over a surface area of 25 x 25 cm 2 with a sub-millimeter resolution. Commercially available chemi-luminescent - and enhancer molecules are compared and their reaction conditions optimized for best signal-to-noise ratios. Double labelling was performed to verify quantification with radioactive probes. (authors)

  11. AtRTD2: A Reference Transcript Dataset for accurate quantification of alternative splicing and expression changes in Arabidopsis thaliana RNA-seq data

    KAUST Repository

    Zhang, Runxuan

    2016-05-06

    Background Alternative splicing is the major post-transcriptional mechanism by which gene expression is regulated and affects a wide range of processes and responses in most eukaryotic organisms. RNA-sequencing (RNA-seq) can generate genome-wide quantification of individual transcript isoforms to identify changes in expression and alternative splicing. RNA-seq is an essential modern tool but its ability to accurately quantify transcript isoforms depends on the diversity, completeness and quality of the transcript information. Results We have developed a new Reference Transcript Dataset for Arabidopsis (AtRTD2) for RNA-seq analysis containing over 82k non-redundant transcripts, whereby 74,194 transcripts originate from 27,667 protein-coding genes. A total of 13,524 protein-coding genes have at least one alternatively spliced transcript in AtRTD2 such that about 60% of the 22,453 protein-coding, intron-containing genes in Arabidopsis undergo alternative splicing. More than 600 putative U12 introns were identified in more than 2,000 transcripts. AtRTD2 was generated from transcript assemblies of ca. 8.5 billion pairs of reads from 285 RNA-seq data sets obtained from 129 RNA-seq libraries and merged along with the previous version, AtRTD, and Araport11 transcript assemblies. AtRTD2 increases the diversity of transcripts and through application of stringent filters represents the most extensive and accurate transcript collection for Arabidopsis to date. We have demonstrated a generally good correlation of alternative splicing ratios from RNA-seq data analysed by Salmon and experimental data from high resolution RT-PCR. However, we have observed inaccurate quantification of transcript isoforms for genes with multiple transcripts which have variation in the lengths of their UTRs. This variation is not effectively corrected in RNA-seq analysis programmes and will therefore impact RNA-seq analyses generally. To address this, we have tested different genome

  12. Collagen Quantification in Tissue Specimens.

    Science.gov (United States)

    Coentro, João Quintas; Capella-Monsonís, Héctor; Graceffa, Valeria; Wu, Zhuning; Mullen, Anne Maria; Raghunath, Michael; Zeugolis, Dimitrios I

    2017-01-01

    Collagen is the major extracellular protein in mammals. Accurate quantification of collagen is essential in the biomaterials (e.g., reproducible collagen scaffold fabrication), drug discovery (e.g., assessment of collagen in pathophysiologies, such as fibrosis), and tissue engineering (e.g., quantification of cell-synthesized collagen) fields. Although measuring hydroxyproline content is the most widely used method to quantify collagen in biological specimens, the process is very laborious. To this end, the Sircol™ Collagen Assay is widely used due to its inherent simplicity and convenience. However, this method leads to overestimation of collagen content due to the interaction of Sirius red with basic amino acids of non-collagenous proteins. Herein, we describe the addition of an ultrafiltration purification step in the process to accurately determine collagen content in tissues.

  13. Reliability and safety engineering

    CERN Document Server

    Verma, Ajit Kumar; Karanki, Durga Rao

    2016-01-01

    Reliability and safety are core issues that must be addressed throughout the life cycle of engineering systems. Reliability and Safety Engineering presents an overview of the basic concepts, together with simple and practical illustrations. The authors present reliability terminology in various engineering fields, viz.,electronics engineering, software engineering, mechanical engineering, structural engineering and power systems engineering. The book describes the latest applications in the area of probabilistic safety assessment, such as technical specification optimization, risk monitoring and risk informed in-service inspection. Reliability and safety studies must, inevitably, deal with uncertainty, so the book includes uncertainty propagation methods: Monte Carlo simulation, fuzzy arithmetic, Dempster-Shafer theory and probability bounds. Reliability and Safety Engineering also highlights advances in system reliability and safety assessment including dynamic system modeling and uncertainty management. Cas...

  14. Evolution of the cAMP-dependent protein kinase (PKA catalytic subunit isoforms.

    Directory of Open Access Journals (Sweden)

    Kristoffer Søberg

    Full Text Available The 3',5'-cyclic adenosine monophosphate (cAMP-dependent protein kinase, or protein kinase A (PKA, pathway is one of the most versatile and best studied signaling pathways in eukaryotic cells. The two paralogous PKA catalytic subunits Cα and Cβ, encoded by the genes PRKACA and PRKACB, respectively, are among the best understood model kinases in signal transduction research. In this work, we explore and elucidate the evolution of the alternative 5' exons and the splicing pattern giving rise to the numerous PKA catalytic subunit isoforms. In addition to the universally conserved Cα1/Cβ1 isoforms, we find kinase variants with short N-termini in all main vertebrate classes, including the sperm-specific Cα2 isoform found to be conserved in all mammals. We also describe, for the first time, a PKA Cα isoform with a long N-terminus, paralogous to the PKA Cβ2 N-terminus. An analysis of isoform-specific variation highlights residues and motifs that are likely to be of functional importance.

  15. Analysis of the synaptotagmin family during reconstituted membrane fusion. Uncovering a class of inhibitory isoforms.

    Science.gov (United States)

    Bhalla, Akhil; Chicka, Michael C; Chapman, Edwin R

    2008-08-01

    Ca(2+)-triggered exocytosis in neurons and neuroendocrine cells is regulated by the Ca(2+)-binding protein synaptotagmin (syt) I. Sixteen additional isoforms of syt have been identified, but little is known concerning their biochemical or functional properties. Here, we assessed the abilities of fourteen syt isoforms to directly regulate SNARE (soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor)-catalyzed membrane fusion. One group of isoforms stimulated neuronal SNARE-mediated fusion in response to Ca(2+), while another set inhibited SNARE catalyzed fusion in both the absence and presence of Ca(2+). Biochemical analysis revealed a strong correlation between the ability of syt isoforms to bind 1,2-dioleoyl phosphatidylserine (PS) and t-SNAREs in a Ca(2+)-promoted manner with their abilities to enhance fusion, further establishing PS and SNAREs as critical effectors for syt action. The ability of syt I to efficiently stimulate fusion was specific for certain SNARE pairs, suggesting that syts might contribute to the specificity of intracellular membrane fusion reactions. Finally, a subset of inhibitory syts down-regulated the ability of syt I to activate fusion, demonstrating that syt isoforms can modulate the function of each other.

  16. Elevated serum tartrate-resistant acid phosphatase isoform 5a levels in metabolic syndrome.

    Science.gov (United States)

    Huang, Yi-Jhih; Huang, Tsai-Wang; Chao, Tsu-Yi; Sun, Yu-Shan; Chen, Shyi-Jou; Chu, Der-Ming; Chen, Wei-Liang; Wu, Li-Wei

    2017-09-29

    Tartrate-resistant phosphatase isoform 5a is expressed in tumor-associated macrophages and is a biomarker of chronic inflammation. Herein, we correlated serum tartrate-resistant phosphatase isoform 5a levels with metabolic syndrome status and made comparisons with traditional markers of inflammation, including c-reactive protein and interleukin-6. One hundred healthy volunteers were randomly selected, and cut-off points for metabolic syndrome related inflammatory biomarkers were determined using receiver operating characteristic curves. Linear and logistic regression models were subsequently used to correlate inflammatory markers with the risk of metabolic syndrome. Twenty-two participants met the criteria for metabolic syndrome, and serum tartrate-resistant phosphatase isoform 5a levels of >5.8 μg/L were associated with metabolic syndrome (c-statistics, 0.730; p = 0.001; 95% confidence interval, 0.618-0.842). In addition, 1 μg/L increases in tartrate-resistant phosphatase isoform 5a levels were indicative of a 1.860 fold increase in the risk of metabolic syndrome (p = 0.012). Elevated serum tartrate-resistant phosphatase isoform 5a levels are associated with the risk of metabolic syndrome, with a cut-off level of 5.8 μg/L.

  17. Novel frataxin isoforms may contribute to the pathological mechanism of Friedreich ataxia.

    Directory of Open Access Journals (Sweden)

    Haiyan Xia

    Full Text Available Friedreich ataxia (FRDA is an inherited neurodegenerative disease caused by frataxin (FXN deficiency. The nervous system and heart are the most severely affected tissues. However, highly mitochondria-dependent tissues, such as kidney and liver, are not obviously affected, although the abundance of FXN is normally high in these tissues. In this study we have revealed two novel FXN isoforms (II and III, which are specifically expressed in affected cerebellum and heart tissues, respectively, and are functional in vitro and in vivo. Increasing the abundance of the heart-specific isoform III significantly increased the mitochondrial aconitase activity, while over-expression of the cerebellum-specific isoform II protected against oxidative damage of Fe-S cluster-containing aconitase. Further, we observed that the protein level of isoform III decreased in FRDA patient heart, while the mRNA level of isoform II decreased more in FRDA patient cerebellum compared to total FXN mRNA. Our novel findings are highly relevant to understanding the mechanism of tissue-specific pathology in FRDA.

  18. Nesprin-2 epsilon: A novel nesprin isoform expressed in human ovary and Ntera-2 cells

    International Nuclear Information System (INIS)

    Lam, Le Thanh; Boehm, Sabrina V.; Roberts, Roland G.; Morris, Glenn E.

    2011-01-01

    Highlights: → A novel epsilon isoform of nesprin-2 has been discovered. → This 120 kDa protein was predicted by bioinformatic analysis, but has not previously been observed. → It is the main isoform expressed in a teratocarcinoma cell line and is also found in ovary. → Like other nesprins, it is located at the nuclear envelope. → We suggest it may have a role in very early development or in some ovary-specific function. -- Abstract: The nuclear envelope-associated cytoskeletal protein, nesprin-2, is encoded by a large gene containing several internal promoters that produce shorter isoforms. In a study of Ntera-2 teratocarcinoma cells, a novel isoform, nesprin-2-epsilon, was found to be the major mRNA and protein product of the nesprin-2 gene. Its existence was predicted by bioinformatic analysis, but this is the first direct demonstration of both the mRNA and the 120 kDa protein which is located at the nuclear envelope. In a panel of 21 adult and foetal human tissues, the nesprin-2-epsilon mRNA was strongly expressed in ovary but was a minor isoform elsewhere. The expression pattern suggests a possible link with very early development and a likely physiological role in ovary.

  19. Discovery of novel isoforms of huntingtin reveals a new hominid-specific exon.

    Directory of Open Access Journals (Sweden)

    Albert Ruzo

    Full Text Available Huntington's disease (HD is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT. HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell are still not well understood. Scrutiny of HTT function has been focused on a single, full length mRNA. In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HTT-expanded human embryonic stem cell (hESC lines as well as in cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease.

  20. Discovery of Novel Isoforms of Huntingtin Reveals a New Hominid-Specific Exon

    Science.gov (United States)

    Popowski, Melissa; Haremaki, Tomomi; Croft, Gist F.; Deglincerti, Alessia; Brivanlou, Ali H.

    2015-01-01

    Huntington’s disease (HD) is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT). HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell are still not well understood. Scrutiny of HTT function has been focused on a single, full length mRNA. In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HTT-expanded human embryonic stem cell (hESC) lines as well as in cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease. PMID:26010866

  1. Functional divergence of platelet protein kinase C (PKC) isoforms in thrombus formation on collagen.

    Science.gov (United States)

    Gilio, Karen; Harper, Matthew T; Cosemans, Judith M E M; Konopatskaya, Olga; Munnix, Imke C A; Prinzen, Lenneke; Leitges, Michael; Liu, Qinghang; Molkentin, Jeffery D; Heemskerk, Johan W M; Poole, Alastair W

    2010-07-23

    Arterial thrombosis, a major cause of myocardial infarction and stroke, is initiated by activation of blood platelets by subendothelial collagen. The protein kinase C (PKC) family centrally regulates platelet activation, and it is becoming clear that the individual PKC isoforms play distinct roles, some of which oppose each other. Here, for the first time, we address all four of the major platelet-expressed PKC isoforms, determining their comparative roles in regulating platelet adhesion to collagen and their subsequent activation under physiological flow conditions. Using mouse gene knock-out and pharmacological approaches in human platelets, we show that collagen-dependent alpha-granule secretion and thrombus formation are mediated by the conventional PKC isoforms, PKCalpha and PKCbeta, whereas the novel isoform, PKC, negatively regulates these events. PKCdelta also negatively regulates thrombus formation but not alpha-granule secretion. In addition, we demonstrate for the first time that individual PKC isoforms differentially regulate platelet calcium signaling and exposure of phosphatidylserine under flow. Although platelet deficient in PKCalpha or PKCbeta showed reduced calcium signaling and phosphatidylserine exposure, these responses were enhanced in the absence of PKC. In summary therefore, this direct comparison between individual subtypes of PKC, by standardized methodology under flow conditions, reveals that the four major PKCs expressed in platelets play distinct non-redundant roles, where conventional PKCs promote and novel PKCs inhibit thrombus formation on collagen.

  2. Functional Divergence of Platelet Protein Kinase C (PKC) Isoforms in Thrombus Formation on Collagen*

    Science.gov (United States)

    Gilio, Karen; Harper, Matthew T.; Cosemans, Judith M. E. M.; Konopatskaya, Olga; Munnix, Imke C. A.; Prinzen, Lenneke; Leitges, Michael; Liu, Qinghang; Molkentin, Jeffery D.; Heemskerk, Johan W. M.; Poole, Alastair W.

    2010-01-01

    Arterial thrombosis, a major cause of myocardial infarction and stroke, is initiated by activation of blood platelets by subendothelial collagen. The protein kinase C (PKC) family centrally regulates platelet activation, and it is becoming clear that the individual PKC isoforms play distinct roles, some of which oppose each other. Here, for the first time, we address all four of the major platelet-expressed PKC isoforms, determining their comparative roles in regulating platelet adhesion to collagen and their subsequent activation under physiological flow conditions. Using mouse gene knock-out and pharmacological approaches in human platelets, we show that collagen-dependent α-granule secretion and thrombus formation are mediated by the conventional PKC isoforms, PKCα and PKCβ, whereas the novel isoform, PKCθ, negatively regulates these events. PKCδ also negatively regulates thrombus formation but not α-granule secretion. In addition, we demonstrate for the first time that individual PKC isoforms differentially regulate platelet calcium signaling and exposure of phosphatidylserine under flow. Although platelet deficient in PKCα or PKCβ showed reduced calcium signaling and phosphatidylserine exposure, these responses were enhanced in the absence of PKCθ. In summary therefore, this direct comparison between individual subtypes of PKC, by standardized methodology under flow conditions, reveals that the four major PKCs expressed in platelets play distinct non-redundant roles, where conventional PKCs promote and novel PKCs inhibit thrombus formation on collagen. PMID:20479008

  3. Proteomic Analysis of Parkin Isoforms Expression in Different Rat Brain Areas.

    Science.gov (United States)

    D'Amico, Agata Grazia; Maugeri, Grazia; Reitano, Rita; Cavallaro, Sebastiano; D'Agata, Velia

    2016-10-01

    PARK2 gene's mutations are related to the familial form of juvenile Parkinsonism, also known as the autosomic recessive juvenile Parkinsonism. This gene encodes for parkin, a 465-amino acid protein. To date, a large number of parkin isoforms, generated by an alternative splicing mechanism, have been described. Currently, Gene Bank lists 27 rat PARK2 transcripts, which matches to 20 exclusive parkin alternative splice variants. Despite the existence of these isoforms, most of the studies carried out so far, have been focused only on the originally cloned parkin. In this work we have analyzed the expression profile of parkin isoforms in some rat brain areas including prefrontal cortex, hippocampus, substantia nigra and cerebellum. To discriminate among these isoforms, we detected their localization through the use of two antibodies that are able to identify different domains of the parkin canonical sequence. Our analysis has revealed that at least fourteen parkin isoforms are expressed in rat brain with a various distribution in the regions analyzed. Our study might help to elucidate the pathophysiological role of these proteins in the central nervous system.

  4. Allosteric Mutant IDH1 Inhibitors Reveal Mechanisms for IDH1 Mutant and Isoform Selectivity

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Xiaoling; Baird, Daniel; Bowen, Kimberly; Capka, Vladimir; Chen, Jinyun; Chenail, Gregg; Cho, YoungShin; Dooley, Julia; Farsidjani, Ali; Fortin, Pascal; Kohls, Darcy; Kulathila, Raviraj; Lin, Fallon; McKay, Daniel; Rodrigues, Lindsey; Sage, David; Touré, B. Barry; van der Plas, Simon; Wright, Kirk; Xu, Ming; Yin, Hong; Levell, Julian; Pagliarini, Raymond A. (Novartis)

    2017-03-01

    Oncogenic IDH1 and IDH2 mutations contribute to cancer via production of R-2-hydroxyglutarate (2-HG). Here, we characterize two structurally distinct mutant- and isoform-selective IDH1 inhibitors that inhibit 2-HG production. Both bind to an allosteric pocket on IDH1, yet shape it differently, highlighting the plasticity of this site. Oncogenic IDH1R132H mutation destabilizes an IDH1 “regulatory segment,” which otherwise restricts compound access to the allosteric pocket. Regulatory segment destabilization in wild-type IDH1 promotes inhibitor binding, suggesting that destabilization is critical for mutant selectivity. We also report crystal structures of oncogenic IDH2 mutant isoforms, highlighting the fact that the analogous segment of IDH2 is not similarly destabilized. This intrinsic stability of IDH2 may contribute to observed inhibitor IDH1 isoform selectivity. Moreover, discrete residues in the IDH1 allosteric pocket that differ from IDH2 may also guide IDH1 isoform selectivity. These data provide a deeper understanding of how IDH1 inhibitors achieve mutant and isoform selectivity.

  5. Expression of a novel cardiac-specific tropomyosin isoform in humans

    International Nuclear Information System (INIS)

    Denz, Christopher R.; Narshi, Aruna; Zajdel, Robert W.; Dube, Dipak K.

    2004-01-01

    Tropomyosins are a family of actin binding proteins encoded by a group of highly conserved genes. Humans have four tropomyosin-encoding genes: TPM1, TPM2, TPM3, and TPM4, each of which is known to generate multiple isoforms by alternative splicing, promoters, and 3 ' end processing. TPM1 is the most versatile and encodes a variety of tissue specific isoforms. The TPM1 isoform specific to striated muscle, designated TPM1α, consists of 10 exons: 1a, 2b, 3, 4, 5, 6b, 7, 8, and 9a/b. In this study, using RT-PCR with adult and fetal human RNAs, we present evidence for the expression of a novel isoform of the TPM1 gene that is specifically expressed in cardiac tissues. The new isoform is designated TPM1κ and contains exon 2a instead of 2b. Ectopic expression of human GFP.TPM1κ fusion protein can promote myofibrillogenesis in cardiac mutant axolotl hearts that are lacking in tropomyosin

  6. Multiple isoforms for the catalytic subunit of PKA in the basal fungal lineage Mucor circinelloides.

    Science.gov (United States)

    Fernández Núñez, Lucas; Ocampo, Josefina; Gottlieb, Alexandra M; Rossi, Silvia; Moreno, Silvia

    2016-12-01

    Protein kinase A (PKA) activity is involved in dimorphism of the basal fungal lineage Mucor. From the recently sequenced genome of Mucor circinelloides we could predict ten catalytic subunits of PKA. From sequence alignment and structural prediction we conclude that the catalytic core of the isoforms is conserved, and the difference between them resides in their amino termini. This high number of isoforms is maintained in the subdivision Mucoromycotina. Each paralogue, when compared to the ones form other fungi is more homologous to one of its orthologs than to its paralogs. All of these fungal isoforms cannot be included in the class I or II in which fungal protein kinases have been classified. mRNA levels for each isoform were measured during aerobic and anaerobic growth. The expression of each isoform is differential and associated to a particular growth stage. We reanalyzed the sequence of PKAC (GI 20218944), the only cloned sequence available until now for a catalytic subunit of M. circinelloides. PKAC cannot be classified as a PKA because of its difference in the conserved C-tail; it shares with PKB a conserved C2 domain in the N-terminus. No catalytic activity could be measured for this protein nor predicted bioinformatically. It can thus be classified as a pseudokinase. Its importance can not be underestimated since it is expressed at the mRNA level in different stages of growth, and its deletion is lethal. Copyright © 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  7. MAPA distinguishes genotype-specific variability of highly similar regulatory protein isoforms in potato tuber.

    Science.gov (United States)

    Hoehenwarter, Wolfgang; Larhlimi, Abdelhalim; Hummel, Jan; Egelhofer, Volker; Selbig, Joachim; van Dongen, Joost T; Wienkoop, Stefanie; Weckwerth, Wolfram

    2011-07-01

    Mass Accuracy Precursor Alignment is a fast and flexible method for comparative proteome analysis that allows the comparison of unprecedented numbers of shotgun proteomics analyses on a personal computer in a matter of hours. We compared 183 LC-MS analyses and more than 2 million MS/MS spectra and could define and separate the proteomic phenotypes of field grown tubers of 12 tetraploid cultivars of the crop plant Solanum tuberosum. Protein isoforms of patatin as well as other major gene families such as lipoxygenase and cysteine protease inhibitor that regulate tuber development were found to be the primary source of variability between the cultivars. This suggests that differentially expressed protein isoforms modulate genotype specific tuber development and the plant phenotype. We properly assigned the measured abundance of tryptic peptides to different protein isoforms that share extensive stretches of primary structure and thus inferred their abundance. Peptides unique to different protein isoforms were used to classify the remaining peptides assigned to the entire subset of isoforms based on a common abundance profile using multivariate statistical procedures. We identified nearly 4000 proteins which we used for quantitative functional annotation making this the most extensive study of the tuber proteome to date.

  8. DNA imaging and quantification using chemi-luminescent probes; Imagerie et quantification d`ADN par chimiluminescence

    Energy Technology Data Exchange (ETDEWEB)

    Dorner, G; Redjdal, N; Laniece, P; Siebert, R; Tricoire, H; Valentin, L [Groupe I.P.B., Experimental Research Division, Inst. de Physique Nucleaire, Paris-11 Univ., 91 - Orsay (France)

    1999-11-01

    During this interdisciplinary study we have developed an ultra sensitive and reliable imaging system of DNA labelled by chemiluminescence. Based on a liquid nitrogen cooled CCD, the system achieves sensitivities down to 10 fg/mm{sup 2} labelled DNA over a surface area of 25 x 25 cm{sup 2} with a sub-millimeter resolution. Commercially available chemi-luminescent - and enhancer molecules are compared and their reaction conditions optimized for best signal-to-noise ratios. Double labelling was performed to verify quantification with radioactive probes. (authors) 1 fig.

  9. Human reliability analysis

    International Nuclear Information System (INIS)

    Dougherty, E.M.; Fragola, J.R.

    1988-01-01

    The authors present a treatment of human reliability analysis incorporating an introduction to probabilistic risk assessment for nuclear power generating stations. They treat the subject according to the framework established for general systems theory. Draws upon reliability analysis, psychology, human factors engineering, and statistics, integrating elements of these fields within a systems framework. Provides a history of human reliability analysis, and includes examples of the application of the systems approach

  10. Reliability of electronic systems

    International Nuclear Information System (INIS)

    Roca, Jose L.

    2001-01-01

    Reliability techniques have been developed subsequently as a need of the diverse engineering disciplines, nevertheless they are not few those that think they have been work a lot on reliability before the same word was used in the current context. Military, space and nuclear industries were the first ones that have been involved in this topic, however not only in these environments it is that it has been carried out this small great revolution in benefit of the increase of the reliability figures of the products of those industries, but rather it has extended to the whole industry. The fact of the massive production, characteristic of the current industries, drove four decades ago, to the fall of the reliability of its products, on one hand, because the massively itself and, for other, to the recently discovered and even not stabilized industrial techniques. Industry should be changed according to those two new requirements, creating products of medium complexity and assuring an enough reliability appropriated to production costs and controls. Reliability began to be integral part of the manufactured product. Facing this philosophy, the book describes reliability techniques applied to electronics systems and provides a coherent and rigorous framework for these diverse activities providing a unifying scientific basis for the entire subject. It consists of eight chapters plus a lot of statistical tables and an extensive annotated bibliography. Chapters embrace the following topics: 1- Introduction to Reliability; 2- Basic Mathematical Concepts; 3- Catastrophic Failure Models; 4-Parametric Failure Models; 5- Systems Reliability; 6- Reliability in Design and Project; 7- Reliability Tests; 8- Software Reliability. This book is in Spanish language and has a potentially diverse audience as a text book from academic to industrial courses. (author)

  11. The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4216, Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

    Science.gov (United States)

    Appukuttan, Binoy; McFarland, Trevor J.; Stempel, Andrew; Kassem, Jean B.; Hartzell, Matthew; Zhang, Yi; Bond, Derek; West, Kelsey; Wilson, Reid; Stout, Andrew; Pan, Yuzhen; Ilias, Hoda; Robertson, Kathryn; Klein, Michael L.; Wilson, David; Smith, Justine R.; Stout, J. Timothy

    2012-01-01

    Increased cellular production of vascular endothelial growth factor (VEGF) is responsible for the development and progression of multiple cancers and other neovascular conditions, and therapies targeting post-translational VEGF products are used in the treatment of these diseases. Development of methods to control and modify the transcription of the VEGF gene is an alternative approach that may have therapeutic potential. We have previously shown that isoforms of the transcriptional enhancer factor 1-related (TEAD4) protein can enhance the production of VEGF. In this study we describe a new TEAD4 isoform, TEAD4216, which represses VEGF promoter activity. The TEAD4216 isoform inhibits human VEGF promoter activity and does not require the presence of the hypoxia responsive element (HRE), which is the sequence critical to hypoxia inducible factor (HIF)-mediated effects. The TEAD4216 protein is localized to the cytoplasm, whereas the enhancer isoforms are found within the nucleus. The TEAD4216 isoform can competitively repress the stimulatory activity of the TEAD4434 and TEAD4148 enhancers. Synthesis of the native VEGF165 protein and cellular proliferation is suppressed by the TEAD4216 isoform. Mutational analysis indicates that nuclear or cytoplasmic localization of any isoform determines whether it acts as an enhancer or repressor, respectively. The TEAD4216 isoform appears to inhibit VEGF production independently of the HRE required activity by HIF, suggesting that this alternatively spliced isoform of TEAD4 may provide a novel approach to treat VEGF-dependent diseases. PMID:22761647

  12. Operational safety reliability research

    International Nuclear Information System (INIS)

    Hall, R.E.; Boccio, J.L.

    1986-01-01

    Operating reactor events such as the TMI accident and the Salem automatic-trip failures raised the concern that during a plant's operating lifetime the reliability of systems could degrade from the design level that was considered in the licensing process. To address this concern, NRC is sponsoring the Operational Safety Reliability Research project. The objectives of this project are to identify the essential tasks of a reliability program and to evaluate the effectiveness and attributes of such a reliability program applicable to maintaining an acceptable level of safety during the operating lifetime at the plant

  13. Circuit design for reliability

    CERN Document Server

    Cao, Yu; Wirth, Gilson

    2015-01-01

    This book presents physical understanding, modeling and simulation, on-chip characterization, layout solutions, and design techniques that are effective to enhance the reliability of various circuit units.  The authors provide readers with techniques for state of the art and future technologies, ranging from technology modeling, fault detection and analysis, circuit hardening, and reliability management. Provides comprehensive review on various reliability mechanisms at sub-45nm nodes; Describes practical modeling and characterization techniques for reliability; Includes thorough presentation of robust design techniques for major VLSI design units; Promotes physical understanding with first-principle simulations.

  14. Development of Accident Scenarios and Quantification Methodology for RAON Accelerator

    International Nuclear Information System (INIS)

    Lee, Yongjin; Jae, Moosung

    2014-01-01

    The RIsp (Rare Isotope Science Project) plans to provide neutron-rich isotopes (RIs) and stable heavy ion beams. The accelerator is defined as radiation production system according to Nuclear Safety Law. Therefore, it needs strict operate procedures and safety assurance to prevent radiation exposure. In order to satisfy this condition, there is a need for evaluating potential risk of accelerator from the design stage itself. Though some of PSA researches have been conducted for accelerator, most of them focus on not general accident sequence but simple explanation of accident. In this paper, general accident scenarios are developed by Event Tree and deduce new quantification methodology of Event Tree. In this study, some initial events, which may occur in the accelerator, are selected. Using selected initial events, the accident scenarios of accelerator facility are developed with Event Tree. These results can be used as basic data of the accelerator for future risk assessments. After analyzing the probability of each heading, it is possible to conduct quantification and evaluate the significance of the accident result. If there is a development of the accident scenario for external events, risk assessment of entire accelerator facility will be completed. To reduce the uncertainty of the Event Tree, it is possible to produce a reliable data via the presented quantification techniques

  15. Differential expression of a new isoform of DLG2 in renal oncocytoma

    International Nuclear Information System (INIS)

    Zubakov, Dmitry; Stupar, Zorica; Kovacs, Gyula

    2006-01-01

    Renal oncocytoma, a benign tumour of the kidney, may pose a differential diagnostic problem due to overlapping phenotype with chromophobe renal cell carcinoma or other types of renal cell tumours. Therefore, identification of molecular markers would be of great value for molecular diagnostics of this tumour type. In the current study we applied various techniques, including Affymetrix microarray hybridization and semiquantitative RT-PCR, to identify genes expressed differentially in renal oncocytomas. Subsequently, we used RACE and Northern blot hybridization to characterize the potential candidates for molecular diagnosis. We have identified new isoform of DLG2 gene, which contains 3'-end exons of the known DLG2 gene along with the hypothetical gene FLJ37266. The new isoform is specifically upregulated in renal oncocytoma, whereas the known DLG2 gene is downregulated in this type of kidney tumour. The new isoform of DLG2 is the promising candidate gene for molecular differential diagnostics of renal oncocytoma

  16. The ROCK isoforms differentially regulate the morphological characteristics of carcinoma cells.

    Science.gov (United States)

    Jerrell, Rachel J; Leih, Mitchell J; Parekh, Aron

    2017-06-26

    Rho-associated kinase (ROCK) activity drives cell migration via actomyosin contractility. During invasion, individual cancer cells can transition between 2 modes of migration, mesenchymal and amoeboid. Changes in ROCK activity can cause a switch between these migration phenotypes which are defined by distinct morphologies. However, recent studies have shown that the ROCK isoforms are not functionally redundant as previously thought. Therefore, it is unclear whether the ROCK isoforms play different roles in regulating migration phenotypes. Here, we found that ROCK1 and ROCK2 differentially regulate carcinoma cell morphology resulting in intermediate phenotypes that share some mesenchymal and amoeboid characteristics. These findings suggest that the ROCK isoforms play unique roles in the phenotypic plasticity of mesenchymal carcinoma cells which may have therapeutic implications.

  17. Branchial Expression Patterns of Claudin Isoforms in Atlantic Salmon During Seawater Acclimation and Smoltification

    DEFF Research Database (Denmark)

    Tipsmark, Christian K; Kiilerich, Pia; Nilsen, Tom O

    2008-01-01

    in epithelia. We identified Atlantic salmon genes belonging to the claudin family by screening expressed sequence tag libraries available at NCBI and classification was performed with aid of maximum likelihood and neighbour-joining analysis. In gill libraries, five isoforms (10e, 27a, 28a, 28b and 30) were...... present and QPCR analysis confirmed tissue-specific expression in gill when compared to kidney, intestine, heart, muscle, brain and liver. Expression patterns during acclimation of freshwater salmon to seawater (SW) and during the smoltification process were examined. Acclimation to SW reduced...... induced no significant changes in expression of the other isoforms. This study demonstrates the expression of an array of salmon claudin isoforms and shows that SW acclimation involves inverse regulation, in the gill, of claudin 10e versus claudin 27a and 30. It is possible, that claudin 10e...

  18. TAPAS: tools to assist the targeted protein quantification of human alternative splice variants.

    Science.gov (United States)

    Yang, Jae-Seong; Sabidó, Eduard; Serrano, Luis; Kiel, Christina

    2014-10-15

    In proteomes of higher eukaryotes, many alternative splice variants can only be detected by their shared peptides. This makes it highly challenging to use peptide-centric mass spectrometry to distinguish and to quantify protein isoforms resulting from alternative splicing events. We have developed two complementary algorithms based on linear mathematical models to efficiently compute a minimal set of shared and unique peptides needed to quantify a set of isoforms and splice variants. Further, we developed a statistical method to estimate the splice variant abundances based on stable isotope labeled peptide quantities. The algorithms and databases are integrated in a web-based tool, and we have experimentally tested the limits of our quantification method using spiked proteins and cell extracts. The TAPAS server is available at URL http://davinci.crg.es/tapas/. luis.serrano@crg.eu or christina.kiel@crg.eu Supplementary data are available at Bioinformatics online. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  19. Different characteristics and nucleotide binding properties of inosine monophosphate dehydrogenase (IMPDH isoforms.

    Directory of Open Access Journals (Sweden)

    Elaine C Thomas

    Full Text Available We recently reported that Inosine Monophosphate Dehydrogenase (IMPDH, a rate-limiting enzyme in de novo guanine nucleotide biosynthesis, clustered into macrostructures in response to decreased nucleotide levels and that there were differences between the IMPDH isoforms, IMPDH1 and IMPDH2. We hypothesised that the Bateman domains, which are present in both isoforms and serve as energy-sensing/allosteric modules in unrelated proteins, would contribute to isoform-specific differences and that mutations situated in and around this domain in IMPDH1 which give rise to retinitis pigmentosa (RP would compromise regulation. We employed immuno-electron microscopy to investigate the ultrastructure of IMPDH macrostructures and live-cell imaging to follow clustering of an IMPDH2-GFP chimera in real-time. Using a series of IMPDH1/IMPDH2 chimera we demonstrated that the propensity to cluster was conferred by the N-terminal 244 amino acids, which includes the Bateman domain. A protease protection assay suggested isoform-specific purine nucleotide binding characteristics, with ATP protecting IMPDH1 and AMP protecting IMPDH2, via a mechanism involving conformational changes upon nucleotide binding to the Bateman domain without affecting IMPDH catalytic activity. ATP binding to IMPDH1 was confirmed in a nucleotide binding assay. The RP-causing mutation, R224P, abolished ATP binding and nucleotide protection and this correlated with an altered propensity to cluster. Collectively these data demonstrate that (i the isoforms are differentially regulated by AMP and ATP by a mechanism involving the Bateman domain, (ii communication occurs between the Bateman and catalytic domains and (iii the RP-causing mutations compromise such regulation. These findings support the idea that the IMPDH isoforms are subject to distinct regulation and that regulatory defects contribute to human disease.

  20. In vivo human apolipoprotein E isoform fractional turnover rates in the CNS.

    Directory of Open Access Journals (Sweden)

    Kristin R Wildsmith

    Full Text Available Apolipoprotein E (ApoE is the strongest genetic risk factor for Alzheimer's disease and has been implicated in the risk for other neurological disorders. The three common ApoE isoforms (ApoE2, E3, and E4 each differ by a single amino acid, with ApoE4 increasing and ApoE2 decreasing the risk of Alzheimer's disease (AD. Both the isoform and amount of ApoE in the brain modulate AD pathology by altering the extent of amyloid beta (Aβ peptide deposition. Therefore, quantifying ApoE isoform production and clearance rates may advance our understanding of the role of ApoE in health and disease. To measure the kinetics of ApoE in the central nervous system (CNS, we applied in vivo stable isotope labeling to quantify the fractional turnover rates of ApoE isoforms in 18 cognitively-normal adults and in ApoE3 and ApoE4 targeted-replacement mice. No isoform-specific differences in CNS ApoE3 and ApoE4 turnover rates were observed when measured in human CSF or mouse brain. However, CNS and peripheral ApoE isoform turnover rates differed substantially, which is consistent with previous reports and suggests that the pathways responsible for ApoE metabolism are different in the CNS and the periphery. We also demonstrate a slower turnover rate for CSF ApoE than that for amyloid beta, another molecule critically important in AD pathogenesis.

  1. Mesenchymal Stromal Cells for Sphincter Regeneration: Role of Laminin Isoforms upon Myogenic Differentiation

    Science.gov (United States)

    Seeger, Tanja; Hart, Melanie; Patarroyo, Manuel; Rolauffs, Bernd; Aicher, Wilhelm K.; Klein, Gerd

    2015-01-01

    Multipotent mesenchymal stromal cells (MSCs) are well known for their tri-lineage potential and ability to differentiate in vitro into osteogenic, chondrogenic or adipogenic lineages. By selecting appropriate conditions MSCs can also be differentiated in vitro into the myogenic lineage and are therefore a promising option for cell-based regeneration of muscle tissue such as an aged or damaged sphincter muscle. For the differentiation into the myogenic lineage there is still a need to evaluate the effects of extracellular matrix proteins such as laminins (LM) which are crucial for different stem cell types and for normal muscle function. The laminin family consists of 16 functionally different isoforms with LM-211 being the most abundant isoform of adult muscle tissues. In the sphincter tissue a strong expression of the isoforms LM-211/221, LM-411/421 and LM-511/521 can be detected in the different cell layers. Bone marrow-derived MSCs in culture, however, mainly express the isoforms LM-411 and LM-511, but not LM-211. Even after myogenic differentiation, LM-211 can hardly be detected. All laminin isoforms tested (LM-211, LM-411, LM-511 and LM-521) showed a significant inhibition of the proliferation of undifferentiated MSCs but, with the exception of LM-521, they had no influence on the proliferation of MSCs cultivated in myogenic medium. The strongest cellular adhesion of MSCs was to LM-511 and LM-521, whereas LM-211 was only a weakly-adhesive substrate for MSCs. Myogenic differentiation of MSCs even reduced the interaction with LM-211, but it did not affect the interaction with LM-511 and LM-521. Since during normal myogenesis the latter two isoforms are the major laminins surrounding developing myogenic progenitors, α5 chain-containing laminins are recommended for further improvements of myogenic differentiation protocols of MSCs into smooth muscle cells. PMID:26406476

  2. Mesenchymal Stromal Cells for Sphincter Regeneration: Role of Laminin Isoforms upon Myogenic Differentiation.

    Directory of Open Access Journals (Sweden)

    Tanja Seeger

    Full Text Available Multipotent mesenchymal stromal cells (MSCs are well known for their tri-lineage potential and ability to differentiate in vitro into osteogenic, chondrogenic or adipogenic lineages. By selecting appropriate conditions MSCs can also be differentiated in vitro into the myogenic lineage and are therefore a promising option for cell-based regeneration of muscle tissue such as an aged or damaged sphincter muscle. For the differentiation into the myogenic lineage there is still a need to evaluate the effects of extracellular matrix proteins such as laminins (LM which are crucial for different stem cell types and for normal muscle function. The laminin family consists of 16 functionally different isoforms with LM-211 being the most abundant isoform of adult muscle tissues. In the sphincter tissue a strong expression of the isoforms LM-211/221, LM-411/421 and LM-511/521 can be detected in the different cell layers. Bone marrow-derived MSCs in culture, however, mainly express the isoforms LM-411 and LM-511, but not LM-211. Even after myogenic differentiation, LM-211 can hardly be detected. All laminin isoforms tested (LM-211, LM-411, LM-511 and LM-521 showed a significant inhibition of the proliferation of undifferentiated MSCs but, with the exception of LM-521, they had no influence on the proliferation of MSCs cultivated in myogenic medium. The strongest cellular adhesion of MSCs was to LM-511 and LM-521, whereas LM-211 was only a weakly-adhesive substrate for MSCs. Myogenic differentiation of MSCs even reduced the interaction with LM-211, but it did not affect the interaction with LM-511 and LM-521. Since during normal myogenesis the latter two isoforms are the major laminins surrounding developing myogenic progenitors, α5 chain-containing laminins are recommended for further improvements of myogenic differentiation protocols of MSCs into smooth muscle cells.

  3. Differential regulation of protein phosphatase 1 (PP1) isoforms in human heart failure and atrial fibrillation.

    Science.gov (United States)

    Meyer-Roxlau, Stefanie; Lämmle, Simon; Opitz, Annett; Künzel, Stephan; Joos, Julius P; Neef, Stefan; Sekeres, Karolina; Sossalla, Samuel; Schöndube, Friedrich; Alexiou, Konstantin; Maier, Lars S; Dobrev, Dobromir; Guan, Kaomei; Weber, Silvio; El-Armouche, Ali

    2017-07-01

    Protein phosphatase 1 (PP1) is a key regulator of important cardiac signaling pathways. Dysregulation of PP1 has been heavily implicated in cardiac dysfunctions. Accordingly, pharmacological targeting of PP1 activity is considered for therapeutic intervention in human cardiomyopathies. Recent evidence from animal models implicated previously unrecognized, isoform-specific activities of PP1 in the healthy and diseased heart. Therefore, this study examined the expression of the distinct PP1 isoforms PP1α, β, and γ in human heart failure (HF) and atrial fibrillation (AF) and addressed the consequences of β-adrenoceptor blocker (beta-blocker) therapy for HF patients with reduced ejection fraction on PP1 isoform expression. Using western blot analysis, we found greater abundance of PP1 isoforms α and γ but unaltered PP1β levels in left ventricular myocardial tissues from HF patients as compared to non-failing controls. However, expression of all three PP1 isoforms was higher in atrial appendages from patients with AF compared to patients with sinus rhythm. Moreover, we found that in human failing ventricles, beta-blocker therapy was associated with lower PP1α abundance and activity, as indicated by higher phosphorylation of the PP1α-specific substrate eIF2α. Greater eIF2α phosphorylation is a known repressor of protein translation, and accordingly, we found lower levels of the endoplasmic reticulum (ER) stress marker Grp78 in the very same samples. We propose that isoform-specific targeting of PP1α activity may be a novel and innovative therapeutic strategy for the treatment of human cardiac diseases by reducing ER stress conditions.

  4. GSK3β isoform-selective regulation of depression, memory and hippocampal cell proliferation.

    Science.gov (United States)

    Pardo, M; Abrial, E; Jope, R S; Beurel, E

    2016-03-01

    Abnormally active glycogen synthase kinase-3 (GSK3) contributes to pathological processes in multiple psychiatric and neurological disorders. Modeled in mice, this includes increasing susceptibility to dysregulation of mood-relevant behaviors, impairing performance in several cognitive tasks and impairing adult hippocampal neural precursor cell (NPC) proliferation. These deficits are all evident in GSK3α/β knockin mice, in which serine-to-alanine mutations block the inhibitory serine phosphorylation regulation of both GSK3 isoforms, leaving GSK3 hyperactive. It was unknown if both GSK3 isoforms perform redundant actions in these processes, or if hyperactivity of one GSK3 isoform has a predominant effect. To test this, we examined GSK3α or GSK3β knockin mice in which only one isoform was mutated to a hyperactive form. Only GSK3β, not GSK3α, knockin mice displayed heightened vulnerability to the learned helplessness model of depression-like behavior. Three cognitive measures impaired in GSK3α/β knockin mice showed differential regulation by GSK3 isoforms. Novel object recognition was impaired in GSK3β, not in GSK3α, knockin mice, whereas temporal order memory was not impaired in GSK3α or GSK3β knockin mice, and co-ordinate spatial processing was impaired in both GSK3α and GSK3β knockin mice. Adult hippocampal NPC proliferation was severely impaired in GSK3β knockin mice, but not impaired in GSK3α knockin mice. Increased activity of GSK3β, in the absence of overexpression or disease pathology, is sufficient to impair mood regulation, novel object recognition and hippocampal NPC proliferation, whereas hyperactive GSK3α individually does not impair these processes. These results show that hyperactivity of the two GSK3 isoforms execute non-redundant effects on these processes. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

  5. BORIS/CTCFL mRNA isoform expression and epigenetic regulation in epithelial ovarian cancer

    Science.gov (United States)

    Link, Petra A.; Zhang, Wa; Odunsi, Kunle; Karpf, Adam R.

    2013-01-01

    Cancer germline (CG) genes are normally expressed in germ cells and aberrantly expressed in a variety of cancers; their immunogenicity has led to the widespread development of cancer vaccines targeting these antigens. BORIS/CTCFL is an autosomal CG antigen and promising cancer vaccine target. BORIS is the only known paralog of CTCF, a gene intimately involved in genomic imprinting, chromatin insulation, and nuclear regulation. We have previously shown that BORIS is expressed in epithelial ovarian cancer (EOC) and that its expression coincides with promoter and global DNA hypomethylation. Recently, 23 different BORIS mRNA variants have been described, and have been functionally grouped into six BORIS isoform families (sf1–sf6). In the present study, we have characterized the expression of BORIS isoform families in normal ovary (NO) and EOC, the latter of which were selected to include two groups with widely varying global DNA methylation status. We find selective expression of BORIS isoform families in NO, which becomes altered in EOC, primarily by the activation of BORIS sf1 in EOC. When comparing EOC samples based on methylation status, we find that BORIS sf1 and sf2 isoform families are selectively activated in globally hypomethylated tumors. In contrast, CTCF is downregulated in EOC, and the ratio of BORIS sf1, sf2, and sf6 isoform families as a function of CTCF is elevated in hypomethylated tumors. Finally, the expression of all BORIS isoform families was induced to varying extents by epigenetic modulatory drugs in EOC cell lines, particularly when DNMT and HDAC inhibitors were used in combination. PMID:23390377

  6. Expression of Metallothionein and Vascular Endothelial Growth Factor Isoforms in Breast Cancer Cells.

    Science.gov (United States)

    Wierzowiecka, Barbara; Gomulkiewicz, Agnieszka; Cwynar-Zajac, Lucja; Olbromski, Mateusz; Grzegrzolka, Jedrzej; Kobierzycki, Christopher; Podhorska-Okolow, Marzenna; Dziegiel, Piotr

    2016-01-01

    Metallothioneins (MTs) are low-molecular-weight and cysteine-rich proteins that bind heavy metal ions and oxygen-free radicals. MTs are commonly expressed in various tissues of mammals and are involved in regulation of cell proliferation and differentiation, and may be engaged in angiogenesis. Expression of MTs has been studied in many cancer types, especially breast cancer. The research results indicate that MTs may play important, although not yet fully known, roles in cancer angiogenesis. The aim of this study was to analyze the level of gene expression of selected MT isoforms induced with zinc ions in correlation with vascular endothelial growth factor (VEGF) isoforms in in vitro models of breast cancer. The studies were carried out in three breast cancer cell lines (MCF-7, SK-BR-3, MDA-MB-231). An epithelial cell line derived from normal breast tissue (Me16c) was used as a control. The levels of expression of selected MT isoforms and selected genes involved in angiogenesis were studied with real-time PCR. Expression of different MT isoforms was induced by zinc ions to differing degrees in individual breast cancer cell lines. An increase in the expression of some MT isoforms was associated with a slight increase in the level of expression of VEGFA. The research results may indicate certain correlation between an increased expression of selected MT isoforms and a pro-angiogenic factor VEGF in specific types of breast cancer cells. Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  7. Diagnostic Accuracy of Cerebrospinal Fluid Amyloid-β Isoforms for Early and Differential Dementia Diagnosis.

    Science.gov (United States)

    Struyfs, Hanne; Van Broeck, Bianca; Timmers, Maarten; Fransen, Erik; Sleegers, Kristel; Van Broeckhoven, Christine; De Deyn, Peter P; Streffer, Johannes R; Mercken, Marc; Engelborghs, Sebastiaan

    2015-01-01

    Overlapping cerebrospinal fluid biomarkers (CSF) levels between Alzheimer's disease (AD) and non-AD patients decrease differential diagnostic accuracy of the AD core CSF biomarkers. Amyloid-β (Aβ) isoforms might improve the AD versus non-AD differential diagnosis. To determine the added diagnostic value of Aβ isoforms, Aβ(1-37), Aβ(1-38), and Aβ(1-40), as compared to the AD CSF biomarkers Aβ(1-42), T-tau, and P-tau(181P). CSF from patients with dementia due to AD (n = 50), non-AD dementias (n = 50), mild cognitive impairment due to AD (n = 50) and non-demented controls (n = 50) was analyzed with a prototype multiplex assay using MSD detection technology. The non-AD group consisted of frontotemporal dementia (FTD; n = 17), dementia with Lewy bodies (DLB; n = 17), and vascular dementia (n = 16). Aβ(1-37) and Aβ(1-38) increased accuracy to differentiate AD from FTD or DLB. Aβ(1-37), Aβ(1-38), and Aβ(1-40) levels correlated with Mini-Mental State Examination scores and disease duration in dementia due to AD. The Aβ(1-42)/Aβ(1-40) ratio improved diagnostic performance of Aβ(1-42) in most differential diagnostic situations. Aβ(1-42) levels were lower in APOE ε4 carriers compared to non-carriers. Aβ isoforms help to differentiate AD from FTD and DLB. Aβ isoforms increase diagnostic performance of Aβ(1-42). In contrast to Aβ1-42, Aβ isoforms seem to be correlated with disease severity in AD. Adding the Aβ isoforms to the current biomarker panel could enhance diagnostic accuracy.

  8. Transcriptome-wide identification and characterization of CAD isoforms specific for podophyllotoxin biosynthesis from Podophyllum hexandrum.

    Science.gov (United States)

    Bhattacharyya, Dipto; Hazra, Saptarshi; Banerjee, Anindyajit; Datta, Riddhi; Kumar, Deepak; Chakrabarti, Saikat; Chattopadhyay, Sharmila

    2016-09-01

    Podophyllotoxin (ptox) is a therapeutically important lignan derived from Podophyllum hexandrum and is used as a precursor for the synthesis of anticancer drugs etoposide, teniposide and etopophose. In spite of its enormous economic significance, genomic information on this endangered medicinal herb is scarce. We have performed de novo transcriptome analysis of methyl jasmonate (MeJA)-treated P. hexandrum cell cultures exhibiting enhanced ptox accumulation. The results revealed the maximum up-regulation of several isoforms of cinnamyl alcohol dehydrogenase (CAD). CAD catalyzes the synthesis of coniferyl alcohol and sinapyl alcohol from coniferaldehyde (CAld) and sinapaldehyde respectively. Coniferyl alcohol can produce both lignin and lignan while sinapyl alcohol produces only lignin. To isolate the CAD isoforms favoring ptox, we deduced full length cDNA sequences of four CAD isoforms: PhCAD1, PhCAD2, PhCAD3 and PhCAD4 from the contigs of the transcriptome data. In vitro enzyme assays indicated a higher affinity for CAld over sinapaldehyde for each isoform. In silico molecular docking analyses also suggested that PhCAD3 has a higher binding preference with CAld over sinapaldehyde, followed by PhCAD4, PhCAD2, and PhCAD1, respectively. The transgenic cell cultures overexpressing these isoforms independently revealed that PhCAD3 favored the maximum accumulation of ptox as compared to lignin followed by PhCAD4 and PhCAD2, whereas, PhCAD1 favored both equally. Together, our study reveals transcriptome-wide identification and characterization of ptox specific CAD isoforms from P. hexandrum. It provides a useful resource for future research not only on the ptox biosynthetic pathway but on overall P. hexandrum, an endangered medicinal herb with immense therapeutic importance.

  9. The α and Δ isoforms of CREB1 are required to maintain normal pulmonary vascular resistance.

    Directory of Open Access Journals (Sweden)

    Lili Li

    Full Text Available Chronic hypoxia causes pulmonary hypertension associated with structural alterations in pulmonary vessels and sustained vasoconstriction. The transcriptional mechanisms responsible for these distinctive changes are unclear. We have previously reported that CREB1 is activated in the lung in response to alveolar hypoxia but not in other organs. To directly investigate the role of α and Δ isoforms of CREB1 in the regulation of pulmonary vascular resistance we examined the responses of mice in which these isoforms of CREB1 had been inactivated by gene mutation, leaving only the β isoform intact (CREB(αΔ mice. Here we report that expression of CREB regulated genes was altered in the lungs of CREB(αΔ mice. CREB(αΔ mice had greater pulmonary vascular resistance than wild types, both basally in normoxia and following exposure to hypoxic conditions for three weeks. There was no difference in rho kinase mediated vasoconstriction between CREB(αΔ and wild type mice. Stereological analysis of pulmonary vascular structure showed characteristic wall thickening and lumen reduction in hypoxic wild-type mice, with similar changes observed in CREB(αΔ. CREB(αΔ mice had larger lungs with reduced epithelial surface density suggesting increased pulmonary compliance. These findings show that α and Δ isoforms of CREB1 regulate homeostatic gene expression in the lung and that normal activity of these isoforms is essential to maintain low pulmonary vascular resistance in both normoxic and hypoxic conditions and to maintain the normal alveolar structure. Interventions that enhance the actions of α and Δ isoforms of CREB1 warrant further investigation in hypoxic lung diseases.

  10. Hawaii Electric System Reliability

    Energy Technology Data Exchange (ETDEWEB)

    Loose, Verne William [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Silva Monroy, Cesar Augusto [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2012-08-01

    This report addresses Hawaii electric system reliability issues; greater emphasis is placed on short-term reliability but resource adequacy is reviewed in reference to electric consumers’ views of reliability “worth” and the reserve capacity required to deliver that value. The report begins with a description of the Hawaii electric system to the extent permitted by publicly available data. Electrical engineering literature in the area of electric reliability is researched and briefly reviewed. North American Electric Reliability Corporation standards and measures for generation and transmission are reviewed and identified as to their appropriateness for various portions of the electric grid and for application in Hawaii. Analysis of frequency data supplied by the State of Hawaii Public Utilities Commission is presented together with comparison and contrast of performance of each of the systems for two years, 2010 and 2011. Literature tracing the development of reliability economics is reviewed and referenced. A method is explained for integrating system cost with outage cost to determine the optimal resource adequacy given customers’ views of the value contributed by reliable electric supply. The report concludes with findings and recommendations for reliability in the State of Hawaii.

  11. Hawaii electric system reliability.

    Energy Technology Data Exchange (ETDEWEB)

    Silva Monroy, Cesar Augusto; Loose, Verne William

    2012-09-01

    This report addresses Hawaii electric system reliability issues; greater emphasis is placed on short-term reliability but resource adequacy is reviewed in reference to electric consumers' views of reliability %E2%80%9Cworth%E2%80%9D and the reserve capacity required to deliver that value. The report begins with a description of the Hawaii electric system to the extent permitted by publicly available data. Electrical engineering literature in the area of electric reliability is researched and briefly reviewed. North American Electric Reliability Corporation standards and measures for generation and transmission are reviewed and identified as to their appropriateness for various portions of the electric grid and for application in Hawaii. Analysis of frequency data supplied by the State of Hawaii Public Utilities Commission is presented together with comparison and contrast of performance of each of the systems for two years, 2010 and 2011. Literature tracing the development of reliability economics is reviewed and referenced. A method is explained for integrating system cost with outage cost to determine the optimal resource adequacy given customers' views of the value contributed by reliable electric supply. The report concludes with findings and recommendations for reliability in the State of Hawaii.

  12. Improving machinery reliability

    CERN Document Server

    Bloch, Heinz P

    1998-01-01

    This totally revised, updated and expanded edition provides proven techniques and procedures that extend machinery life, reduce maintenance costs, and achieve optimum machinery reliability. This essential text clearly describes the reliability improvement and failure avoidance steps practiced by best-of-class process plants in the U.S. and Europe.

  13. LED system reliability

    NARCIS (Netherlands)

    Driel, W.D. van; Yuan, C.A.; Koh, S.; Zhang, G.Q.

    2011-01-01

    This paper presents our effort to predict the system reliability of Solid State Lighting (SSL) applications. A SSL system is composed of a LED engine with micro-electronic driver(s) that supplies power to the optic design. Knowledge of system level reliability is not only a challenging scientific

  14. Integrated system reliability analysis

    DEFF Research Database (Denmark)

    Gintautas, Tomas; Sørensen, John Dalsgaard

    Specific targets: 1) The report shall describe the state of the art of reliability and risk-based assessment of wind turbine components. 2) Development of methodology for reliability and risk-based assessment of the wind turbine at system level. 3) Describe quantitative and qualitative measures...

  15. Reliability of neural encoding

    DEFF Research Database (Denmark)

    Alstrøm, Preben; Beierholm, Ulrik; Nielsen, Carsten Dahl

    2002-01-01

    The reliability with which a neuron is able to create the same firing pattern when presented with the same stimulus is of critical importance to the understanding of neuronal information processing. We show that reliability is closely related to the process of phaselocking. Experimental results f...

  16. Considerations on the elements of quantifying human reliability

    International Nuclear Information System (INIS)

    Straeter, Oliver

    2004-01-01

    This paper attempts to provide a contribution for the discussion of what the term 'data' means and how the qualitative perspective can be linked with the quantitative one. It will argue that the terms 'quantitative data' and 'qualitative data' are not distinct but a continuum that spans over the entire spectrum of the expertise that has to be involved in the HRA process. It elaborates the rational behind any human reliability quantification figure and suggests a scientific way forward to better data for human reliability assessment

  17. Crystal structures of a halophilic archaeal malate synthase from Haloferax volcanii and comparisons with isoforms A and G

    Science.gov (United States)

    2011-01-01

    Background Malate synthase, one of the two enzymes unique to the glyoxylate cycle, is found in all three domains of life, and is crucial to the utilization of two-carbon compounds for net biosynthetic pathways such as gluconeogenesis. In addition to the main isoforms A and G, so named because of their differential expression in E. coli grown on either acetate or glycolate respectively, a third distinct isoform has been identified. These three isoforms differ considerably in size and sequence conservation. The A isoform (MSA) comprises ~530 residues, the G isoform (MSG) is ~730 residues, and this third isoform (MSH-halophilic) is ~430 residues in length. Both isoforms A and G have been structurally characterized in detail, but no structures have been reported for the H isoform which has been found thus far only in members of the halophilic Archaea. Results We have solved the structure of a malate synthase H (MSH) isoform member from Haloferax volcanii in complex with glyoxylate at 2.51 Å resolution, and also as a ternary complex with acetyl-coenzyme A and pyruvate at 1.95 Å. Like the A and G isoforms, MSH is based on a β8/α8 (TIM) barrel. Unlike previously solved malate synthase structures which are all monomeric, this enzyme is found in the native state as a trimer/hexamer equilibrium. Compared to isoforms A and G, MSH displays deletion of an N-terminal domain and a smaller deletion at the C-terminus. The MSH active site is closely superimposable with those of MSA and MSG, with the ternary complex indicating a nucleophilic attack on pyruvate by the enolate intermediate of acetyl-coenzyme A. Conclusions The reported structures of MSH from Haloferax volcanii allow a detailed analysis and comparison with previously solved structures of isoforms A and G. These structural comparisons provide insight into evolutionary relationships among these isoforms, and also indicate that despite the size and sequence variation, and the truncated C-terminal domain of the H

  18. Design reliability engineering

    International Nuclear Information System (INIS)

    Buden, D.; Hunt, R.N.M.

    1989-01-01

    Improved design techniques are needed to achieve high reliability at minimum cost. This is especially true of space systems where lifetimes of many years without maintenance are needed and severe mass limitations exist. Reliability must be designed into these systems from the start. Techniques are now being explored to structure a formal design process that will be more complete and less expensive. The intent is to integrate the best features of design, reliability analysis, and expert systems to design highly reliable systems to meet stressing needs. Taken into account are the large uncertainties that exist in materials, design models, and fabrication techniques. Expert systems are a convenient method to integrate into the design process a complete definition of all elements that should be considered and an opportunity to integrate the design process with reliability, safety, test engineering, maintenance and operator training. 1 fig

  19. Bayesian methods in reliability

    Science.gov (United States)

    Sander, P.; Badoux, R.

    1991-11-01

    The present proceedings from a course on Bayesian methods in reliability encompasses Bayesian statistical methods and their computational implementation, models for analyzing censored data from nonrepairable systems, the traits of repairable systems and growth models, the use of expert judgment, and a review of the problem of forecasting software reliability. Specific issues addressed include the use of Bayesian methods to estimate the leak rate of a gas pipeline, approximate analyses under great prior uncertainty, reliability estimation techniques, and a nonhomogeneous Poisson process. Also addressed are the calibration sets and seed variables of expert judgment systems for risk assessment, experimental illustrations of the use of expert judgment for reliability testing, and analyses of the predictive quality of software-reliability growth models such as the Weibull order statistics.

  20. Effect of renal replacement therapy on retinol-binding protein 4 isoforms

    DEFF Research Database (Denmark)

    Frey, Simone K; Henze, Andrea; Nagl, Britta

    2009-01-01

    Retinol-binding protein 4 (RBP4) levels are elevated in the serum of patients with kidney dysfunction. We recently showed that RBP4 isoforms including apo-RBP4 (RBP4 not bound to retinol) and RBP4 truncated at the C-terminus (RBP4-L, RBP4-LL) are increased in the serum of patients with kidney dis...... diseases but not in serum of patients with various liver diseases. The aim of this study was to investigate the effect of renal replacement therapy on RBP4 isoforms....

  1. Protein kinase C isoforms at the neuromuscular junction: localization and specific roles in neurotransmission and development.

    Science.gov (United States)

    Lanuza, Maria A; Santafe, Manel M; Garcia, Neus; Besalduch, Núria; Tomàs, Marta; Obis, Teresa; Priego, Mercedes; Nelson, Phillip G; Tomàs, Josep

    2014-01-01

    The protein kinase C family (PKC) regulates a variety of neural functions including neurotransmitter release. The selective activation of a wide range of PKC isoforms in different cells and domains is likely to contribute to the functional diversity of PKC phosphorylating activity. In this review, we describe the isoform localization, phosphorylation function, regulation and signalling of the PKC family at the neuromuscular junction. Data show the involvement of the PKC family in several important functions at the neuromuscular junction and in particular in the maturation of the synapse and the modulation of neurotransmission in the adult. © 2013 Anatomical Society.

  2. VEGF121b and VEGF165b are weakly angiogenic isoforms of VEGF-A

    Directory of Open Access Journals (Sweden)

    Pio Ruben

    2010-12-01

    Full Text Available Abstract Background Different isoforms of VEGF-A (mainly VEGF121, VEGF165 and VEGF189 have been shown to display particular angiogenic properties in the generation of a functional tumor vasculature. Recently, a novel class of VEGF-A isoforms, designated as VEGFxxxb, generated through alternative splicing, have been described. Previous studies have suggested that these isoforms may inhibit angiogenesis. In the present work we have produced recombinant VEGF121/165b proteins in the yeast Pichia pastoris and constructed vectors to overexpress these isoforms and assess their angiogenic potential. Results Recombinant VEGF121/165b proteins generated either in yeasts or mammalian cells activated VEGFR2 and its downstream effector ERK1/2, although to a lesser extent than VEGF165. Furthermore, treatment of endothelial cells with VEGF121/165b increased cell proliferation compared to untreated cells, although such stimulation was lower than that induced by VEGF165. Moreover, in vivo angiogenesis assays confirmed angiogenesis stimulation by VEGF121/165b isoforms. A549 and PC-3 cells overexpressing VEGF121b or VEGF165b (or carrying the PCDNA3.1 empty vector, as control and xenotransplanted into nude mice showed increased tumor volume and angiogenesis compared to controls. To assess whether the VEGFxxxb isoforms are differentially expressed in tumors compared to healthy tissues, immunohistochemical analysis was conducted on a breast cancer tissue microarray. A significant increase (p xxxb and total VEGF-A protein expression in infiltrating ductal carcinomas compared to normal breasts was observed. A positive significant correlation (r = 0.404, p = 0.033 between VEGFxxxb and total VEGF-A was found. Conclusions Our results demonstrate that VEGF121/165b are not anti-angiogenic, but weakly angiogenic isoforms of VEGF-A. In addition, VEGFxxxb isoforms are up-regulated in breast cancer in comparison with non malignant breast tissues. These results are to be taken

  3. Complex p63 mRNA isoform expression patterns in squamous cell carcinoma of the head and neck

    DEFF Research Database (Denmark)

    Thurfjell, N.; Coates, P.J.; Uusitalo, T.

    2004-01-01

    on the role of p63 expression in human tumours, we used quantitative real-time RT-PCR to study individual p63 isoforms in squamous cell carcinomas of the head and neck (SCCHN). In keeping with previous reports, expression of the deltaN- and p63alpha-isoforms predominated and deltaNp63 mRNA was expressed...

  4. Lipoprotein(a) levels, apo(a) isoform size, and coronary heart disease risk in the Framingham Offspring Study

    Science.gov (United States)

    The aim of this study was to assess the independent contributions of plasma levels of lipoprotein(a) [Lp(a)], Lp(a) cholesterol, and of apo(a) isoform size to prospective coronary heart disease (CHD) risk. Plasma Lp(a) and Lp(a) cholesterol levels, and apo(a) isoform size were measured at examinati...

  5. Distinct transthyretin oxidation isoform profile in spinal fluid from patients with Alzheimer’s disease and mild cognitive impairment

    DEFF Research Database (Denmark)

    Poulsen, Keld; Bahl, Justyna Mc; Simonsen, Anja H

    2014-01-01

    BACKGROUND: Transthyretin (TTR), an abundant protein in cerebrospinal fluid (CSF), contains a free, oxidation-prone cysteine residue that gives rise to TTR isoforms. These isoforms may reflect conditions in vivo. Since increased oxidative stress has been linked to neurodegenerative disorders such...

  6. MicroRNA-281 regulates the expression of ecdysone receptor (EcR) isoform B in the silkworm, Bombyx mori

    Science.gov (United States)

    Hundreds of Bombyx mori miRNAs had been identified in recent years, but their function in vivo remains poorly understood. The silkworm EcR gene (BmEcR) has three transcriptional isoforms, A, B1 and B2. Isoform sequences are different in the 3’UTR region of the gene, which is the case only in insects...

  7. Laminin isoforms: biological roles and effects on the intracellular distribution of nuclear proteins in intestinal epithelial cells

    International Nuclear Information System (INIS)

    Turck, Natacha; Gross, Isabelle; Gendry, Patrick; Stutzmann, Jeanne; Freund, Jean-Noel; Kedinger, Michele; Simon-Assmann, Patricia; Launay, Jean-Francois

    2005-01-01

    Laminins are structurally and functionally major components of the extracellular matrix. Four isoforms of laminins (laminin-1, -2, -5 and -10) are expressed in a specific pattern along the crypt-villus axis of the intestine. Previous works indicated that expression of these isoforms is developmentally regulated and that laminins could modulate the behaviour of intestinal cells, but the exact role of each isoform remained unclear. Here, we report the first systematic analysis of the cellular functions of the four isoforms using the human colon adenocarcinoma Caco2/TC7 cell line as a model. We compared the respective abilities of each isoform to modulate adhesion, proliferation and differentiation of intestinal epithelial cells. We found that the isoforms were functionally distinct, with laminin-10 being the most adhesive substratum, laminin-2, laminin-5 and laminin-10 enhancing cellular proliferation and at the opposite, laminin-1 stimulating intestinal cell differentiation. To begin to characterise the molecular events induced by the different isoforms, we examined by immunofluorescence the intracellular distribution of several nuclear proteins, recently highlighted by a nuclear proteomic approach. We observed clear nucleocytoplasmic redistribution of these proteins, which depended on the laminin isoform. These results provide evidence for a distinct functional role of laminins in intestinal cell functions characterised by specific localisation of nuclear proteins

  8. Reliability of construction materials

    International Nuclear Information System (INIS)

    Merz, H.

    1976-01-01

    One can also speak of reliability with respect to materials. While for reliability of components the MTBF (mean time between failures) is regarded as the main criterium, this is replaced with regard to materials by possible failure mechanisms like physical/chemical reaction mechanisms, disturbances of physical or chemical equilibrium, or other interactions or changes of system. The main tasks of the reliability analysis of materials therefore is the prediction of the various failure reasons, the identification of interactions, and the development of nondestructive testing methods. (RW) [de

  9. Structural Reliability Methods

    DEFF Research Database (Denmark)

    Ditlevsen, Ove Dalager; Madsen, H. O.

    The structural reliability methods quantitatively treat the uncertainty of predicting the behaviour and properties of a structure given the uncertain properties of its geometry, materials, and the actions it is supposed to withstand. This book addresses the probabilistic methods for evaluation...... of structural reliability, including the theoretical basis for these methods. Partial safety factor codes under current practice are briefly introduced and discussed. A probabilistic code format for obtaining a formal reliability evaluation system that catches the most essential features of the nature...... of the uncertainties and their interplay is the developed, step-by-step. The concepts presented are illustrated by numerous examples throughout the text....

  10. Reliability and mechanical design

    International Nuclear Information System (INIS)

    Lemaire, Maurice

    1997-01-01

    A lot of results in mechanical design are obtained from a modelisation of physical reality and from a numerical solution which would lead to the evaluation of needs and resources. The goal of the reliability analysis is to evaluate the confidence which it is possible to grant to the chosen design through the calculation of a probability of failure linked to the retained scenario. Two types of analysis are proposed: the sensitivity analysis and the reliability analysis. Approximate methods are applicable to problems related to reliability, availability, maintainability and safety (RAMS)

  11. RTE - 2013 Reliability Report

    International Nuclear Information System (INIS)

    Denis, Anne-Marie

    2014-01-01

    RTE publishes a yearly reliability report based on a standard model to facilitate comparisons and highlight long-term trends. The 2013 report is not only stating the facts of the Significant System Events (ESS), but it moreover underlines the main elements dealing with the reliability of the electrical power system. It highlights the various elements which contribute to present and future reliability and provides an overview of the interaction between the various stakeholders of the Electrical Power System on the scale of the European Interconnected Network. (author)

  12. SMRT has tissue-specific isoform profiles that include a form containing one CoRNR box

    International Nuclear Information System (INIS)

    Short, Stephen; Malartre, Marianne; Sharpe, Colin

    2005-01-01

    SMRT acts as a corepressor for a range of transcription factors. The amino-terminal part of the protein includes domains that mainly mediate transcriptional repression whilst the carboxy-terminal part includes domains that interact with nuclear receptors using up to three motifs called CoRNR boxes. The region of the SMRT primary transcript encoding the interaction domains is subject to alternative splicing that varies the inclusion of the third CoRNR box. The profile in mice includes an abundant, novel SMRT isoform that possesses just one CoRNR box. Mouse tissues therefore express SMRT isoforms containing one, two or three CoRNR boxes. In frogs, the SMRT isoform profile is tissue-specific. The mouse also shows distinct profiles generated by differential expression levels of the SMRT transcript isoforms. The formation of multiple SMRT isoforms and their tissue-specific regulation indicates a mechanism, whereby cells can define the repertoire of transcription factors regulated by SMRT

  13. The characterization of soybean oil body integral oleosin isoforms and the effects of alkaline pH on them.

    Science.gov (United States)

    Cao, Yanyun; Zhao, Luping; Ying, Yusang; Kong, Xiangzhen; Hua, Yufei; Chen, Yeming

    2015-06-15

    Oil body, an organelle in seed cell (naturally pre-emulsified oil), has great potentials to be used in food, cosmetics, pharmaceutical and other applications requiring stable oil-in-water emulsions. Researchers have tried to extract oil body by alkaline buffers, which are beneficial for removing contaminated proteins. But it is not clear whether alkaline buffers could remove oil body integral proteins (mainly oleosins), which could keep oil body integrity and stability. In this study, seven oleosin isoforms were identified for soybean oil body (three isoforms, 24 kDa; three isoforms, 18 kDa; one isoform, 16kDa). Oleosins were not glycoproteins and 24 kDa oleosin isoforms possessed less thiol groups than 18 kDa ones. It was found that alkaline pH not only removed contaminated proteins but also oleosins, and more and more oleosins were removed with increasing alkaline pH. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. The fair value of operational reliability

    Energy Technology Data Exchange (ETDEWEB)

    Patino-Echeverri, Dalia; Morel, Benoit

    2005-12-15

    Information about the uncertainties that surround the operation of the power system can be used to enlighten the debate of how much reliability should be pursued and how resources should be allocated to pursue it. In this paper we present a method to determine the value of having flexible generators to react to load fluctuations. This value can be seen as the value of hedging against the uncertainty on the load due to the volatility of the demand and the possibility of congestion. Because having this flexibility can be related to a financial option, we will use an extension of options theory and in particular the risk-neutral valuation method, to find a risk neutral quantification of its value. We illustrate our point valuing the flexibility that leads to ''operational reliability'' in the PJM market. Our formula for that value is what we call ''the fair value'' of operational reliability. (Author)

  15. Sensitivity analysis in a structural reliability context

    International Nuclear Information System (INIS)

    Lemaitre, Paul

    2014-01-01

    This thesis' subject is sensitivity analysis in a structural reliability context. The general framework is the study of a deterministic numerical model that allows to reproduce a complex physical phenomenon. The aim of a reliability study is to estimate the failure probability of the system from the numerical model and the uncertainties of the inputs. In this context, the quantification of the impact of the uncertainty of each input parameter on the output might be of interest. This step is called sensitivity analysis. Many scientific works deal with this topic but not in the reliability scope. This thesis' aim is to test existing sensitivity analysis methods, and to propose more efficient original methods. A bibliographical step on sensitivity analysis on one hand and on the estimation of small failure probabilities on the other hand is first proposed. This step raises the need to develop appropriate techniques. Two variables ranking methods are then explored. The first one proposes to make use of binary classifiers (random forests). The second one measures the departure, at each step of a subset method, between each input original density and the density given the subset reached. A more general and original methodology reflecting the impact of the input density modification on the failure probability is then explored. The proposed methods are then applied on the CWNR case, which motivates this thesis. (author)

  16. Data Used in Quantified Reliability Models

    Science.gov (United States)

    DeMott, Diana; Kleinhammer, Roger K.; Kahn, C. J.

    2014-01-01

    Data is the crux to developing quantitative risk and reliability models, without the data there is no quantification. The means to find and identify reliability data or failure numbers to quantify fault tree models during conceptual and design phases is often the quagmire that precludes early decision makers consideration of potential risk drivers that will influence design. The analyst tasked with addressing a system or product reliability depends on the availability of data. But, where is does that data come from and what does it really apply to? Commercial industries, government agencies, and other international sources might have available data similar to what you are looking for. In general, internal and external technical reports and data based on similar and dissimilar equipment is often the first and only place checked. A common philosophy is "I have a number - that is good enough". But, is it? Have you ever considered the difference in reported data from various federal datasets and technical reports when compared to similar sources from national and/or international datasets? Just how well does your data compare? Understanding how the reported data was derived, and interpreting the information and details associated with the data is as important as the data itself.

  17. Approach to reliability assessment

    International Nuclear Information System (INIS)

    Green, A.E.; Bourne, A.J.

    1975-01-01

    Experience has shown that reliability assessments can play an important role in the early design and subsequent operation of technological systems where reliability is at a premium. The approaches to and techniques for such assessments, which have been outlined in the paper, have been successfully applied in variety of applications ranging from individual equipments to large and complex systems. The general approach involves the logical and systematic establishment of the purpose, performance requirements and reliability criteria of systems. This is followed by an appraisal of likely system achievment based on the understanding of different types of variational behavior. A fundamental reliability model emerges from the correlation between the appropriate Q and H functions for performance requirement and achievement. This model may cover the complete spectrum of performance behavior in all the system dimensions

  18. Comparison of inhibition capability of scutellarein and scutellarin towards important liver UDP-glucuronosyltransferase (UGT) isoforms.

    Science.gov (United States)

    Ma, Guang-You; Cao, Yun-Feng; Hu, Cui-Min; Fang, Zhong-Ze; Sun, Xiao-Yu; Hong, Mo; Zhu, Zhi-Tu

    2014-03-01

    Scutellarin is an important bioactive flavonoid extracted from Erigeron breviscapus (Vant.) Hand-Mazz, and scutellarein is the corresponding aglycone of scutellarin. The present study aims to compare the inhibition potential of scutellarin and scutellarein towards several important UDP-glucuronosyltransferase (UGT) isoforms, including UGT1A1, UGT1A6, UGT1A9 and UGT2B7. It was demonstrated that scutellarein exerted stronger inhibition towards the tested UGT isoforms than scutellarin. Furthermore, the inhibition kinetic type and parameters (Ki ) were determined for the scutellarein's inhibition towards these UGT isoforms. Competitive inhibition of scutellarein towards all these UGT isoforms was demonstrated, and the Ki values were calculated to be 0.02, 5.0, 5.8 and 35.9 μM for UGT1A1, 1A6, 1A9 and 2B7, respectively. Using in vivo maximum plasma concentration of scutellarein in rat, the in vitro-in vivo extrapolation was performed to predict in vivo situation, indicating the most possible in vivo adverse effects due to the inhibition of scutellarein towards UGT1A1. All these results remind us to monitor the utilization of scutellarin and scutellarein, and the herbs containing these two components. Copyright © 2013 John Wiley & Sons, Ltd.

  19. Frataxin mRNA Isoforms in FRDA Patients and Normal Subjects: Effect of Tocotrienol Supplementation

    Directory of Open Access Journals (Sweden)

    Provvidenza Maria Abruzzo

    2013-01-01

    Full Text Available Friedreich’s ataxia (FRDA is caused by deficient expression of the mitochondrial protein frataxin involved in the formation of iron-sulphur complexes and by consequent oxidative stress. We analysed low-dose tocotrienol supplementation effects on the expression of the three splice variant isoforms (FXN-1, FXN-2, and FXN-3 in mononuclear blood cells of FRDA patients and healthy subjects. In FRDA patients, tocotrienol leads to a specific and significant increase of FXN-3 expression while not affecting FXN-1 and FXN-2 expression. Since no structural and functional details were available for FNX-2 and FXN-3, 3D models were built. FXN-1, the canonical isoform, was then docked on the human iron-sulphur complex, and functional interactions were computed; when FXN-1 was replaced by FXN-2 or FNX-3, we found that the interactions were maintained, thus suggesting a possible biological role for both isoforms in human cells. Finally, in order to evaluate whether tocotrienol enhancement of FXN-3 was mediated by an increase in peroxisome proliferator-activated receptor-γ (PPARG, PPARG expression was evaluated. At a low dose of tocotrienol, the increase of FXN-3 expression appeared to be independent of PPARG expression. Our data show that it is possible to modulate the mRNA expression of the minor frataxin isoforms and that they may have a functional role.

  20. HPLC separation of human serum albumin isoforms based on their isoelectric points

    Science.gov (United States)

    Bonilla, Lucía; Torres, María José; Schopfer, Francisco; Freeman, Bruce A.; Armas, Larissa; Ricciardi, Alejandro; Alvarez, Beatriz; Radi, Rafael

    2014-01-01

    Human serum albumin (HSA) is the most abundant protein in plasma. Cys34, the only free Cys residue, is the predominant plasma thiol and a relevant sacrificial antioxidant. Both in vivo circulating HSA and pharmaceutical preparations are heterogeneous with respect to the oxidation state of Cys34. In this work, we developed an external pH gradient chromatofocusing procedure that allows the analysis of the oxidation status of HSA in human plasma and biopharmaceutical products based on the different apparent isoelectric points and chemical properties of the redox isoforms. Specifically, reduced-mercury blocked HSA (HSA–SHg+), HSA with Cys34 oxidized to sulfenic acid (HSA–SOH) and HSA oxidized to sulfinate anion (HSA–SO2−) can be separated with resolutions of 1.4 and 3.1 (first and last pair) and hence quantified and purified. In addition, an N-terminally degraded isoform (HSA3–585) in different redox states can be resolved as well. Confirmation of the identity of the chromatofocusing isolated isoforms was achieved by high resolution whole protein MS. It is proposed that the chromatofocusing procedure can be used to produce more exact and complete descriptions of the redox status of HSA in vivo and in vitro. Finally, the scalability capabilities of the chromatofocusing procedure allow for the preparation of highly pure standards of several redox isoforms of HSA PMID:24316526

  1. Isoform 1 of TPD52 (PC-1) promotes neuroendocrine transdifferentiation in prostate cancer cells

    KAUST Repository

    Moritz, Tom; Venz, Simone; Junker, Heike; Kreuz, Sarah; Walther, Reinhard; Zimmermann, Uwe

    2016-01-01

    The tumour protein D52 isoform 1 (PC-1), a member of the tumour protein D52 (TPD52) protein family, is androgen-regulated and prostate-specific expressed. Previous studies confirmed that PC-1 contributes to malignant progression in prostate cancer

  2. Generating Isoform-Specific Antibodies : Lessons from Nucleocytoplasmic Glycoprotein Skp1

    NARCIS (Netherlands)

    West, Christopher M.; Van Der Wel, Hanke; Chinoy, Zoiesha; Boons, Geert Jan; Gauthier, Ted J.; Taylor, Carol M.; Xu, Yuechi

    2015-01-01

    Antibodies that discriminate protein isoforms differing by modifications at specific amino acids have revolutionized studies of their functions. Skp1 is a novel nucleocytoplasmic glycoprotein that is hydroxylated at proline-143 and then O-glycosylated by a pentasaccharide attached via a GlcNAcα1,

  3. Muscle-Type Specific Autophosphorylation of CaMKII Isoforms after Paced Contractions

    NARCIS (Netherlands)

    Eilers, W.; Gevers, W.; van Overbeek, D.; de Haan, A.; Jaspers, R.T.; Hilbers, P.A.; van Riel, A.C.R.; Flueck, M.

    2014-01-01

    We explored to what extent isoforms of the regulator of excitation-contraction and excitation-transcription coupling, calcium/calmodulin protein kinase II (CaMKII) contribute to the specificity of myocellular calcium sensing between muscle types and whether concentration transients in its

  4. Distribution of protein kinase Mzeta and the complete protein kinase C isoform family in rat brain

    DEFF Research Database (Denmark)

    Naik, M U; Benedikz, Eirikur; Hernandez, I

    2000-01-01

    Protein kinase C (PKC) is a multigene family of at least ten isoforms, nine of which are expressed in brain (alpha, betaI, betaII, gamma, delta, straightepsilon, eta, zeta, iota/lambda). Our previous studies have shown that many of these PKCs participate in synaptic plasticity in the CA1 region...

  5. Differential regulation by AMP and ADP of AMPK complexes containing different γ subunit isoforms

    DEFF Research Database (Denmark)

    Ross, Fiona A; Jensen, Thomas Elbenhardt; Hardie, D Grahame

    2016-01-01

    The g subunits of heterotrimeric AMPK complexes contain the binding sites for the regulatory adenine nucleotides AMP, ADP and ATP. We addressed whether complexes containing different g isoforms display different responses to adenine nucleotides by generating cells stably expressing FLAG-tagged ve...

  6. Biochemical Characteristics of Three Laccase Isoforms from the Basidiomycete Pleurotus nebrodensis

    Directory of Open Access Journals (Sweden)

    Xianghe Yuan

    2016-02-01

    Full Text Available The characterization of three laccase isoforms from Pleurotus nebrodensis is described. Isoenzymes Lac1, Lac2 and Lac3 were purified to homogeneity using ion exchange chromatography on DEAE-cellulose, CM-cellulose and Q-Sepharose and a gel filtration step on Superdex 75. The molecular weights of the purified laccases were estimated to be 68, 64 and 51 kDa, respectively. The isoenzymes demonstrated the same optimum pH at 3.0 but slightly different temperature optima: 50–60 °C for Lac1 and Lac3 and 60 °C for Lac2. Lac2 was always more stable than the other two isoforms and exposure to 50 °C for 120 min caused 30% loss in activity. Lac2 was relatively less stable than the other two isoforms when exposed to the pH range of 3.0–8.0 for 24 h, but inactivation only occurred initially, with around 70% residual activity being maintained during the whole process. Oxidative ability towards aromatic compounds varied substantially among the isoforms and each of them displayed preference toward some substrates. Kinetic constants (Km, Kcat were determined by using a 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid diammonium salt (ABTS assay, with Lac3 showing the best affinity and Lac2 displaying the highest catalytic efficiency. Amino acid sequences from peptides derived from digestion of isoenzymes showed great consistency with laccases in the databases.

  7. Quarternary structure and enzymological properties of the different hormone-sensitive lipase (HSL) isoforms

    DEFF Research Database (Denmark)

    Krintel, Christian; Klint, Cecilia; Lindvall, Håkan

    2010-01-01

    Hormone-sensitive lipase (HSL) is a key enzyme in the mobilization of energy in the form of fatty acids from intracellular stores of neutral lipids. The enzyme has been shown to exist in different isoforms with different molecular masses (84 kDa, 89 kDa and 117 kDa) expressed in a tissue-dependen...

  8. Role of the AMPKgamma3 isoform in hypoxia-stimulated glucose transport in glycolytic skeletal muscle

    DEFF Research Database (Denmark)

    Deshmukh, Atul S; Glund, Stephan; Tom, Robby Z

    2009-01-01

    , phosphorylation of CaMKII, AMPK, ACC, and TBC1D1/D4 as well as isoform-specific AMPK activity was determined. Basal and hypoxia-mediated phosphorylation of CaMKII, AMPK, and ACC as well as alpha1- and alpha2-associated AMPK activity was comparable between AMPKgamma3-KO and wild-type mice. KN-93 reduced hypoxia...

  9. Over-expression in Escherichia coli and characterization of two recombinant isoforms of human FAD synthetase

    International Nuclear Information System (INIS)

    Brizio, Carmen; Galluccio, Michele; Wait, Robin; Torchetti, Enza Maria; Bafunno, Valeria; Accardi, Rosita; Gianazza, Elisabetta; Indiveri, Cesare; Barile, Maria

    2006-01-01

    FAD synthetase (FADS) (EC 2.7.7.2) is a key enzyme in the metabolic pathway that converts riboflavin into the redox cofactor FAD. Two hypothetical human FADSs, which are the products of FLAD1 gene, were over-expressed in Escherichia coli and identified by ESI-MS/MS. Isoform 1 was over-expressed as a T7-tagged protein which had a molecular mass of 63 kDa on SDS-PAGE. Isoform 2 was over-expressed as a 6-His-tagged fusion protein, carrying an extra 84 amino acids at the N-terminal with an apparent molecular mass of 60 kDa on SDS-PAGE. It was purified near to homogeneity from the soluble cell fraction by one-step affinity chromatography. Both isoforms possessed FADS activity and had a strict requirement for MgCl 2 , as demonstrated using both spectrophotometric and chromatographic methods. The purified recombinant isoform 2 showed a specific activity of 6.8 ± 1.3 nmol of FAD synthesized/min/mg protein and exhibited a K M value for FMN of 1.5 ± 0.3 μM. This is First report on characterization of human FADS, and First cloning and over-expression of FADS from an organism higher than yeast

  10. TGF-β's delay skeletal muscle progenitor cell differentiation in an isoform-independent manner

    International Nuclear Information System (INIS)

    Schabort, Elske J.; Merwe, Mathilde van der; Loos, Benjamin; Moore, Frances P.; Niesler, Carola U.

    2009-01-01

    Satellite cells are a quiescent heterogenous population of mononuclear stem and progenitor cells which, once activated, differentiate into myotubes and facilitate skeletal muscle repair or growth. The Transforming Growth Factor-β (TGF-β) superfamily members are elevated post-injury and their importance in the regulation of myogenesis and wound healing has been demonstrated both in vitro and in vivo. Most studies suggest a negative role for TGF-β on satellite cell differentiation. However, none have compared the effect of these three isoforms on myogenesis in vitro. This is despite known isoform-specific effects of TGF-β1, -β2 and -β3 on wound repair in other tissues. In the current study we compared the effect of TGF-β1, -β2 and -β3 on proliferation and differentiation of the C2C12 myoblast cell-line. We found that, irrespective of the isoform, TGF-β increased proliferation of C2C12 cells by changing the cellular localisation of PCNA to promote cell division and prevent cell cycle exit. Concomitantly, TGF-β1, -β2 and -β3 delayed myogenic commitment by increasing MyoD degradation and decreasing myogenin expression. Terminal differentiation, as measured by a decrease in myosin heavy chain (MHC) expression, was also delayed. These results demonstrate that TGF-β promotes proliferation and delays differentiation of C2C12 myoblasts in an isoform-independent manner

  11. Thick filament length and isoform composition determine self-organized contractile units in actomyosin bundles.

    Science.gov (United States)

    Thoresen, Todd; Lenz, Martin; Gardel, Margaret L

    2013-02-05

    Diverse myosin II isoforms regulate contractility of actomyosin bundles in disparate physiological processes by variations in both motor mechanochemistry and the extent to which motors are clustered into thick filaments. Although the role of mechanochemistry is well appreciated, the extent to which thick filament length regulates actomyosin contractility is unknown. Here, we study the contractility of minimal actomyosin bundles formed in vitro by mixtures of F-actin and thick filaments of nonmuscle, smooth, and skeletal muscle myosin isoforms with varied length. Diverse myosin II isoforms guide the self-organization of distinct contractile units within in vitro bundles with shortening rates similar to those of in vivo myofibrils and stress fibers. The tendency to form contractile units increases with the thick filament length, resulting in a bundle shortening rate proportional to the length of constituent myosin thick filament. We develop a model that describes our data, providing a framework in which to understand how diverse myosin II isoforms regulate the contractile behaviors of disordered actomyosin bundles found in muscle and nonmuscle cells. These experiments provide insight into physiological processes that use dynamic regulation of thick filament length, such as smooth muscle contraction. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  12. Neuronal Glucose Transporter Isoform 3 Deficient Mice Demonstrate Features of Autism Spectrum Disorders

    OpenAIRE

    Zhao, Yuanzi; Fung, Camille; Shin, Don; Shin, Bo-Chul; Thamotharan, Shanthie; Sankar, Raman; Ehninger, Dan; Silva, Alcino; Devaskar, Sherin U.

    2009-01-01

    Neuronal glucose transporter (GLUT) isoform 3 deficiency in null heterozygous mice led to abnormal spatial learning and working memory but normal acquisition and retrieval during contextual conditioning, abnormal cognitive flexibility with intact gross motor ability, electroencephalographic seizures, perturbed social behavior with reduced vocalization and stereotypies at low frequency. This phenotypic expression is unique as it combines the neurobehavioral with the epileptiform characteristic...

  13. Alternative NF-κB Isoforms in the Drosophila Neuromuscular Junction and Brain.

    Directory of Open Access Journals (Sweden)

    Bo Zhou

    Full Text Available The Drosophila NF-κB protein Dorsal is expressed at the larval neuromuscular junction, where its expression appears unrelated to known Dorsal functions in embryonic patterning and innate immunity. Using confocal microscopy with domain-specific antisera, we demonstrate that larval muscle expresses only the B isoform of Dorsal, which arises by intron retention. We find that Dorsal B interacts with and stabilizes Cactus at the neuromuscular junction, but exhibits Cactus independent localization and an absence of detectable nuclear translocation. We further find that the Dorsal-related immune factor Dif encodes a B isoform, reflecting a conservation of B domains across a range of insect NF-κB proteins. Carrying out mutagenesis of the Dif locus via a site-specific recombineering approach, we demonstrate that Dif B is the major, if not sole, Dif isoform in the mushroom bodies of the larval brain. The Dorsal and Dif B isoforms thus share a specific association with nervous system tissues as well as an alternative protein structure.

  14. Bilirubin UDP-glucuronosyltransferase 1 is the only relevant bilirubin glucuronidating isoform in man

    NARCIS (Netherlands)

    Bosma, P. J.; Seppen, J.; Goldhoorn, B.; Bakker, C.; Oude Elferink, R. P.; Chowdhury, J. R.; Chowdhury, N. R.; Jansen, P. L.

    1994-01-01

    Crigler-Najjar syndrome type I (CN-I) is caused by an inherited absence of UDP-glucuronosyltransferase activity toward bilirubin (B-UGT), resulting in severe non-hemolytic unconjugated hyperbilirubinemia. Based on the expression of cDNAs in COS cells, two UGT isoforms in human liver, B-UGT1 and

  15. Identification of a novel CoA synthase isoform, which is primarily expressed in Brain

    International Nuclear Information System (INIS)

    Nemazanyy, Ivan; Panasyuk, Ganna; Breus, Oksana; Zhyvoloup, Alexander; Filonenko, Valeriy; Gout, Ivan T.

    2006-01-01

    CoA and its derivatives Acetyl-CoA and Acyl-CoA are important players in cellular metabolism and signal transduction. CoA synthase is a bifunctional enzyme which mediates the final stages of CoA biosynthesis. In previous studies, we have reported molecular cloning, biochemical characterization, and subcellular localization of CoA synthase (CoASy). Here, we describe the existence of a novel CoA synthase isoform, which is the product of alternative splicing and possesses a 29aa extension at the N-terminus. We termed it CoASy β and originally identified CoA synthase, CoASy α. The transcript specific for CoASy β was identified by electronic screening and by RT-PCR analysis of various rat tissues. The existence of this novel isoform was further confirmed by immunoblot analysis with antibodies directed to the N-terminal peptide of CoASy β. In contrast to CoASy α, which shows ubiquitous expression, CoASy β is primarily expressed in Brain. Using confocal microscopy, we demonstrated that both isoforms are localized on mitochondria. The N-terminal extension does not affect the activity of CoA synthase, but possesses a proline-rich sequence which can bring the enzyme into complexes with signalling proteins containing SH3 or WW domains. The role of this novel isoform in CoA biosynthesis, especially in Brain, requires further elucidation

  16. The novel protein kinase C epsilon isoform modulates acetylcholine release in the rat neuromuscular junction.

    Science.gov (United States)

    Obis, Teresa; Hurtado, Erica; Nadal, Laura; Tomàs, Marta; Priego, Mercedes; Simon, Anna; Garcia, Neus; Santafe, Manel M; Lanuza, Maria A; Tomàs, Josep

    2015-12-01

    Various protein kinase C (PKC) isoforms contribute to the phosphorylating activity that modulates neurotransmitter release. In previous studies we showed that nPKCε is confined in the presynaptic site of the neuromuscular junction and its presynaptic function is activity-dependent. Furthermore, nPKCε regulates phorbol ester-induced acetylcholine release potentiation, which further indicates that nPKCε is involved in neurotransmission. The present study is designed to examine the nPKCε involvement in transmitter release at the neuromuscular junction. We use the specific nPKCε translocation inhibitor peptide εV1-2 and electrophysiological experiments to investigate the involvement of this isoform in acetylcholine release. We observed that nPKCε membrane translocation is key to the synaptic potentiation of NMJ, being involved in several conditions that upregulate PKC isoforms coupling to acetylcholine (ACh) release (incubation with high Ca(2+), stimulation with phorbol esters and protein kinase A, stimulation with adenosine 3',5'-cyclic monophosphorothioate, 8-Bromo-, Rp-isomer, sodium salt -Sp-8-BrcAMP-). In all these conditions, preincubation with the nPKCε translocation inhibitor peptide (εV1-2) impairs PKC coupling to acetylcholine release potentiation. In addition, the inhibition of nPKCε translocation and therefore its activity impedes that presynaptic muscarinic autoreceptors and adenosine autoreceptors modulate transmitter secretion. Together, these results point to the importance of nPKCε isoform in the control of acetylcholine release in the neuromuscular junction.

  17. The Invasion and Metastasis Promotion Role of CD97 Small Isoform in Gastric Carcinoma

    DEFF Research Database (Denmark)

    Liu, Daren; Trojanowicz, Bogusz; Ye, Longyun

    2012-01-01

    CD97 is over-expressed in the majority of gastric adenocarcinomas and is associated with its dedifferentiation and aggressiveness. Our previous results demonstrated that out of three CD97 isoforms tested, only the small one was able to promote increased invasiveness in vitro. Based on these data ...

  18. Roles of SGK Isoform Signaling in Breast Cancer Migration and Invasion

    Science.gov (United States)

    2011-04-01

    significant number of overlapping substrates, and deregulation in breast carcinoma (5,6). To date, no studies have investigated any role for SGK in cell...isoforms in breast carcinoma cell lines (months 2-3) To insure specificity of SGK knockdown in breast cancer cell lines I made two different specific

  19. The activity and isoforms of NADP-malic enzyme in Nicotiana benthamiana plants under biotic stress

    Czech Academy of Sciences Publication Activity Database

    Doubnerová, V.; Jirásková, A.; Janošková, M.; Müller, Karel; Baťková, Petra; Synková, Helena; Čeřovská, Noemi; Ryšlavá, H.

    2007-01-01

    Roč. 26, č. 4 (2007), s. 281-289 ISSN 0231-5882 Institutional research plan: CEZ:AV0Z50380511 Keywords : NADP * malic enzyme isoforms * Nicotiana benthamiana Subject RIV: EF - Botanics Impact factor: 1.286, year: 2007 http://www.gpb.sav.sk/2007-4.htm

  20. Opioid precursor protein isoform is targeted to the cell nuclei in the human brain

    DEFF Research Database (Denmark)

    Kononenko, Olga; Bazov, Igor; Watanabe, Hiroyuki

    2017-01-01

    to the cell nuclei in a model cellular system. This may be driven by bipartite nuclear localization signal (NLS) that is cryptic in the full-length PDYN molecule and becomes functional when signal peptide is truncated. Nuclear PDYN isoform was identified by western blot and radioimmunoassay in neuronal nuclei...

  1. Cryptocephal, the Drosophila melanogaster ATF4, is a specific coactivator for ecdysone receptor isoform B2.

    Directory of Open Access Journals (Sweden)

    Sebastien A Gauthier

    Full Text Available The ecdysone receptor is a heterodimer of two nuclear receptors, the Ecdysone receptor (EcR and Ultraspiracle (USP. In Drosophila melanogaster, three EcR isoforms share common DNA and ligand-binding domains, but these proteins differ in their most N-terminal regions and, consequently, in the activation domains (AF1s contained therein. The transcriptional coactivators for these domains, which impart unique transcriptional regulatory properties to the EcR isoforms, are unknown. Activating transcription factor 4 (ATF4 is a basic-leucine zipper transcription factor that plays a central role in the stress response of mammals. Here we show that Cryptocephal (CRC, the Drosophila homolog of ATF4, is an ecdysone receptor coactivator that is specific for isoform B2. CRC interacts with EcR-B2 to promote ecdysone-dependent expression of ecdysis-triggering hormone (ETH, an essential regulator of insect molting behavior. We propose that this interaction explains some of the differences in transcriptional properties that are displayed by the EcR isoforms, and similar interactions may underlie the differential activities of other nuclear receptors with distinct AF1-coactivators.

  2. A novel MCPH1 isoform complements the defective chromosome condensation of human MCPH1-deficient cells.

    Directory of Open Access Journals (Sweden)

    Ioannis Gavvovidis

    Full Text Available Biallelic mutations in MCPH1 cause primary microcephaly (MCPH with the cellular phenotype of defective chromosome condensation. MCPH1 encodes a multifunctional protein that notably is involved in brain development, regulation of chromosome condensation, and DNA damage response. In the present studies, we detected that MCPH1 encodes several distinct transcripts, including two major forms: full-length MCPH1 (MCPH1-FL and a second transcript lacking the six 3' exons (MCPH1Δe9-14. Both variants show comparable tissue-specific expression patterns, demonstrate nuclear localization that is mediated independently via separate NLS motifs, and are more abundant in certain fetal than adult organs. In addition, the expression of either isoform complements the chromosome condensation defect found in genetically MCPH1-deficient or MCPH1 siRNA-depleted cells, demonstrating a redundancy of both MCPH1 isoforms for the regulation of chromosome condensation. Strikingly however, both transcripts are regulated antagonistically during cell-cycle progression and there are functional differences between the isoforms with regard to the DNA damage response; MCPH1-FL localizes to phosphorylated H2AX repair foci following ionizing irradiation, while MCPH1Δe9-14 was evenly distributed in the nucleus. In summary, our results demonstrate here that MCPH1 encodes different isoforms that are differentially regulated at the transcript level and have different functions at the protein level.

  3. The rating reliability calculator

    Directory of Open Access Journals (Sweden)

    Solomon David J

    2004-04-01

    Full Text Available Abstract Background Rating scales form an important means of gathering evaluation data. Since important decisions are often based on these evaluations, determining the reliability of rating data can be critical. Most commonly used methods of estimating reliability require a complete set of ratings i.e. every subject being rated must be rated by each judge. Over fifty years ago Ebel described an algorithm for estimating the reliability of ratings based on incomplete data. While his article has been widely cited over the years, software based on the algorithm is not readily available. This paper describes an easy-to-use Web-based utility for estimating the reliability of ratings based on incomplete data using Ebel's algorithm. Methods The program is available public use on our server and the source code is freely available under GNU General Public License. The utility is written in PHP, a common open source imbedded scripting language. The rating data can be entered in a convenient format on the user's personal computer that the program will upload to the server for calculating the reliability and other statistics describing the ratings. Results When the program is run it displays the reliability, number of subject rated, harmonic mean number of judges rating each subject, the mean and standard deviation of the averaged ratings per subject. The program also displays the mean, standard deviation and number of ratings for each subject rated. Additionally the program will estimate the reliability of an average of a number of ratings for each subject via the Spearman-Brown prophecy formula. Conclusion This simple web-based program provides a convenient means of estimating the reliability of rating data without the need to conduct special studies in order to provide complete rating data. I would welcome other researchers revising and enhancing the program.

  4. Structural systems reliability analysis

    International Nuclear Information System (INIS)

    Frangopol, D.

    1975-01-01

    For an exact evaluation of the reliability of a structure it appears necessary to determine the distribution densities of the loads and resistances and to calculate the correlation coefficients between loads and between resistances. These statistical characteristics can be obtained only on the basis of a long activity period. In case that such studies are missing the statistical properties formulated here give upper and lower bounds of the reliability. (orig./HP) [de

  5. Reliability and maintainability

    International Nuclear Information System (INIS)

    1994-01-01

    Several communications in this conference are concerned with nuclear plant reliability and maintainability; their titles are: maintenance optimization of stand-by Diesels of 900 MW nuclear power plants; CLAIRE: an event-based simulation tool for software testing; reliability as one important issue within the periodic safety review of nuclear power plants; design of nuclear building ventilation by the means of functional analysis; operation characteristic analysis for a power industry plant park, as a function of influence parameters

  6. Reliability data book

    International Nuclear Information System (INIS)

    Bento, J.P.; Boerje, S.; Ericsson, G.; Hasler, A.; Lyden, C.O.; Wallin, L.; Poern, K.; Aakerlund, O.

    1985-01-01

    The main objective for the report is to improve failure data for reliability calculations as parts of safety analyses for Swedish nuclear power plants. The work is based primarily on evaluations of failure reports as well as information provided by the operation and maintenance staff of each plant. In the report are presented charts of reliability data for: pumps, valves, control rods/rod drives, electrical components, and instruments. (L.E.)

  7. Selective expression of myosin IC Isoform A in mouse and human cell lines and mouse prostate cancer tissues.

    Directory of Open Access Journals (Sweden)

    Ivanna Ihnatovych

    Full Text Available Myosin IC is a single headed member of the myosin superfamily. We recently identified a novel isoform and showed that the MYOIC gene in mammalian cells encodes three isoforms (isoforms A, B, and C. Furthermore, we demonstrated that myosin IC isoform A but not isoform B exhibits a tissue specific expression pattern. In this study, we extended our analysis of myosin IC isoform expression patterns by analyzing the protein and mRNA expression in various mammalian cell lines and in various prostate specimens and tumor tissues from the transgenic mouse prostate (TRAMP model by immunoblotting, qRT-PCR, and by indirect immunohistochemical staining of paraffin embedded prostate specimen. Analysis of a panel of mammalian cell lines showed an increased mRNA and protein expression of specifically myosin IC isoform A in a panel of human and mouse prostate cancer cell lines but not in non-cancer prostate or other (non-prostate- cancer cell lines. Furthermore, we demonstrate that myosin IC isoform A expression is significantly increased in TRAMP mouse prostate samples with prostatic intraepithelial neoplasia (PIN lesions and in distant site metastases in lung and liver when compared to matched normal tissues. Our observations demonstrate specific changes in the expression of myosin IC isoform A that are concurrent with the occurrence of prostate cancer in the TRAMP mouse prostate cancer model that closely mimics clinical prostate cancer. These data suggest that elevated levels of myosin IC isoform A may be a potential marker for the detection of prostate cancer.

  8. Multidisciplinary System Reliability Analysis

    Science.gov (United States)

    Mahadevan, Sankaran; Han, Song; Chamis, Christos C. (Technical Monitor)

    2001-01-01

    The objective of this study is to develop a new methodology for estimating the reliability of engineering systems that encompass multiple disciplines. The methodology is formulated in the context of the NESSUS probabilistic structural analysis code, developed under the leadership of NASA Glenn Research Center. The NESSUS code has been successfully applied to the reliability estimation of a variety of structural engineering systems. This study examines whether the features of NESSUS could be used to investigate the reliability of systems in other disciplines such as heat transfer, fluid mechanics, electrical circuits etc., without considerable programming effort specific to each discipline. In this study, the mechanical equivalence between system behavior models in different disciplines are investigated to achieve this objective. A new methodology is presented for the analysis of heat transfer, fluid flow, and electrical circuit problems using the structural analysis routines within NESSUS, by utilizing the equivalence between the computational quantities in different disciplines. This technique is integrated with the fast probability integration and system reliability techniques within the NESSUS code, to successfully compute the system reliability of multidisciplinary systems. Traditional as well as progressive failure analysis methods for system reliability estimation are demonstrated, through a numerical example of a heat exchanger system involving failure modes in structural, heat transfer and fluid flow disciplines.

  9. Kalrn promoter usage and isoform expression respond to chronic cocaine exposure

    Directory of Open Access Journals (Sweden)

    Ma Xin-Ming

    2011-02-01

    Full Text Available Abstract Background The long-term effects of cocaine on behavior are accompanied by structural changes in excitatory glutamatergic synapses onto the medium spiny neurons of the striatum. The Kalrn gene encodes several functionally distinct isoforms; these multidomain guanine nucleotide exchange factors (GEFs contain additional domains known to interact with phosphatidylinositides as well as with a number of different proteins. Through their activation of Rho proteins and their interactions with other proteins, the different Kalirin isoforms affect cytoskeletal organization. Chronic exposure of adult male rodents to cocaine increases levels of Kalirin 7 in the striatum. When exposed chronically to cocaine, mice lacking Kalirin 7, the major adult isoform, fail to show an increase in dendritic spine density in the nucleus accumbens, show diminished place preference for cocaine, and exhibit increased locomotor activity in response to cocaine. Results The use of alternate promoters and 3'-terminal exons of the mouse Kalrn gene were investigated using real-time quantitative polymerase chain reaction. While the two most distal full-length Kalrn promoters are used equally in the prefrontal cortex, the more proximal of these promoters accounts for most of the transcripts expressed in the nucleus accumbens. The 3'-terminal exon unique to the Kalirin 7 isoform accounts for a greater percentage of the Kalrn transcripts in prefrontal cortex than in nucleus accumbens. Western blot analyses confirmed these differences. Chronic cocaine treatment increases usage of the promoter encoding the Δ-Kalirin isoforms but does not alter full-length Kalirin promoter usage. Usage of the 3'-terminal exon unique to Kalirin 7 increases following chronic cocaine exposure. Conclusions Kalrn promoter and 3'-terminal exon utilization are region-specific. In the nucleus accumbens, cocaine-mediated alterations in promoter usage and 3'-terminal exon usage favor expression of

  10. Redundancy of IL-1 Isoform Signaling and Its Implications for Arterial Remodeling.

    Directory of Open Access Journals (Sweden)

    Marina Beltrami-Moreira

    Full Text Available Mice deficient in IL-1 receptor 1 (hence unresponsive to both IL-1 isoforms α and β have impaired expansive arterial remodeling due to diminished expression of matrix-degrading enzymes, especially MMP-3. Emergence of IL-1 as a target in cardiovascular disease prompted the investigation of the redundancy of IL-1α and IL-1β in the induction of MMP-3 and other matrix-remodeling enzymes in human cells.Human primary vascular smooth muscle cells (VSMCs and carotid endarterectomy specimens were stimulated with equimolar concentrations of IL-1α or IL-1β and analyzed protease expression by immunoblot and ELISA. Either IL-1α or IL-1β increased the expression of pro-MMP-3 in VSMCs, facilitated VSMC migration through Matrigel, and induced MMP-3 production in specimens from atheromatous plaques. VSMCs also secreted MMP-1 and Cathepsin S (CatS upon stimulation with IL-1α or IL-1β. IL-1 isoforms similarly increased MMP-1 and MMP-9 expression in carotid endarterectomy specimens. We examined the expression of MMP-3 and IL-1 isoforms by immunostaining of carotid atheromata, calculated the % positive areas, and tested associations by linear regression. MMP-3 colocalized with IL-1 isoforms in atheromata. MMP-3+ area in plaques positively associated with IL-1α+ (R2 = 0.61, P<0.001 and with IL-1β + areas (R2 = 0.68, P<0.001. MMP-3+ area within atheroma also associated with CD68+ area, but not with α-smooth muscle actin area.Either IL-1α or IL-1β can induce the expression of enzymes implicated in remodeling of the arterial extracellular matrix, and facilitate human VSMC migration in vitro. Human atheromata contain both IL-1 isoforms in association with immunoreactive MMP-3. This redundancy of IL-1 isoforms suggests that selective blocking of one IL-1 isoform should not impair expansive arterial remodeling, a finding with important clinical implications for therapeutic targeting of IL-1 in atherosclerosis.

  11. Two Distinct Isoforms of Matrix Metalloproteinase-2 Are Associated with Human Delayed Kidney Graft Function.

    Directory of Open Access Journals (Sweden)

    Shaynah Wanga

    Full Text Available Delayed graft function (DGF is a frequent complication of renal transplantation, particularly in the setting of transplantation of kidneys derived from deceased donors and expanded-criteria donors. DGF results from tubular epithelial cell injury and has immediate and long term consequences. These include requirement for post-transplantation dialysis, increased incidence of acute rejection, and poorer long-term outcomes. DGF represents one of the clearest clinical examples of renal acute ischemia/reperfusion injury. Experimental studies have demonstrated that ischemia/reperfusion injury induces the synthesis of the full length secreted isoform of matrix metalloproteinase-2 (FL-MMP-2, as well as an intracellular N-terminal truncated MMP-2 isoform (NTT-MMP-2 that initiates an innate immune response. We hypothesized that the two MMP-2 isoforms mediate tubular epithelial cell injury in DGF. Archival renal biopsy sections from 10 protocol biopsy controls and 41 cases with a clinical diagnosis of DGF were analyzed for the extent of tubular injury, expression of the FL-MMP-2 and NTT-MMP-2 isoforms by immunohistochemistry (IHC, in situ hybridization, and qPCR to determine isoform abundance. Differences in transcript abundance were related to tubular injury score. Markers of MMP-2-mediated injury included TUNEL staining and assessment of peritubular capillary density. There was a clear relationship between tubular epithelial cell expression of both FL-MMP-2 and NTT-MMP-2 IHC with the extent of tubular injury. The MMP-2 isoforms were detected in the same tubular segments and were present at sites of tubular injury. qPCR demonstrated highly significant increases in both the FL-MMP-2 and NTT-MMP-2 transcripts. Statistical analysis revealed highly significant associations between FL-MMP-2 and NTT-MMP-2 transcript abundance and the extent of tubular injury, with NTT-MMP-2 having the strongest association. We conclude that two distinct MMP-2 isoforms are

  12. Analysis and Application of Reliability

    International Nuclear Information System (INIS)

    Jeong, Hae Seong; Park, Dong Ho; Kim, Jae Ju

    1999-05-01

    This book tells of analysis and application of reliability, which includes definition, importance and historical background of reliability, function of reliability and failure rate, life distribution and assumption of reliability, reliability of unrepaired system, reliability of repairable system, sampling test of reliability, failure analysis like failure analysis by FEMA and FTA, and cases, accelerated life testing such as basic conception, acceleration and acceleration factor, and analysis of accelerated life testing data, maintenance policy about alternation and inspection.

  13. Integrated Reliability and Risk Analysis System (IRRAS)

    International Nuclear Information System (INIS)

    Russell, K.D.; McKay, M.K.; Sattison, M.B.; Skinner, N.L.; Wood, S.T.; Rasmuson, D.M.

    1992-01-01

    The Integrated Reliability and Risk Analysis System (IRRAS) is a state-of-the-art, microcomputer-based probabilistic risk assessment (PRA) model development and analysis tool to address key nuclear plant safety issues. IRRAS is an integrated software tool that gives the user the ability to create and analyze fault trees and accident sequences using a microcomputer. This program provides functions that range from graphical fault tree construction to cut set generation and quantification. Version 1.0 of the IRRAS program was released in February of 1987. Since that time, many user comments and enhancements have been incorporated into the program providing a much more powerful and user-friendly system. This version has been designated IRRAS 4.0 and is the subject of this Reference Manual. Version 4.0 of IRRAS provides the same capabilities as Version 1.0 and adds a relational data base facility for managing the data, improved functionality, and improved algorithm performance

  14. Standardizing the practice of human reliability analysis

    International Nuclear Information System (INIS)

    Hallbert, B.P.

    1993-01-01

    The practice of human reliability analysis (HRA) within the nuclear industry varies greatly in terms of posited mechanisms that shape human performance, methods of characterizing and analytically modeling human behavior, and the techniques that are employed to estimate the frequency with which human error occurs. This variation has been a source of contention among HRA practitioners regarding the validity of results obtained from different HRA methods. It has also resulted in attempts to develop standard methods and procedures for conducting HRAs. For many of the same reasons, the practice of HRA has not been standardized or has been standardized only to the extent that individual analysts have developed heuristics and consistent approaches in their practice of HRA. From the standpoint of consumers and regulators, this has resulted in a lack of clear acceptance criteria for the assumptions, modeling, and quantification of human errors in probabilistic risk assessments

  15. Results of a Demonstration Assessment of Passive System Reliability Utilizing the Reliability Method for Passive Systems (RMPS)

    Energy Technology Data Exchange (ETDEWEB)

    Bucknor, Matthew; Grabaskas, David; Brunett, Acacia; Grelle, Austin

    2015-04-26

    Advanced small modular reactor designs include many advantageous design features such as passively driven safety systems that are arguably more reliable and cost effective relative to conventional active systems. Despite their attractiveness, a reliability assessment of passive systems can be difficult using conventional reliability methods due to the nature of passive systems. Simple deviations in boundary conditions can induce functional failures in a passive system, and intermediate or unexpected operating modes can also occur. As part of an ongoing project, Argonne National Laboratory is investigating various methodologies to address passive system reliability. The Reliability Method for Passive Systems (RMPS), a systematic approach for examining reliability, is one technique chosen for this analysis. This methodology is combined with the Risk-Informed Safety Margin Characterization (RISMC) approach to assess the reliability of a passive system and the impact of its associated uncertainties. For this demonstration problem, an integrated plant model of an advanced small modular pool-type sodium fast reactor with a passive reactor cavity cooling system is subjected to a station blackout using RELAP5-3D. This paper discusses important aspects of the reliability assessment, including deployment of the methodology, the uncertainty identification and quantification process, and identification of key risk metrics.

  16. Importance of independent and dependent human error to system reliability and plant safety

    International Nuclear Information System (INIS)

    Dach, K.

    1988-08-01

    Uncertainty analysis of the quantification of the unavailability for the emergency core cooling system was made. The reliability analysis of the low pressure injection system (LPIS) of the ECCS of WWER-440 reactor was also performed. Results of reliability analysis proved that LPIS reliability under normal conditions is sufficient and can be increased by two orders of magnitude. This increase in reliability can be achieved by means of simple changes such as securing an opening of the quick-acting fittings at LPIS discharge line. A method for analysis of systems uncertainty with periodic inspected components was elaborated and verified by performing an analysis of the medium size system. Refs, figs and tabs

  17. Deep Sequencing Reveals Uncharted Isoform Heterogeneity of the Protein-Coding Transcriptome in Cerebral Ischemia.

    Science.gov (United States)

    Bhattarai, Sunil; Aly, Ahmed; Garcia, Kristy; Ruiz, Diandra; Pontarelli, Fabrizio; Dharap, Ashutosh

    2018-06-03

    Gene expression in cerebral ischemia has been a subject of intense investigations for several years. Studies utilizing probe-based high-throughput methodologies such as microarrays have contributed significantly to our existing knowledge but lacked the capacity to dissect the transcriptome in detail. Genome-wide RNA-sequencing (RNA-seq) enables comprehensive examinations of transcriptomes for attributes such as strandedness, alternative splicing, alternative transcription start/stop sites, and sequence composition, thus providing a very detailed account of gene expression. Leveraging this capability, we conducted an in-depth, genome-wide evaluation of the protein-coding transcriptome of the adult mouse cortex after transient focal ischemia at 6, 12, or 24 h of reperfusion using RNA-seq. We identified a total of 1007 transcripts at 6 h, 1878 transcripts at 12 h, and 1618 transcripts at 24 h of reperfusion that were significantly altered as compared to sham controls. With isoform-level resolution, we identified 23 splice variants arising from 23 genes that were novel mRNA isoforms. For a subset of genes, we detected reperfusion time-point-dependent splice isoform switching, indicating an expression and/or functional switch for these genes. Finally, for 286 genes across all three reperfusion time-points, we discovered multiple, distinct, simultaneously expressed and differentially altered isoforms per gene that were generated via alternative transcription start/stop sites. Of these, 165 isoforms derived from 109 genes were novel mRNAs. Together, our data unravel the protein-coding transcriptome of the cerebral cortex at an unprecedented depth to provide several new insights into the flexibility and complexity of stroke-related gene transcription and transcript organization.

  18. Isoform-specific proteasomal degradation of Rbfox3 during chicken embryonic development

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Kee K.; Adelstein, Robert S.; Kawamoto, Sachiyo, E-mail: kawamots@mail.nih.gov

    2014-08-08

    Highlights: • Protein stability of Rbfox3 splice isoforms is differentially regulated. • Rbfox3-d31, an Rbfox3 isoform lacking the RRM, is highly susceptible to degradation. • The protein stability of Rbfox3-d31 is regulated by the ubiquitin–proteasome pathway. • Rbfox3-d31 inhibits the nuclear localization of Rbfox2. • Rbfox3-d31 inhibits the splicing activity of Rbfox2. - Abstract: Rbfox3, a neuron-specific RNA-binding protein, plays an important role in neuronal differentiation during development. An isoform Rbfox3-d31, which excludes the 93-nucleotide cassette exon within the RNA recognition motif of chicken Rbfox3, has been previously identified. However, the cellular functions of Rbfox3-d31 remain largely unknown. Here we find that Rbfox3-d31 mRNA is highly expressed during the early developmental stages of the chicken embryo, while Rbfox3-d31 protein is barely detected during the same stage due to its rapid degradation mediated by the ubiquitin–proteasome pathway. Importantly, this degradation is specific to the Rbfox3-d31 isoform and it does not occur with full-length Rbfox3. Furthermore, suppression of Rbfox3-d31 protein degradation with the proteasome inhibitor MG132 attenuates the splicing activity of another Rbfox family member Rbfox2 by altering the subcellular localization of Rbfox2. These results suggest that Rbfox3-d31 functions as a repressor for the splicing activity of the Rbfox family and its protein level is regulated in an isoform-specific manner in vivo.

  19. Myosin isoform switching during assembly of the Drosophila flight muscle thick filament lattice.

    Science.gov (United States)

    Orfanos, Zacharias; Sparrow, John C

    2013-01-01

    During muscle development myosin molecules form symmetrical thick filaments, which integrate with the thin filaments to produce the regular sarcomeric lattice. In Drosophila indirect flight muscles (IFMs) the details of this process can be studied using genetic approaches. The weeP26 transgenic line has a GFP-encoding exon inserted into the single Drosophila muscle myosin heavy chain gene, Mhc. The weeP26 IFM sarcomeres have a unique MHC-GFP-labelling pattern restricted to the sarcomere core, explained by non-translation of the GFP exon following alternative splicing. Characterisation of wild-type IFM MHC mRNA confirmed the presence of an alternately spliced isoform, expressed earlier than the major IFM-specific isoform. The two wild-type IFM-specific MHC isoforms differ by the presence of a C-terminal 'tailpiece' in the minor isoform. The sequential expression and assembly of these two MHCs into developing thick filaments suggest a role for the tailpiece in initiating A-band formation. The restriction of the MHC-GFP sarcomeric pattern in weeP26 is lifted when the IFM lack the IFM-specific myosin binding protein flightin, suggesting that it limits myosin dissociation from thick filaments. Studies of flightin binding to developing thick filaments reveal a progressive binding at the growing thick filament tips and in a retrograde direction to earlier assembled, proximal filament regions. We propose that this flightin binding restricts myosin molecule incorporation/dissociation during thick filament assembly and explains the location of the early MHC isoform pattern in the IFM A-band.

  20. Characterization of p38 MAPK isoforms for drug resistance study using systems biology approach.

    Science.gov (United States)

    Peng, Huiming; Peng, Tao; Wen, Jianguo; Engler, David A; Matsunami, Risë K; Su, Jing; Zhang, Le; Chang, Chung-Che Jeff; Zhou, Xiaobo

    2014-07-01

    p38 mitogen-activated protein kinase activation plays an important role in resistance to chemotherapeutic cytotoxic drugs in treating multiple myeloma (MM). However, how the p38 mitogen-activated protein kinase signaling pathway is involved in drug resistance, in particular the roles that the various p38 isoforms play, remains largely unknown. To explore the underlying mechanisms, we developed a novel systems biology approach by integrating liquid chromatography-mass spectrometry and reverse phase protein array data from human MM cell lines with computational pathway models in which the unknown parameters were inferred using a proposed novel algorithm called modularized factor graph. New mechanisms predicted by our models suggest that combined activation of various p38 isoforms may result in drug resistance in MM via regulating the related pathways including extracellular signal-regulated kinase (ERK) pathway and NFкB pathway. ERK pathway regulating cell growth is synergistically regulated by p38δ isoform, whereas nuclear factor kappa B (NFкB) pathway regulating cell apoptosis is synergistically regulated by p38α isoform. This finding that p38δ isoform promotes the phosphorylation of ERK1/2 in MM cells treated with bortezomib was validated by western blotting. Based on the predicted mechanisms, we further screened drug combinations in silico and found that a promising drug combination targeting ERK1/2 and NFκB might reduce the effects of drug resistance in MM cells. This study provides a framework of a systems biology approach to studying drug resistance and drug combination selection. RPPA experimental Data and Matlab source codes of modularized factor graph for parameter estimation are freely available online at http://ctsb.is.wfubmc.edu/publications/modularized-factor-graph.php. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  1. The Impact of Endurance Training on Human Skeletal Muscle Memory, Global Isoform Expression and Novel Transcripts.

    Directory of Open Access Journals (Sweden)

    Maléne E Lindholm

    2016-09-01

    Full Text Available Regularly performed endurance training has many beneficial effects on health and skeletal muscle function, and can be used to prevent and treat common diseases e.g. cardiovascular disease, type II diabetes and obesity. The molecular adaptation mechanisms regulating these effects are incompletely understood. To date, global transcriptome changes in skeletal muscles have been studied at the gene level only. Therefore, global isoform expression changes following exercise training in humans are unknown. Also, the effects of repeated interventions on transcriptional memory or training response have not been studied before. In this study, 23 individuals trained one leg for three months. Nine months later, 12 of the same subjects trained both legs in a second training period. Skeletal muscle biopsies were obtained from both legs before and after both training periods. RNA sequencing analysis of all 119 skeletal muscle biopsies showed that training altered the expression of 3,404 gene isoforms, mainly associated with oxidative ATP production. Fifty-four genes had isoforms that changed in opposite directions. Training altered expression of 34 novel transcripts, all with protein-coding potential. After nine months of detraining, no training-induced transcriptome differences were detected between the previously trained and untrained legs. Although there were several differences in the physiological and transcriptional responses to repeated training, no coherent evidence of an endurance training induced transcriptional skeletal muscle memory was found. This human lifestyle intervention induced differential expression of thousands of isoforms and several transcripts from unannotated regions of the genome. It is likely that the observed isoform expression changes reflect adaptational mechanisms and processes that provide the functional and health benefits of regular physical activity.

  2. Comparison of five DNA quantification methods

    DEFF Research Database (Denmark)

    Nielsen, Karsten; Mogensen, Helle Smidt; Hedman, Johannes

    2008-01-01

    Six commercial preparations of human genomic DNA were quantified using five quantification methods: UV spectrometry, SYBR-Green dye staining, slot blot hybridization with the probe D17Z1, Quantifiler Human DNA Quantification kit and RB1 rt-PCR. All methods measured higher DNA concentrations than...... Quantification kit in two experiments. The measured DNA concentrations with Quantifiler were 125 and 160% higher than expected based on the manufacturers' information. When the Quantifiler human DNA standard (Raji cell line) was replaced by the commercial human DNA preparation G147A (Promega) to generate the DNA...... standard curve in the Quantifiler Human DNA Quantification kit, the DNA quantification results of the human DNA preparations were 31% higher than expected based on the manufacturers' information. The results indicate a calibration problem with the Quantifiler human DNA standard for its use...

  3. DMPD: The role of C/EBP isoforms in the control of inflammatory and native immunityfunctions. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 9792624 The role of C/EBP isoforms in the control of inflammatory and native immunityfunction...f C/EBP isoforms in the control of inflammatory and native immunityfunctions. PubmedID 9792624 Title The rol...e of C/EBP isoforms in the control of inflammatory and native immunityfunctions.

  4. Crystallization and Identification of the Glycosylated Moieties of Two Isoforms of the Main Allergen Hev b 2 and Preliminary X-ray Analysis of Two Polymorphs of Isoform ll

    Energy Technology Data Exchange (ETDEWEB)

    Fuentes-Silva,D.; Mendoza-Hernandez, G.; Stojanoff, V.; Palomares, L.; Zenteno, E.; Torres-Larios, A.; Rodriguez-Romero, A.

    2007-01-01

    Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a {beta}-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content consisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapor-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4{sub 1} with unit-cell parameters a = b = 150.17, c = 77.41 {angstrom} were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 {angstrom}, {beta}= 113.6{sup o}. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

  5. Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II

    Energy Technology Data Exchange (ETDEWEB)

    Fuentes-Silva, D. [Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Mendoza-Hernández, G. [Facultad de Medicina, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Stojanoff, V. [Brookhaven National Laboratory, National Synchrotron Light Source, Upton, NY (United States); Palomares, L. A. [Instituto de Biotecnología, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Zenteno, E. [Facultad de Medicina, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Torres-Larios, A. [Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Rodríguez-Romero, A., E-mail: adela@servidor.unam.mx [Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico)

    2007-09-01

    Crystallization of important glycoenzymes involved in IgE-mediated latex allergy. Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a β-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content constisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4{sub 1} with unit-cell parameters a = b = 150.17, c = 77.41 Å were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 Å, β = 113.6°. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

  6. Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II

    International Nuclear Information System (INIS)

    Fuentes-Silva, D.; Mendoza-Hernández, G.; Stojanoff, V.; Palomares, L. A.; Zenteno, E.; Torres-Larios, A.; Rodríguez-Romero, A.

    2007-01-01

    Crystallization of important glycoenzymes involved in IgE-mediated latex allergy. Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a β-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content constisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4 1 with unit-cell parameters a = b = 150.17, c = 77.41 Å were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2 1 , with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 Å, β = 113.6°. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units

  7. Safety and reliability criteria

    International Nuclear Information System (INIS)

    O'Neil, R.

    1978-01-01

    Nuclear power plants and, in particular, reactor pressure boundary components have unique reliability requirements, in that usually no significant redundancy is possible, and a single failure can give rise to possible widespread core damage and fission product release. Reliability may be required for availability or safety reasons, but in the case of the pressure boundary and certain other systems safety may dominate. Possible Safety and Reliability (S and R) criteria are proposed which would produce acceptable reactor design. Without some S and R requirement the designer has no way of knowing how far he must go in analysing his system or component, or whether his proposed solution is likely to gain acceptance. The paper shows how reliability targets for given components and systems can be individually considered against the derived S and R criteria at the design and construction stage. Since in the case of nuclear pressure boundary components there is often very little direct experience on which to base reliability studies, relevant non-nuclear experience is examined. (author)

  8. Proposed reliability cost model

    Science.gov (United States)

    Delionback, L. M.

    1973-01-01

    The research investigations which were involved in the study include: cost analysis/allocation, reliability and product assurance, forecasting methodology, systems analysis, and model-building. This is a classic example of an interdisciplinary problem, since the model-building requirements include the need for understanding and communication between technical disciplines on one hand, and the financial/accounting skill categories on the other. The systems approach is utilized within this context to establish a clearer and more objective relationship between reliability assurance and the subcategories (or subelements) that provide, or reenforce, the reliability assurance for a system. Subcategories are further subdivided as illustrated by a tree diagram. The reliability assurance elements can be seen to be potential alternative strategies, or approaches, depending on the specific goals/objectives of the trade studies. The scope was limited to the establishment of a proposed reliability cost-model format. The model format/approach is dependent upon the use of a series of subsystem-oriented CER's and sometimes possible CTR's, in devising a suitable cost-effective policy.

  9. Monoclonal antibodies against muscle actin isoforms: epitope identification and analysis of isoform expression by immunoblot and immunostaining in normal and regenerating skeletal muscle [version 2; referees: 3 approved

    Directory of Open Access Journals (Sweden)

    Christine Chaponnier

    2016-06-01

    Full Text Available Higher vertebrates (mammals and birds express six different highly conserved actin isoforms that can be classified in three subgroups: 1 sarcomeric actins, α-skeletal (α-SKA and α-cardiac (α-CAA, 2 smooth muscle actins (SMAs, α-SMA and γ-SMA, and 3 cytoplasmic actins (CYAs, β-CYA and γ-CYA. The variations among isoactins, in each subgroup, are due to 3-4 amino acid differences located in their acetylated N-decapeptide sequence. The first monoclonal antibody (mAb against an actin isoform (α-SMA was produced and characterized in our laboratory in 1986 (Skalli  et al., 1986 . We have further obtained mAbs against the 5 other isoforms. In this report, we focus on the mAbs anti-α-SKA and anti-α-CAA obtained after immunization of mice with the respective acetylated N-terminal decapeptides using the Repetitive Immunizations at Multiple Sites Strategy (RIMMS. In addition to the identification of their epitope by immunoblotting, we describe the expression of the 2 sarcomeric actins in mature skeletal muscle and during muscle repair after micro-lesions. In particular, we analyze the expression of α-CAA, α-SKA and α-SMA by co-immunostaining in a time course frame during the muscle repair process. Our results indicate that a restricted myocyte population expresses α-CAA and suggest a high capacity of self-regeneration in muscle cells. These antibodies may represent a helpful tool for the follow-up of muscle regeneration and pathological changes.

  10. Reliability Centered Maintenance - Methodologies

    Science.gov (United States)

    Kammerer, Catherine C.

    2009-01-01

    Journal article about Reliability Centered Maintenance (RCM) methodologies used by United Space Alliance, LLC (USA) in support of the Space Shuttle Program at Kennedy Space Center. The USA Reliability Centered Maintenance program differs from traditional RCM programs because various methodologies are utilized to take advantage of their respective strengths for each application. Based on operational experience, USA has customized the traditional RCM methodology into a streamlined lean logic path and has implemented the use of statistical tools to drive the process. USA RCM has integrated many of the L6S tools into both RCM methodologies. The tools utilized in the Measure, Analyze, and Improve phases of a Lean Six Sigma project lend themselves to application in the RCM process. All USA RCM methodologies meet the requirements defined in SAE JA 1011, Evaluation Criteria for Reliability-Centered Maintenance (RCM) Processes. The proposed article explores these methodologies.

  11. Issues in cognitive reliability

    International Nuclear Information System (INIS)

    Woods, D.D.; Hitchler, M.J.; Rumancik, J.A.

    1984-01-01

    This chapter examines some problems in current methods to assess reactor operator reliability at cognitive tasks and discusses new approaches to solve these problems. The two types of human failures are errors in the execution of an intention and errors in the formation/selection of an intention. Topics considered include the types of description, error correction, cognitive performance and response time, the speed-accuracy tradeoff function, function based task analysis, and cognitive task analysis. One problem of human reliability analysis (HRA) techniques in general is the question of what are the units of behavior whose reliability are to be determined. A second problem for HRA is that people often detect and correct their errors. The use of function based analysis, which maps the problem space for plant control, is recommended

  12. A computational Bayesian approach to dependency assessment in system reliability

    International Nuclear Information System (INIS)

    Yontay, Petek; Pan, Rong

    2016-01-01

    Due to the increasing complexity of engineered products, it is of great importance to develop a tool to assess reliability dependencies among components and systems under the uncertainty of system reliability structure. In this paper, a Bayesian network approach is proposed for evaluating the conditional probability of failure within a complex system, using a multilevel system configuration. Coupling with Bayesian inference, the posterior distributions of these conditional probabilities can be estimated by combining failure information and expert opinions at both system and component levels. Three data scenarios are considered in this study, and they demonstrate that, with the quantification of the stochastic relationship of reliability within a system, the dependency structure in system reliability can be gradually revealed by the data collected at different system levels. - Highlights: • A Bayesian network representation of system reliability is presented. • Bayesian inference methods for assessing dependencies in system reliability are developed. • Complete and incomplete data scenarios are discussed. • The proposed approach is able to integrate reliability information from multiple sources at multiple levels of the system.

  13. Reliability issues in PACS

    Science.gov (United States)

    Taira, Ricky K.; Chan, Kelby K.; Stewart, Brent K.; Weinberg, Wolfram S.

    1991-07-01

    Reliability is an increasing concern when moving PACS from the experimental laboratory to the clinical environment. Any system downtime may seriously affect patient care. The authors report on the several classes of errors encountered during the pre-clinical release of the PACS during the past several months and present the solutions implemented to handle them. The reliability issues discussed include: (1) environmental precautions, (2) database backups, (3) monitor routines of critical resources and processes, (4) hardware redundancy (networks, archives), and (5) development of a PACS quality control program.

  14. Reliability Parts Derating Guidelines

    Science.gov (United States)

    1982-06-01

    226-30, October 1974. 66 I, 26. "Reliability of GAAS Injection Lasers", De Loach , B. C., Jr., 1973 IEEE/OSA Conference on Laser Engineering and...Vol. R-23, No. 4, 226-30, October 1974. 28. "Reliability of GAAS Injection Lasers", De Loach , B. C., Jr., 1973 IEEE/OSA Conference on Laser...opnatien ot 󈨊 deg C, mounted on a 4-inach square 0.250~ inch thick al~loy alum~nusi panel.. This mounting technique should be L~ ken into cunoidur~tiou

  15. Reverse transcriptase real-time PCR for detection and quantification of viable Campylobacter jejuni directly from poultry faecal samples

    DEFF Research Database (Denmark)

    Bui, Thanh Xuan; Wolff, Anders; Madsen, Mogens

    2012-01-01

    Campylobacter spp. is the most common cause of bacterial diarrhoea in humans worldwide. Therefore, rapid and reliable methods fordetection and quantification of this pathogen are required. In this study, we have developed a reverse transcription quantitative real-time PCR(RT-qPCR) for detection a...

  16. Improved quantification of alite and belite in anhydrous Portland cements by 29Si MAS NMR: Effects of paramagnetic ions

    DEFF Research Database (Denmark)

    Poulsen, Søren Lundsted; Kocaba, Vanessa; Le Saoût, Gwenn

    2009-01-01

    The applicability, reliability, and repeatability of 29Si MAS NMR for determination of the quantities of alite (Ca3SiO5) and belite (Ca2SiO4) in anhydrous Portland cement was investigated in detail for 11 commercial Portland cements and the results compared with phase quantifications based...

  17. Isoform-specific potentiation of stem and progenitor cell engraftment by AML1/RUNX1.

    Directory of Open Access Journals (Sweden)

    Shinobu Tsuzuki

    2007-05-01

    Full Text Available AML1/RUNX1 is the most frequently mutated gene in leukaemia and is central to the normal biology of hematopoietic stem and progenitor cells. However, the role of different AML1 isoforms within these primitive compartments is unclear. Here we investigate whether altering relative expression of AML1 isoforms impacts the balance between cell self-renewal and differentiation in vitro and in vivo.The human AML1a isoform encodes a truncated molecule with DNA-binding but no transactivation capacity. We used a retrovirus-based approach to transduce AML1a into primitive haematopoietic cells isolated from the mouse. We observed that enforced AML1a expression increased the competitive engraftment potential of murine long-term reconstituting stem cells with the proportion of AML1a-expressing cells increasing over time in both primary and secondary recipients. Furthermore, AML1a expression dramatically increased primitive and committed progenitor activity in engrafted animals as assessed by long-term culture, cobblestone formation, and colony assays. In contrast, expression of the full-length isoform AML1b abrogated engraftment potential. In vitro, AML1b promoted differentiation while AML1a promoted proliferation of progenitors capable of short-term lymphomyeloid engraftment. Consistent with these findings, the relative abundance of AML1a was highest in the primitive stem/progenitor compartment of human cord blood, and forced expression of AML1a in these cells enhanced maintenance of primitive potential both in vitro and in vivo.These data demonstrate that the "a" isoform of AML1 has the capacity to potentiate stem and progenitor cell engraftment, both of which are required for successful clinical transplantation. This activity is consistent with its expression pattern in both normal and leukaemic cells. Manipulating the balance of AML1 isoform expression may offer novel therapeutic strategies, exploitable in the contexts of leukaemia and also in cord blood

  18. Tumour cells expressing single VEGF isoforms display distinct growth, survival and migration characteristics.

    Directory of Open Access Journals (Sweden)

    Chryso Kanthou

    Full Text Available Vascular endothelial growth factor-A (VEGF is produced by most cancer cells as multiple isoforms, which display distinct biological activities. VEGF plays an undisputed role in tumour growth, vascularisation and metastasis; nevertheless the functions of individual isoforms in these processes remain poorly understood. We investigated the effects of three main murine isoforms (VEGF188, 164 and 120 on tumour cell behaviour, using a panel of fibrosarcoma cells we developed that express them individually under endogenous promoter control. Fibrosarcomas expressing only VEGF188 (fs188 or wild type controls (fswt were typically mesenchymal, formed ruffles and displayed strong matrix-binding activity. VEGF164- and VEGF120-producing cells (fs164 and fs120 respectively were less typically mesenchymal, lacked ruffles but formed abundant cell-cell contacts. On 3D collagen, fs188 cells remained mesenchymal while fs164 and fs120 cells adopted rounded/amoeboid and a mix of rounded and elongated morphologies respectively. Consistent with their mesenchymal characteristics, fs188 cells migrated significantly faster than fs164 or fs120 cells on 2D surfaces while contractility inhibitors accelerated fs164 and fs120 cell migration. VEGF164/VEGF120 expression correlated with faster proliferation rates and lower levels of spontaneous apoptosis than VEGF188 expression. Nevertheless, VEGF188 was associated with constitutively active/phosphorylated AKT, ERK1/2 and Stat3 proteins. Differences in proliferation rates and apoptosis could be explained by defective signalling downstream of pAKT to FOXO and GSK3 in fs188 and fswt cells, which also correlated with p27/p21 cyclin-dependent kinase inhibitor over-expression. All cells expressed tyrosine kinase VEGF receptors, but these were not active/activatable suggesting that inherent differences between the cell lines are governed by endogenous VEGF isoform expression through complex interactions that are independent of tyrosine

  19. Conditional expression of CD44 isoforms in lymphoma cells: influence on hyaluronate binding and tumor growth

    Energy Technology Data Exchange (ETDEWEB)

    Fu, J.

    2002-03-01

    CD44 describes a family of surface proteins consisting of many isoforms due to alternative splice of ten 'variant' exons. Members of this family are involved in various processes including hematopoiesis, lymphocyte activation and homing, limb development, wound healing and tumor progression. Clinically, CD44 has been shown to be a prognostic factor for several human cancers. To answer the question which isoform might be relevant for tumor progression and to gain an insight into the mechanism of its function, I established transfectants of the LB lymphoma cell line in which the expression of four CD44 isoforms, namely CD44v3-10, CD44v4-10, CD44v8-10 and CD44s, was controlled by the Tet-off promoter. In the presence of Doxycycline, the expression was repressed. Removal of Doxycycline switched on expression and the maximal CD44 amount was obtained within two days. The transfectants were characterized regarding their ability to bind to the extracellular matrix component hyaluronate (HA). Overexpression of all four CD44 isoforms conferred the ability to bind HA on LB cells. Other glycosaminoglycans (GAGs) were bound in an isotype-specific fashion. CD44v3-10, CD44v4-10 and CD44v8-10 showed high binding affinity to chondroitin A, B and C, and low affinity to heparin, heparan sulfate and keratan sulfate. CD44s could not bind to these GAGs. Among these three variants, the binding ability of CD44v3-10 was the strongest. CD44 clustering seemed to play a crucial role for HA binding. Both CD44s and CD44v8-10 formed reduction-sensitive complexes in LB cells. The complexes are homooligomers or heterooligomers composed of different isoforms. Cys286 in CD44 transmember domain was not responsible for the formation of reduction-sensitive oligomer or for the enhanced HA binding in LB cell line. Using a conditional dimerization system the requirement of CD44 oligomerization for HA binding was directly demonstrated. The induction of oligomerization increased HA binding

  20. Conditional expression of CD44 isoforms in lymphoma cells: influence on hyaluronate binding and tumor growth

    International Nuclear Information System (INIS)

    Fu, J.

    2002-03-01

    CD44 describes a family of surface proteins consisting of many isoforms due to alternative splice of ten 'variant' exons. Members of this family are involved in various processes including hematopoiesis, lymphocyte activation and homing, limb development, wound healing and tumor progression. Clinically, CD44 has been shown to be a prognostic factor for several human cancers. To answer the question which isoform might be relevant for tumor progression and to gain an insight into the mechanism of its function, I established transfectants of the LB lymphoma cell line in which the expression of four CD44 isoforms, namely CD44v3-10, CD44v4-10, CD44v8-10 and CD44s, was controlled by the Tet-off promoter. In the presence of Doxycycline, the expression was repressed. Removal of Doxycycline switched on expression and the maximal CD44 amount was obtained within two days. The transfectants were characterized regarding their ability to bind to the extracellular matrix component hyaluronate (HA). Overexpression of all four CD44 isoforms conferred the ability to bind HA on LB cells. Other glycosaminoglycans (GAGs) were bound in an isotype-specific fashion. CD44v3-10, CD44v4-10 and CD44v8-10 showed high binding affinity to chondroitin A, B and C, and low affinity to heparin, heparan sulfate and keratan sulfate. CD44s could not bind to these GAGs. Among these three variants, the binding ability of CD44v3-10 was the strongest. CD44 clustering seemed to play a crucial role for HA binding. Both CD44s and CD44v8-10 formed reduction-sensitive complexes in LB cells. The complexes are homooligomers or heterooligomers composed of different isoforms. Cys286 in CD44 transmember domain was not responsible for the formation of reduction-sensitive oligomer or for the enhanced HA binding in LB cell line. Using a conditional dimerization system the requirement of CD44 oligomerization for HA binding was directly demonstrated. The induction of oligomerization increased HA binding. Finally, I

  1. Development of Quantification Method for Bioluminescence Imaging

    International Nuclear Information System (INIS)

    Kim, Hyeon Sik; Min, Jung Joon; Lee, Byeong Il; Choi, Eun Seo; Tak, Yoon O; Choi, Heung Kook; Lee, Ju Young

    2009-01-01

    Optical molecular luminescence imaging is widely used for detection and imaging of bio-photons emitted by luminescent luciferase activation. The measured photons in this method provide the degree of molecular alteration or cell numbers with the advantage of high signal-to-noise ratio. To extract useful information from the measured results, the analysis based on a proper quantification method is necessary. In this research, we propose a quantification method presenting linear response of measured light signal to measurement time. We detected the luminescence signal by using lab-made optical imaging equipment of animal light imaging system (ALIS) and different two kinds of light sources. One is three bacterial light-emitting sources containing different number of bacteria. The other is three different non-bacterial light sources emitting very weak light. By using the concept of the candela and the flux, we could derive simplified linear quantification formula. After experimentally measuring light intensity, the data was processed with the proposed quantification function. We could obtain linear response of photon counts to measurement time by applying the pre-determined quantification function. The ratio of the re-calculated photon counts and measurement time present a constant value although different light source was applied. The quantification function for linear response could be applicable to the standard quantification process. The proposed method could be used for the exact quantitative analysis in various light imaging equipment with presenting linear response behavior of constant light emitting sources to measurement time

  2. Advancing agricultural greenhouse gas quantification*

    Science.gov (United States)

    Olander, Lydia; Wollenberg, Eva; Tubiello, Francesco; Herold, Martin

    2013-03-01

    1. Introduction Better information on greenhouse gas (GHG) emissions and mitigation potential in the agricultural sector is necessary to manage these emissions and identify responses that are consistent with the food security and economic development priorities of countries. Critical activity data (what crops or livestock are managed in what way) are poor or lacking for many agricultural systems, especially in developing countries. In addition, the currently available methods for quantifying emissions and mitigation are often too expensive or complex or not sufficiently user friendly for widespread use. The purpose of this focus issue is to capture the state of the art in quantifying greenhouse gases from agricultural systems, with the goal of better understanding our current capabilities and near-term potential for improvement, with particular attention to quantification issues relevant to smallholders in developing countries. This work is timely in light of international discussions and negotiations around how agriculture should be included in efforts to reduce and adapt to climate change impacts, and considering that significant climate financing to developing countries in post-2012 agreements may be linked to their increased ability to identify and report GHG emissions (Murphy et al 2010, CCAFS 2011, FAO 2011). 2. Agriculture and climate change mitigation The main agricultural GHGs—methane and nitrous oxide—account for 10%-12% of anthropogenic emissions globally (Smith et al 2008), or around 50% and 60% of total anthropogenic methane and nitrous oxide emissions, respectively, in 2005. Net carbon dioxide fluxes between agricultural land and the atmosphere linked to food production are relatively small, although significant carbon emissions are associated with degradation of organic soils for plantations in tropical regions (Smith et al 2007, FAO 2012). Population growth and shifts in dietary patterns toward more meat and dairy consumption will lead to

  3. Brain region-specific expression of MeCP2 isoforms correlates with DNA methylation within Mecp2 regulatory elements.

    Directory of Open Access Journals (Sweden)

    Carl O Olson

    Full Text Available MeCP2 is a critical epigenetic regulator in brain and its abnormal expression or compromised function leads to a spectrum of neurological disorders including Rett Syndrome and autism. Altered expression of the two MeCP2 isoforms, MeCP2E1 and MeCP2E2 has been implicated in neurological complications. However, expression, regulation and functions of the two isoforms are largely uncharacterized. Previously, we showed the role of MeCP2E1 in neuronal maturation and reported MeCP2E1 as the major protein isoform in the adult mouse brain, embryonic neurons and astrocytes. Recently, we showed that DNA methylation at the regulatory elements (REs within the Mecp2 promoter and intron 1 impact the expression of Mecp2 isoforms in differentiating neural stem cells. This current study is aimed for a comparative analysis of temporal, regional and cell type-specific expression of MeCP2 isoforms in the developing and adult mouse brain. MeCP2E2 displayed a later expression onset than MeCP2E1 during mouse brain development. In the adult female and male brain hippocampus, both MeCP2 isoforms were detected in neurons, astrocytes and oligodendrocytes. Furthermore, MeCP2E1 expression was relatively uniform in different brain regions (olfactory bulb, striatum, cortex, hippocampus, thalamus, brainstem and cerebellum, whereas MeCP2E2 showed differential enrichment in these brain regions. Both MeCP2 isoforms showed relatively similar distribution in these brain regions, except for cerebellum. Lastly, a preferential correlation was observed between DNA methylation at specific CpG dinucleotides within the REs and Mecp2 isoform-specific expression in these brain regions. Taken together, we show that MeCP2 isoforms display differential expression patterns during brain development and in adult mouse brain regions. DNA methylation patterns at the Mecp2 REs may impact this differential expression of Mecp2/MeCP2 isoforms in brain regions. Our results significantly contribute

  4. SSP: an interval integer linear programming for de novo transcriptome assembly and isoform discovery of RNA-seq reads.

    Science.gov (United States)

    Safikhani, Zhaleh; Sadeghi, Mehdi; Pezeshk, Hamid; Eslahchi, Changiz

    2013-01-01

    Recent advances in the sequencing technologies have provided a handful of RNA-seq datasets for transcriptome analysis. However, reconstruction of full-length isoforms and estimation of the expression level of transcripts with a low cost are challenging tasks. We propose a novel de novo method named SSP that incorporates interval integer linear programming to resolve alternatively spliced isoforms and reconstruct the whole transcriptome from short reads. Experimental results show that SSP is fast and precise in determining different alternatively spliced isoforms along with the estimation of reconstructed transcript abundances. The SSP software package is available at http://www.bioinf.cs.ipm.ir/software/ssp. © 2013.

  5. Uncertainty quantification and error analysis

    Energy Technology Data Exchange (ETDEWEB)

    Higdon, Dave M [Los Alamos National Laboratory; Anderson, Mark C [Los Alamos National Laboratory; Habib, Salman [Los Alamos National Laboratory; Klein, Richard [Los Alamos National Laboratory; Berliner, Mark [OHIO STATE UNIV.; Covey, Curt [LLNL; Ghattas, Omar [UNIV OF TEXAS; Graziani, Carlo [UNIV OF CHICAGO; Seager, Mark [LLNL; Sefcik, Joseph [LLNL; Stark, Philip [UC/BERKELEY; Stewart, James [SNL

    2010-01-01

    UQ studies all sources of error and uncertainty, including: systematic and stochastic measurement error; ignorance; limitations of theoretical models; limitations of numerical representations of those models; limitations on the accuracy and reliability of computations, approximations, and algorithms; and human error. A more precise definition for UQ is suggested below.

  6. A performance study on three qPCR quantification kits and their compatibilities with the 6-dye DNA profiling systems.

    Science.gov (United States)

    Lin, Sze-Wah; Li, Christina; Ip, Stephen C Y

    2018-03-01

    DNA quantification plays an integral role in forensic DNA profiling. Not only does it estimate the total amount of amplifiable human autosomal and male DNA to ensure optimal amplification of target DNA for subsequent analysis, but also assesses the extraction efficiency and purity of the DNA extract. Latest DNA quantification systems even offer an estimate for the degree of DNA degradation in a sample. Here, we report the performance of three new generation qPCR kits, namely Investigator ® Quantiplex HYres Kit from QIAGEN, Quantifiler ® Trio DNA Quantification Kit from Applied Biosystems™, and PowerQuant ® System from Promega, and their compatibilities with three 6-dye DNA profiling systems. Our results have demonstrated that all three kits generate standard curves with satisfactory consistency and reproducibility, and are capable of screening out traces of male DNA in the presence of 30-fold excess of female DNA. They also exhibit a higher tolerance to PCR inhibition than Quantifiler ® Human DNA Quantification Kit from Applied Biosystems™ in autosomal DNA quantification. PowerQuant ® , as compared to Quantiplex HYres and Quantifiler ® Trio, shows a better precision for both autosomal and male DNA quantifications. Quantifiler ® Trio and PowerQuant ® in contrast to Quantiplex HYres offer better correlations with lower discrepancies between autosomal and male DNA quantification, and their additional degradation index features provide a detection platform for inhibited and/or degraded DNA template. Regarding the compatibility between these quantification and profiling systems: (1) both Quantifiler ® Trio and PowerQuant ® work well with GlobalFiler and Fusion 6C, allowing a fairly accurate prediction of their DNA typing results based on the quantification values; (2) Quantiplex HYres offers a fairly reliable IPC system for detecting any potential inhibitions on Investigator 24plex, whereas Quantifiler ® Trio and PowerQuant ® suit better for Global

  7. Sample Size Bounding and Context Ranking as Approaches to the Human Error Quantification Problem

    Energy Technology Data Exchange (ETDEWEB)

    Reer, B

    2004-03-01

    The paper describes a technique denoted as Sub-Sample-Size Bounding (SSSB), which is useable for the statistical derivation of context-specific probabilities from data available in existing reports on operating experience. Applications to human reliability analysis (HRA) are emphasised in the presentation of this technique. Exemplified by a sample of 180 abnormal event sequences, the manner in which SSSB can provide viable input for the quantification of errors of commission (EOCs) are outlined. (author)

  8. Working group of experts on rare events in human error analysis and quantification

    International Nuclear Information System (INIS)

    Goodstein, L.P.

    1977-01-01

    In dealing with the reference problem of rare events in nuclear power plants, the group has concerned itself with the man-machine system and, in particular, with human error analysis and quantification. The Group was requested to review methods of human reliability prediction, to evaluate the extent to which such analyses can be formalized and to establish criteria to be met by task conditions and system design which would permit a systematic, formal analysis. Recommendations are given on the Fessenheim safety system

  9. Sample Size Bounding and Context Ranking as Approaches to the Human Error Quantification Problem

    International Nuclear Information System (INIS)

    Reer, B.

    2004-01-01

    The paper describes a technique denoted as Sub-Sample-Size Bounding (SSSB), which is useable for the statistical derivation of context-specific probabilities from data available in existing reports on operating experience. Applications to human reliability analysis (HRA) are emphasised in the presentation of this technique. Exemplified by a sample of 180 abnormal event sequences, the manner in which SSSB can provide viable input for the quantification of errors of commission (EOCs) are outlined. (author)

  10. Columbus safety and reliability

    Science.gov (United States)

    Longhurst, F.; Wessels, H.

    1988-10-01

    Analyses carried out to ensure Columbus reliability, availability, and maintainability, and operational and design safety are summarized. Failure modes/effects/criticality is the main qualitative tool used. The main aspects studied are fault tolerance, hazard consequence control, risk minimization, human error effects, restorability, and safe-life design.

  11. Reliability versus reproducibility

    International Nuclear Information System (INIS)

    Lautzenheiser, C.E.

    1976-01-01

    Defect detection and reproducibility of results are two separate but closely related subjects. It is axiomatic that a defect must be detected from examination to examination or reproducibility of results is very poor. On the other hand, a defect can be detected on each of subsequent examinations for higher reliability and still have poor reproducibility of results

  12. Power transformer reliability modelling

    NARCIS (Netherlands)

    Schijndel, van A.

    2010-01-01

    Problem description Electrical power grids serve to transport and distribute electrical power with high reliability and availability at acceptable costs and risks. These grids play a crucial though preferably invisible role in supplying sufficient power in a convenient form. Today’s society has

  13. Designing reliability into accelerators

    International Nuclear Information System (INIS)

    Hutton, A.

    1992-08-01

    For the next generation of high performance, high average luminosity colliders, the ''factories,'' reliability engineering must be introduced right at the inception of the project and maintained as a central theme throughout the project. There are several aspects which will be addressed separately: Concept; design; motivation; management techniques; and fault diagnosis

  14. Proof tests on reliability

    International Nuclear Information System (INIS)

    Mishima, Yoshitsugu

    1983-01-01

    In order to obtain public understanding on nuclear power plants, tests should be carried out to prove the reliability and safety of present LWR plants. For example, the aseismicity of nuclear power plants must be verified by using a large scale earthquake simulator. Reliability test began in fiscal 1975, and the proof tests on steam generators and on PWR support and flexure pins against stress corrosion cracking have already been completed, and the results have been internationally highly appreciated. The capacity factor of the nuclear power plant operation in Japan rose to 80% in the summer of 1983, and considering the period of regular inspection, it means the operation of almost full capacity. Japanese LWR technology has now risen to the top place in the world after having overcome the defects. The significance of the reliability test is to secure the functioning till the age limit is reached, to confirm the correct forecast of deteriorating process, to confirm the effectiveness of the remedy to defects and to confirm the accuracy of predicting the behavior of facilities. The reliability of nuclear valves, fuel assemblies, the heat affected zones in welding, reactor cooling pumps and electric instruments has been tested or is being tested. (Kako, I.)

  15. Reliability and code level

    NARCIS (Netherlands)

    Kasperski, M.; Geurts, C.P.W.

    2005-01-01

    The paper describes the work of the IAWE Working Group WBG - Reliability and Code Level, one of the International Codification Working Groups set up at ICWE10 in Copenhagen. The following topics are covered: sources of uncertainties in the design wind load, appropriate design target values for the

  16. Reliability of Plastic Slabs

    DEFF Research Database (Denmark)

    Thoft-Christensen, Palle

    1989-01-01

    In the paper it is shown how upper and lower bounds for the reliability of plastic slabs can be determined. For the fundamental case it is shown that optimal bounds of a deterministic and a stochastic analysis are obtained on the basis of the same failure mechanisms and the same stress fields....

  17. Reliability based structural design

    NARCIS (Netherlands)

    Vrouwenvelder, A.C.W.M.

    2014-01-01

    According to ISO 2394, structures shall be designed, constructed and maintained in such a way that they are suited for their use during the design working life in an economic way. To fulfil this requirement one needs insight into the risk and reliability under expected and non-expected actions. A

  18. Travel time reliability modeling.

    Science.gov (United States)

    2011-07-01

    This report includes three papers as follows: : 1. Guo F., Rakha H., and Park S. (2010), "A Multi-state Travel Time Reliability Model," : Transportation Research Record: Journal of the Transportation Research Board, n 2188, : pp. 46-54. : 2. Park S.,...

  19. Reliability and Model Fit

    Science.gov (United States)

    Stanley, Leanne M.; Edwards, Michael C.

    2016-01-01

    The purpose of this article is to highlight the distinction between the reliability of test scores and the fit of psychometric measurement models, reminding readers why it is important to consider both when evaluating whether test scores are valid for a proposed interpretation and/or use. It is often the case that an investigator judges both the…

  20. Parametric Mass Reliability Study

    Science.gov (United States)

    Holt, James P.

    2014-01-01

    The International Space Station (ISS) systems are designed based upon having redundant systems with replaceable orbital replacement units (ORUs). These ORUs are designed to be swapped out fairly quickly, but some are very large, and some are made up of many components. When an ORU fails, it is replaced on orbit with a spare; the failed unit is sometimes returned to Earth to be serviced and re-launched. Such a system is not feasible for a 500+ day long-duration mission beyond low Earth orbit. The components that make up these ORUs have mixed reliabilities. Components that make up the most mass-such as computer housings, pump casings, and the silicon board of PCBs-typically are the most reliable. Meanwhile components that tend to fail the earliest-such as seals or gaskets-typically have a small mass. To better understand the problem, my project is to create a parametric model that relates both the mass of ORUs to reliability, as well as the mass of ORU subcomponents to reliability.

  1. FRANX. Application for analysis and quantification of the APS fire; FRANK. Aplicacion para el analisis y cuantificacion de los APS de incendios

    Energy Technology Data Exchange (ETDEWEB)

    Snchez, A.; Osorio, F.; Ontoso, N.

    2014-07-01

    The FRANX application has been developed by EPRI within the Risk and Reliability User Group in order to facilitate the process of quantification and updating APS Fire (also covers floods and earthquakes). By applying fire scenarios are quantified in the central integrating the tasks performed during the APS fire. This paper describes the main features of the program to allow quantification of an APS Fire. (Author)

  2. Reliability Approach of a Compressor System using Reliability Block ...

    African Journals Online (AJOL)

    pc

    2018-03-05

    Mar 5, 2018 ... This paper presents a reliability analysis of such a system using reliability ... Keywords-compressor system, reliability, reliability block diagram, RBD .... the same structure has been kept with the three subsystems: air flow, oil flow and .... and Safety in Engineering Design", Springer, 2009. [3] P. O'Connor ...

  3. Deep Proteomics of Mouse Skeletal Muscle Enables Quantitation of Protein Isoforms, Metabolic Pathways, and Transcription Factors*

    Science.gov (United States)

    Deshmukh, Atul S.; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T.; Cox, Jürgen; Mann, Matthias

    2015-01-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms. PMID:25616865

  4. Examination of Gelatinase Isoforms in Rodent Models of Acute Neurodegenerative Diseases Using Two-Dimensional Zymography.

    Science.gov (United States)

    Chen, Shanyan; Meng, Fanjun; Chen, Zhenzhou; Qu, Zhe; Cui, Jiankun; Gu, Zezong

    2017-01-01

    Pathological activation of gelatinases (matrix metalloproteinase-2 and -9; MMP-2/-9) has been shown to cause a number of detrimental outcomes in neurodegenerative diseases. In gel gelatin zymography is a highly sensitive methodology commonly used in revealing levels of gelatinase activity and in separating the proform and active form of gelatinases, based on their different molecular weights. However, this methodology is inadequate in resolving complex enzyme isoforms, because gelatinase expression and activity can be regulated at transcriptional and/or post-translational levels under in vivo conditions resulting in alternation of their isoelectric focusing (IEF) points. In this chapter, we describe an advanced methodology, termed two-dimensional zymography, combining IEF with zymographic electrophoresis under non-reducing conditions to achieve significant improvement in separation of the gelatinase isoforms in both cell-based and in vivo models for acute brain injuries and neuroinflammation.

  5. Algal Toxin Azaspiracid-1 Induces Early Neuronal Differentiation and Alters Peripherin Isoform Stoichiometry

    Directory of Open Access Journals (Sweden)

    Linda V. Hjørnevik

    2015-12-01

    Full Text Available Azaspiracid-1 is an algal toxin that accumulates in edible mussels, and ingestion may result in human illness as manifested by vomiting and diarrhoea. When injected into mice, it causes neurotoxicological symptoms and death. Although it is well known that azaspiracid-1 is toxic to most cells and cell lines, little is known about its biological target(s. A rat PC12 cell line, commonly used as a model for the peripheral nervous system, was used to study the neurotoxicological effects of azaspiracid-1. Azaspiracid-1 induced differentiation-related morphological changes followed by a latter cell death. The differentiated phenotype showed peripherin-labelled neurite-like processes simultaneously as a specific isoform of peripherin was down-regulated. The precise mechanism behind this down-regulation remains uncertain. However, this study provides new insights into the neurological effects of azaspiracid-1 and into the biological significance of specific isoforms of peripherin.

  6. Hemoglobin isoform differentiation and allosteric regulation of oxygen binding in the turtle, Trachemys scripta

    DEFF Research Database (Denmark)

    Damsgaard, Christian; Storz, Jay F.; Hoffmann, Federico G.

    2013-01-01

    When freshwater turtles acclimatize to winter hibernation, there is a gradual transition from aerobic to anaerobic metabolism, which may require adjustments of blood O2 transport before turtles become anoxic. Here, we report the effects of protons, anionic cofactors, and temperature on the O2......-binding properties of isolated hemoglobin (Hb) isoforms, HbA and HbD, in the turtle Trachemys scripta. We determined the primary structures of the constituent subunits of the two Hb isoforms, and we related the measured functional properties to differences in O2 affinity between untreated hemolysates from...... turtles that were acclimated to normoxia and anoxia. Our data show that HbD has a consistently higher O2 affinity compared with HbA, whereas Bohr and temperature effects, as well as thiol reactivity, are similar. Although sequence data show amino acid substitutions at two known β-chain ATP-binding site...

  7. alpha isoforms of soluble and membrane-linked folate-binding protein in human blood

    DEFF Research Database (Denmark)

    Hoier-Madsen, M.; Holm, J.; Hansen, S.I.

    2008-01-01

    supported the hypothesis that serum FBP (29 kDa) mainly originates from neutrophils. The presence of FBP/FR alpha isoforms were established for the first time in human blood using antibodies specifically directed against human milk FBP alpha. The alpha isoforms identified on erythrocyte membranes......, and in granulocytes and serum, only constituted an almost undetectable fraction of the functional FBP The FBP alpha in neutrophil granulocytes was identified as a cytoplasmic component by indirect immunofluorescence. Gel filtration of serum revealed a peak of FBP alpha (>120 kDa), which could represent receptor...... fragments from decomposed erythrocytes and granulocytes. The soluble FBPs may exert bacteriostatic effects and protect folates in plasma from biological degradation, whereas FRs on the surface of blood cells could be involved in intracellular folate uptake or serve as signal proteins. The latter receptors...

  8. Neuronal glucose transporter isoform 3 deficient mice demonstrate features of autism spectrum disorders.

    Science.gov (United States)

    Zhao, Y; Fung, C; Shin, D; Shin, B-C; Thamotharan, S; Sankar, R; Ehninger, D; Silva, A; Devaskar, S U

    2010-03-01

    Neuronal glucose transporter (GLUT) isoform 3 deficiency in null heterozygous mice led to abnormal spatial learning and working memory but normal acquisition and retrieval during contextual conditioning, abnormal cognitive flexibility with intact gross motor ability, electroencephalographic seizures, perturbed social behavior with reduced vocalization and stereotypies at low frequency. This phenotypic expression is unique as it combines the neurobehavioral with the epileptiform characteristics of autism spectrum disorders. This clinical presentation occurred despite metabolic adaptations consisting of an increase in microvascular/glial GLUT1, neuronal GLUT8 and monocarboxylate transporter isoform 2 concentrations, with minimal to no change in brain glucose uptake but an increase in lactate uptake. Neuron-specific glucose deficiency has a negative impact on neurodevelopment interfering with functional competence. This is the first description of GLUT3 deficiency that forms a possible novel genetic mechanism for pervasive developmental disorders, such as the neuropsychiatric autism spectrum disorders, requiring further investigation in humans.

  9. Synaptic activity-related classical protein kinase C isoform localization in the adult rat neuromuscular synapse.

    Science.gov (United States)

    Besalduch, Núria; Tomàs, Marta; Santafé, Manel M; Garcia, Neus; Tomàs, Josep; Lanuza, Maria Angel

    2010-01-10

    Protein kinase C (PKC) is essential for signal transduction in a variety of cells, including neurons and myocytes, and is involved in both acetylcholine release and muscle fiber contraction. Here, we demonstrate that the increases in synaptic activity by nerve stimulation couple PKC to transmitter release in the rat neuromuscular junction and increase the level of alpha, betaI, and betaII isoforms in the membrane when muscle contraction follows the stimulation. The phosphorylation activity of these classical PKCs also increases. It seems that the muscle has to contract in order to maintain or increase classical PKCs in the membrane. We use immunohistochemistry to show that PKCalpha and PKCbetaI were located in the nerve terminals, whereas PKCalpha and PKCbetaII were located in the postsynaptic and the Schwann cells. Stimulation and contraction do not change these cellular distributions, but our results show that the localization of classical PKC isoforms in the membrane is affected by synaptic activity.

  10. FoxP2 isoforms delineate spatiotemporal transcriptional networks for vocal learning in the zebra finch

    Science.gov (United States)

    Day, Nancy F; Kimball, Todd Haswell; Aamodt, Caitlin M; Heston, Jonathan B; Hilliard, Austin T; Xiao, Xinshu; White, Stephanie A

    2018-01-01

    Human speech is one of the few examples of vocal learning among mammals yet ~half of avian species exhibit this ability. Its neurogenetic basis is largely unknown beyond a shared requirement for FoxP2 in both humans and zebra finches. We manipulated FoxP2 isoforms in Area X, a song-specific region of the avian striatopallidum analogous to human anterior striatum, during a critical period for song development. We delineate, for the first time, unique contributions of each isoform to vocal learning. Weighted gene coexpression network analysis of RNA-seq data revealed gene modules correlated to singing, learning, or vocal variability. Coexpression related to singing was found in juvenile and adult Area X whereas coexpression correlated to learning was unique to juveniles. The confluence of learning and singing coexpression in juvenile Area X may underscore molecular processes that drive vocal learning in young zebra finches and, by analogy, humans. PMID:29360038

  11. Accuracy and Precision of Radioactivity Quantification in Nuclear Medicine Images

    Science.gov (United States)

    Frey, Eric C.; Humm, John L.; Ljungberg, Michael

    2012-01-01

    The ability to reliably quantify activity in nuclear medicine has a number of increasingly important applications. Dosimetry for targeted therapy treatment planning or for approval of new imaging agents requires accurate estimation of the activity in organs, tumors, or voxels at several imaging time points. Another important application is the use of quantitative metrics derived from images, such as the standard uptake value commonly used in positron emission tomography (PET), to diagnose and follow treatment of tumors. These measures require quantification of organ or tumor activities in nuclear medicine images. However, there are a number of physical, patient, and technical factors that limit the quantitative reliability of nuclear medicine images. There have been a large number of improvements in instrumentation, including the development of hybrid single-photon emission computed tomography/computed tomography and PET/computed tomography systems, and reconstruction methods, including the use of statistical iterative reconstruction methods, which have substantially improved the ability to obtain reliable quantitative information from planar, single-photon emission computed tomography, and PET images. PMID:22475429

  12. Human reliability assessors guide: an overview

    International Nuclear Information System (INIS)

    Humphreys, P.

    1988-01-01

    The Human Reliability Assessors Guide 1 provides a review of techniques currently available for the quantification of Human Error Probabilities. The Guide has two main objectives. The first is to provide a clear and comprehensive description of eight major techniques which can be used to assess human reliability. This is supplemented by case studies taken from practical applications of each technique to industrial problems. The second objective is to provide practical guidelines for the selection of techniques. The selection process is aided by reference to a set of criteria against which each of the eight techniques have been evaluated. Utilising the criteria and critiques, a selection method is presented. This is designed to assist the potential user in choosing the technique, or combination of techniques, most suited to answering the users requirements. For each of the eight selected techniques, a summary of the origins of the technique is provided, together with a method description, detailed case studies, abstracted case studies and supporting references. (author)

  13. Guided Wave Delamination Detection and Quantification With Wavefield Data Analysis

    Science.gov (United States)

    Tian, Zhenhua; Campbell Leckey, Cara A.; Seebo, Jeffrey P.; Yu, Lingyu

    2014-01-01

    Unexpected damage can occur in aerospace composites due to impact events or material stress during off-nominal loading events. In particular, laminated composites are susceptible to delamination damage due to weak transverse tensile and inter-laminar shear strengths. Developments of reliable and quantitative techniques to detect delamination damage in laminated composites are imperative for safe and functional optimally-designed next-generation composite structures. In this paper, we investigate guided wave interactions with delamination damage and develop quantification algorithms by using wavefield data analysis. The trapped guided waves in the delamination region are observed from the wavefield data and further quantitatively interpreted by using different wavenumber analysis methods. The frequency-wavenumber representation of the wavefield shows that new wavenumbers are present and correlate to trapped waves in the damage region. These new wavenumbers are used to detect and quantify the delamination damage through the wavenumber analysis, which can show how the wavenumber changes as a function of wave propagation distance. The location and spatial duration of the new wavenumbers can be identified, providing a useful means not only for detecting the presence of delamination damage but also allowing for estimation of the delamination size. Our method has been applied to detect and quantify real delamination damage with complex geometry (grown using a quasi-static indentation technique). The detection and quantification results show the location, size, and shape of the delamination damage.

  14. Quantification of margins and uncertainties: Alternative representations of epistemic uncertainty

    International Nuclear Information System (INIS)

    Helton, Jon C.; Johnson, Jay D.

    2011-01-01

    In 2001, the National Nuclear Security Administration of the U.S. Department of Energy in conjunction with the national security laboratories (i.e., Los Alamos National Laboratory, Lawrence Livermore National Laboratory and Sandia National Laboratories) initiated development of a process designated Quantification of Margins and Uncertainties (QMU) for the use of risk assessment methodologies in the certification of the reliability and safety of the nation's nuclear weapons stockpile. A previous presentation, 'Quantification of Margins and Uncertainties: Conceptual and Computational Basis,' describes the basic ideas that underlie QMU and illustrates these ideas with two notional examples that employ probability for the representation of aleatory and epistemic uncertainty. The current presentation introduces and illustrates the use of interval analysis, possibility theory and evidence theory as alternatives to the use of probability theory for the representation of epistemic uncertainty in QMU-type analyses. The following topics are considered: the mathematical structure of alternative representations of uncertainty, alternative representations of epistemic uncertainty in QMU analyses involving only epistemic uncertainty, and alternative representations of epistemic uncertainty in QMU analyses involving a separation of aleatory and epistemic uncertainty. Analyses involving interval analysis, possibility theory and evidence theory are illustrated with the same two notional examples used in the presentation indicated above to illustrate the use of probability to represent aleatory and epistemic uncertainty in QMU analyses.

  15. Antibiotic Resistome: Improving Detection and Quantification Accuracy for Comparative Metagenomics.

    Science.gov (United States)

    Elbehery, Ali H A; Aziz, Ramy K; Siam, Rania

    2016-04-01

    The unprecedented rise of life-threatening antibiotic resistance (AR), combined with the unparalleled advances in DNA sequencing of genomes and metagenomes, has pushed the need for in silico detection of the resistance potential of clinical and environmental metagenomic samples through the quantification of AR genes (i.e., genes conferring antibiotic resistance). Therefore, determining an optimal methodology to quantitatively and accurately assess AR genes in a given environment is pivotal. Here, we optimized and improved existing AR detection methodologies from metagenomic datasets to properly consider AR-generating mutations in antibiotic target genes. Through comparative metagenomic analysis of previously published AR gene abundance in three publicly available metagenomes, we illustrate how mutation-generated resistance genes are either falsely assigned or neglected, which alters the detection and quantitation of the antibiotic resistome. In addition, we inspected factors influencing the outcome of AR gene quantification using metagenome simulation experiments, and identified that genome size, AR gene length, total number of metagenomics reads and selected sequencing platforms had pronounced effects on the level of detected AR. In conclusion, our proposed improvements in the current methodologies for accurate AR detection and resistome assessment show reliable results when tested on real and simulated metagenomic datasets.

  16. Reliability in the utility computing era: Towards reliable Fog computing

    DEFF Research Database (Denmark)

    Madsen, Henrik; Burtschy, Bernard; Albeanu, G.

    2013-01-01

    This paper considers current paradigms in computing and outlines the most important aspects concerning their reliability. The Fog computing paradigm as a non-trivial extension of the Cloud is considered and the reliability of the networks of smart devices are discussed. Combining the reliability...... requirements of grid and cloud paradigms with the reliability requirements of networks of sensor and actuators it follows that designing a reliable Fog computing platform is feasible....

  17. Revealing the functions of the transketolase enzyme isoforms in Rhodopseudomonas palustris using a systems biology approach.

    Directory of Open Access Journals (Sweden)

    Chia-Wei Hu

    Full Text Available BACKGROUND: Rhodopseudomonas palustris (R. palustris is a purple non-sulfur anoxygenic phototrophic bacterium that belongs to the class of proteobacteria. It is capable of absorbing atmospheric carbon dioxide and converting it to biomass via the process of photosynthesis and the Calvin-Benson-Bassham (CBB cycle. Transketolase is a key enzyme involved in the CBB cycle. Here, we reveal the functions of transketolase isoforms I and II in R. palustris using a systems biology approach. METHODOLOGY/PRINCIPAL FINDINGS: By measuring growth ability, we found that transketolase could enhance the autotrophic growth and biomass production of R. palustris. Microarray and real-time quantitative PCR revealed that transketolase isoforms I and II were involved in different carbon metabolic pathways. In addition, immunogold staining demonstrated that the two transketolase isoforms had different spatial localizations: transketolase I was primarily associated with the intracytoplasmic membrane (ICM but transketolase II was mostly distributed in the cytoplasm. Comparative proteomic analysis and network construction of transketolase over-expression and negative control (NC strains revealed that protein folding, transcriptional regulation, amino acid transport and CBB cycle-associated carbon metabolism were enriched in the transketolase I over-expressed strain. In contrast, ATP synthesis, carbohydrate transport, glycolysis-associated carbon metabolism and CBB cycle-associated carbon metabolism were enriched in the transketolase II over-expressed strain. Furthermore, ATP synthesis assays showed a significant increase in ATP synthesis in the transketolase II over-expressed strain. A PEPCK activity assay showed that PEPCK activity was higher in transketolase over-expressed strains than in the negative control strain. CONCLUSIONS/SIGNIFICANCE: Taken together, our results indicate that the two isoforms of transketolase in R. palustris could affect photoautotrophic growth

  18. Muscle-Type Specific Autophosphorylation of CaMKII Isoforms after Paced Contractions

    Directory of Open Access Journals (Sweden)

    Wouter Eilers

    2014-01-01

    Full Text Available We explored to what extent isoforms of the regulator of excitation-contraction and excitation-transcription coupling, calcium/calmodulin protein kinase II (CaMKII contribute to the specificity of myocellular calcium sensing between muscle types and whether concentration transients in its autophosphorylation can be simulated. CaMKII autophosphorylation at Thr287 was assessed in three muscle compartments of the rat after slow or fast motor unit-type stimulation and was compared against a computational model (CaMuZclE coupling myocellular calcium dynamics with CaMKII Thr287 phosphorylation. Qualitative differences existed between fast- (gastrocnemius medialis and slow-type muscle (soleus for the expression pattern of CaMKII isoforms. Phospho-Thr287 content of δA CaMKII, associated with nuclear functions, demonstrated a transient and compartment-specific increase after excitation, which contrasted to the delayed autophosphorylation of the sarcoplasmic reticulum-associated βM CaMKII. In soleus muscle, excitation-induced δA CaMKII autophosphorylation demonstrated frequency dependence (P = 0.02. In the glycolytic compartment of gastrocnemius medialis, CaMKII autophosphorylation after excitation was blunted. In silico assessment emphasized the importance of mitochondrial calcium buffer capacity for excitation-induced CaMKII autophosphorylation but did not predict its isoform specificity. The findings expose that CaMKII autophosphorylation with paced contractions is regulated in an isoform and muscle type-specific fashion and highlight properties emerging for phenotype-specific regulation of CaMKII.

  19. Heterogeneity of serum gelatinases MMP-2 and MMP-9 isoforms and charge variants

    Science.gov (United States)

    Rossano, Rocco; Larocca, Marilena; Riviello, Lea; Coniglio, Maria Gabriella; Vandooren, Jennifer; Liuzzi, Grazia Maria; Opdenakker, Ghislain; Riccio, Paolo

    2014-01-01

    The matrix metalloproteinases (MMPs) gelatinase A (MMP-2) and gelatinase B (MMP-9) are mediators of brain injury in multiple sclerosis (MS) and valuable biomarkers of disease activity. We applied bidimensional zymography (2-DZ) as an extension of classic monodimensional zymography (1-DZ) to analyse the complete pattern of isoforms and post-translational modifications of both MMP-9 and MMP-2 present in the sera of MS patients. The enzymes were separated on the basis of their isoelectric points (pI) and apparent molecular weights (Mw) and identified both by comparison with standard enzyme preparations and by Western blot analysis. Two MMP-2 isoforms, and at least three different isoforms and two different states of organization of MMP-9 (the multimeric MMP-9 and the N-GAL-MMP-9 complex) were observed. In addition, 2-DZ revealed for the first time that all MMP-9 and MMP-2 isoforms actually exist in the form of charge variants: four or five variants in the N-GAL complex, more charge variants in the case of MMP-9; and five to seven charge variants for MMP-2. Charge variants were also observed in recombinant enzymes and, after concentration, also in sera from healthy individuals. Sialylation (MMP-9) and phosphorylation (MMP-2) contributed to molecular heterogeneity. The detection of charge variants of MMP-9 and MMP-2 in MS serum samples illustrates the power of 2-DZ and demonstrates that in previous studies MMP mixtures, rather than single molecules, were analysed. These observations open perspectives for better diagnosis and prognosis of many diseases and need to be critically interpreted when applying other methods for MS and other diseases. PMID:24616914

  20. Cancer metabolism meets systems biology: Pyruvate kinase isoform PKM2 is a metabolic master regulator

    OpenAIRE

    Fabian V Filipp

    2013-01-01

    Pyruvate kinase activity is controlled by a tightly woven regulatory network. The oncofetal isoform of pyruvate kinase (PKM2) is a master regulator of cancer metabolism. PKM2 engages in parallel, feed-forward, positive and negative feedback control contributing to cancer progression. Besides its metabolic role, non-metabolic functions of PKM2 as protein kinase and transcriptional coactivator for c-MYC and hypoxia-inducible factor 1-alpha are essential for epidermal growth factor receptor acti...

  1. Characterization of Alien isoforms in vertebrates : Caracterización de isoformas de Alien en vertebrados

    OpenAIRE

    Tenbaum, Stephan

    2002-01-01

    Alien protein isoforms have been described to be involved in a number of biological processes. Alienalpha is a corepressor of the thyroid hormone receptor mediating transcriptional repression in a ligand-sensitive manner. Furthermore, Alienalpha is a corepressor for the orphan receptor DAX1 and the vitamin-D3 receptor. Alienbetta/CSN2 is part of the COP9-signalosome complex that acts in protein phosphorylation, protein degradation and cell cycle regulation. The major goal of this...

  2. Signal transduction by normal isoforms and W mutant variants of the Kit receptor tyrosine kinase.

    OpenAIRE

    Reith, A D; Ellis, C; Lyman, S D; Anderson, D M; Williams, D E; Bernstein, A; Pawson, T

    1991-01-01

    Germline mutations at the Dominant White Spotting (W) and Steel (Sl) loci have provided conclusive genetic evidence that c-kit mediated signal transduction pathways are essential for normal mouse development. We have analysed the interactions of normal and mutant W/c-kit gene products with cytoplasmic signalling proteins, using transient c-kit expression assays in COS cells. In addition to the previously identified c-kit gene product (Kit+), a second normal Kit isoform (KitA+) containing an i...

  3. Role of L-type Ca2+ channel isoforms in the extinction of conditioned fear.

    Science.gov (United States)

    Busquet, Perrine; Hetzenauer, Alfred; Sinnegger-Brauns, Martina J; Striessnig, Jörg; Singewald, Nicolas

    2008-05-01

    Dihydropyridine (DHP) L-type Ca(2+) channel (LTCC) antagonists, such as nifedipine, have been reported to impair the extinction of conditioned fear without interfering with its acquisition. Identification of the LTCC isoforms mediating this DHP effect is an essential basis to reveal their role as potential drug targets for the treatment of specific anxiety disorders. Ca(V)1.2 and Ca(V)1.3 are the predominant LTCCs in the mammalian brain. However, since no isoform-selective DHP blockers are available, their individual contribution to fear memory extinction is unknown. We used a novel mouse model expressing DHP-insensitive Ca(V)1.2 LTCCs (Ca(V)1.2DHP(-/-) mice) to address this question. In line with previous studies, wild-type (WT) mice treated with systemic nifedipine displayed markedly impaired fear extinction. This DHP effect was completely abolished in Ca(V)1.2DHP(-/-) mice, indicating that it is mediated by Ca(V)1.2, but not by Ca(V)1.3 LTCCs. Supporting this conclusion, Ca(V)1.3-deficient mice (Ca(V)1.3(-/-)) showed extinction identical to the respective WT mice. The inhibition of fear extinction was not observed after intracerebroventricular (i.c.v.) application of different doses of nifedipine, suggesting that this effect is secondary to inhibition of peripheral Ca(V)1.2 channels. The LTCC activator BayK, which lacks neurotoxic effects in Ca(V)1.2DHP(-/-) mice, did not influence the extinction time course. In summary, we demonstrate that LTCC signaling through the Ca(V)1.2 isoform of LTCCs interferes with fear memory extinction, presumably via a peripherally mediated mechanism. Activation of other LTCC isoforms (predominantly Ca(V)1.3) is not sufficient to accelerate extinction of conditioned fear in mice.

  4. Heterogeneous effects of M-CSF isoforms on the progression of MLL-AF9 leukemia.

    Science.gov (United States)

    Wang, Rong; Feng, Wenli; Yang, Feifei; Yang, Xiao; Wang, Lina; Chen, Chong; Hu, Yuting; Ren, Qian; Zheng, Guoguang

    2018-02-01

    Macrophage colony-stimulating factor (M-CSF) regulates both malignant cells and microenvironmental cells. Its splicing isoforms show functional heterogeneity. However, their roles on leukemia have not been well established. Here, the expression of total M-CSF in patients with hematopoietic malignancies was analyzed. The roles of M-CSF isoforms on the progression of acute myeloid leukemia (AML) were studied by establishing MLL-AF9-induced mouse AML models with high level membrane-bound M-CSF (mM-CSF) or soluble M-CSF (sM-CSF). Total M-CSF was highly expressed in myeloid leukemia patients. Furthermore, mM-CSF but not sM-CSF prolonged the survival of leukemia mice. While sM-CSF was more potent to promote proliferation and self-renew, mM-CSF was more potent to promote differentiation. Moreover, isoforms had different effects on leukemia-associated macrophages (LAMs) though they both increase monocytes/macrophages by growth-promoting and recruitment effects. In addition, mM-CSF promoted specific phagocytosis of leukemia cells by LAMs. RNA-seq analysis revealed that mM-CSF enhanced phagocytosis-associated genes and activated oxidative phosphorylation and metabolism pathway. These results highlight heterogeneous effects of M-CSF isoforms on AML progression and the mechanisms of mM-CSF, that is, intrinsically promoting AML cell differentiation and extrinsically enhancing infiltration of macrophages and phagocytosis by macrophages, which may provide potential clues for clinical diagnosis and therapy. © 2017 Australasian Society for Immunology Inc.

  5. Role of cyclooxygenase isoforms in the altered excitatory motor pathways of human colon with diverticular disease.

    Science.gov (United States)

    Fornai, M; Colucci, R; Antonioli, L; Ippolito, C; Segnani, C; Buccianti, P; Marioni, A; Chiarugi, M; Villanacci, V; Bassotti, G; Blandizzi, C; Bernardini, N

    2014-08-01

    The COX isoforms (COX-1, COX-2) regulate human gut motility, although their role under pathological conditions remains unclear. This study examines the effects of COX inhibitors on excitatory motility in colonic tissue from patients with diverticular disease (DD). Longitudinal muscle preparations, from patients with DD or uncomplicated cancer (controls), were set up in organ baths and connected to isotonic transducers. Indomethacin (COX-1/COX-2 inhibitor), SC-560 (COX-1 inhibitor) or DFU (COX-2 inhibitor) were assayed on electrically evoked, neurogenic, cholinergic and tachykininergic contractions, or carbachol- and substance P (SP)-induced myogenic contractions. Distribution and expression of COX isoforms in the neuromuscular compartment were assessed by RT-PCR, Western blot and immunohistochemical analysis. In control preparations, neurogenic cholinergic contractions were enhanced by COX inhibitors, whereas tachykininergic responses were blunted. Carbachol-evoked contractions were increased by indomethacin or SC-560, but not DFU, whereas all inhibitors reduced SP-induced motor responses. In preparations from DD patients, COX inhibitors did not affect electrically evoked cholinergic contractions. Both indomethacin and DFU, but not SC-560, decreased tachykininergic responses. COX inhibitors did not modify carbachol-evoked motor responses, whereas they counteracted SP-induced contractions. COX-1 expression was decreased in myenteric neurons, whereas COX-2 was enhanced in glial cells and smooth muscle. In control colon, COX-1 and COX-2 down-regulate cholinergic motility, whereas both isoforms enhance tachykininergic motor activity. In the presence of DD, there is a loss of modulation by both COX isoforms on the cholinergic system, whereas COX-2 displays an enhanced facilitatory control on tachykininergic contractile activity. © 2014 The British Pharmacological Society.

  6. Isoform-specific regulation of osteogenic factors by polypeptide N-Acetylgalactosaminyltransferases 1 and 4

    International Nuclear Information System (INIS)

    Tang, Juan; Zheng, Hanxi; Chen, Ling; Gao, Shangshang; Shi, Xiaorui; Liu, Jingjing; Xu, Lan

    2017-01-01

    The family of UDP-GalNAc polypeptide: N-Acetylgalactosaminlytransfersases (ppGalNAcTs) catalyzes the initial step of O-linked protein glycosylation. Mucin-type O-glycoproteins are abundant in the bone and may play an important role in osteogenesis. Herein, we examined the effects of ppGalNAc-T isoforms on osteogenesis of MC3T3-E1 pre-osteoblasts. We found that ppGalNAc-T1 and -T4 isoforms were highly expressed during osteogenesis of MC3T3-E1 and their knockdown by short hairpin RNA (shRNA) decreased osteoblast formation and bone mineralization. Knockdown of ppGalNAc-T1 or -T4 decreased mRNA and protein levels of bone sialoprotein (BSP). Knockdown of ppGalNAc-T1decreased mRNA levels of osteocalcin (OC), osteoprotegerin (OPG). Knockdown ofppGalNAc-T4 isoform decreased mRNA levels of OC, OPG and vitamin D receptor (VDR). While knockdown of T1 or T4 isoforms did not change the expression of osteopontin (OPN), COLLI, receptor activator for nuclear factor-κB ligand (RANKL) and transforming growth factor-β (TGF-β). Our results demonstrated that the ppGalNAc-T4 was highly expressed in MC3T3-E1 cells during osteogenesis for the first time. We also found that ppGalNAc-T1 and -T4 affected the expression of different osteogenic factors, suggesting distinct roles ppGalNAc-T isoformsplay in regulating osteogenesis in vitro. - Highlights: • ppGalNAc-T1 and T4 are highly expressed during MC3T3 cell osteogenesis. • Knockdown of ppGalNAc-T1 and -T4 decreases osteogenic differentiation and mineralization. • Expression of osteogenic factors are differentially affected by decreased ppGalNAc-T1 and -T4 expression.

  7. Smoking specifically induces metallothionein-2 isoform in human placenta at term

    International Nuclear Information System (INIS)

    Ronco, Ana Maria; Garrido, Fernando; Llanos, Miguel N.

    2006-01-01

    Recently, we reported the presence of higher levels of metallothionein (MT) in placentas of smokers compared to non-smokers. In the present study, we designed experiments to separate and evaluate two isoforms of MT (MT-1 and MT-2) in placentas of smokers and non-smokers. Metallothionein was extracted and separated by ion-exchange high performance liquid chromatography (HPLC), previous saturation with cadmium chloride. Two peaks eluting at 6 and 12.5 min, corresponding to MT-1 and MT-2, respectively, were obtained. Metallothionein present in both peaks was identified by Western blot analysis using a monoclonal antibody directed against MT-1 and MT-2. Each isoform concentration was calculated after measuring its cadmium content by atomic absorption spectrometry with inductively coupled-plasma. In placentas of smokers, MT-2 levels increased by seven-fold compared to non-smokers, whereas MT-1 was not changed. Total placental cadmium and zinc concentrations, determined by atomic absorption spectrometry and neutron activation analysis, respectively, were higher in smokers. Metallothioneins levels were clearly in excess to bind all cadmium ions present in placentas. However, most of placental zinc remains unbound to MTs, although as much as twice zinc ions could be bound to MT in smokers. In conclusion, MT-2 is the main isoform induced by smoking, suggesting that this isoform could be involved in placental cadmium and zinc retention. This fact, which could contribute to reduce the transference of zinc to the fetus, may be associated to detrimental effects on fetal growth and development

  8. The Role of a Novel Myosin Isoform in Prostate Cancer Metastasis

    Science.gov (United States)

    2013-10-01

    2013 Accepted 14 February 2013 Available online 21 February 2013 Keywords: Myosin IC Isoforms Nucleolar localization signal Nucleolus Nucleus RNA...polymerase I Fibrillarinnt matter & 2013 Elsevier 1016/j.yexcr.2013.02.008 S, nucleolar localization ; No, nucleolus ; N, nucle bovine serum albumin; S...the nucleus, and the nucleolus . In the cytoplasm, myosin IC associ- ates with membranes and is involved in vesicle transport of membrane proteins [2

  9. Deregulation of the endogenous C/EBPβ LIP isoform predisposes to tumorigenesis.

    Science.gov (United States)

    Bégay, Valérie; Smink, Jeske J; Loddenkemper, Christoph; Zimmermann, Karin; Rudolph, Cornelia; Scheller, Marina; Steinemann, Doris; Leser, Ulf; Schlegelberger, Brigitte; Stein, Harald; Leutz, Achim

    2015-01-01

    Two long and one truncated isoforms (termed LAP*, LAP, and LIP, respectively) of the transcription factor CCAAT enhancer binding protein beta (C/EBPβ) are expressed from a single intronless Cebpb gene by alternative translation initiation. Isoform expression is sensitive to mammalian target of rapamycin (mTOR)-mediated activation of the translation initiation machinery and relayed through an upstream open reading frame (uORF) on the C/EBPβ mRNA. The truncated C/EBPβ LIP, initiated by high mTOR activity, has been implied in neoplasia, but it was never shown whether endogenous C/EBPβ LIP may function as an oncogene. In this study, we examined spontaneous tumor formation in C/EBPβ knockin mice that constitutively express only the C/EBPβ LIP isoform from its own locus. Our data show that deregulated C/EBPβ LIP predisposes to oncogenesis in many tissues. Gene expression profiling suggests that C/EBPβ LIP supports a pro-tumorigenic microenvironment, resistance to apoptosis, and alteration of cytokine/chemokine expression. The results imply that enhanced translation reinitiation of C/EBPβ LIP promotes tumorigenesis. Accordingly, pharmacological restriction of mTOR function might be a therapeutic option in tumorigenesis that involves enhanced expression of the truncated C/EBPβ LIP isoform. Elevated C/EBPβ LIP promotes cancer in mice. C/EBPβ LIP is upregulated in B-NHL. Deregulated C/EBPβ LIP alters apoptosis and cytokine/chemokine networks. Deregulated C/EBPβ LIP may support a pro-tumorigenic microenvironment.

  10. VEGF111b, a new member of VEGFxxxb isoforms and induced by mitomycin C, inhibits angiogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Fang; Li, Xiuli [Department of Obstetrics and Gynecology, Chinese PLA General Hospital, Beijing (China); Kong, Jian [Department of Hepatobiliary Surgery, Beijing Chaoyang Hospital, Capital Medical University, Beijing (China); Pan, Bing [The Institute of Cardiovascular Sciences, Peking University Health Science Center, Key Laboratory of Molecular Cardiovascular Sciences of Education Ministry, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides of Health Ministry, Beijing (China); Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Key Laboratory of Molecular Cardiovascular Sciences of Education Ministry, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides of Health Ministry, Beijing (China); Sun, Min [Department of Obstetrics and Gynecology, Tangdu Hospital, Fourth Military Medical University, Xian (China); Zheng, Lemin, E-mail: zhengl@bjmu.edu.cn [The Institute of Cardiovascular Sciences, Peking University Health Science Center, Key Laboratory of Molecular Cardiovascular Sciences of Education Ministry, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides of Health Ministry, Beijing (China); Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Key Laboratory of Molecular Cardiovascular Sciences of Education Ministry, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides of Health Ministry, Beijing (China); Yao, Yuanqing, E-mail: yqyao@126.com [Department of Obstetrics and Gynecology, Chinese PLA General Hospital, Beijing (China)

    2013-11-08

    Highlights: •We discovered a new member of VEGFxxxb family-VEGF111b. •We found VEGF111b mRNA and protein can be induced by mitomycin C. •We confirmed VEGF111b over-expression inhibits angiogenesis. •VEGF111b inhibits angiogenesis through inhibiting VEGF-R2/PI3K/Akt and VEGF-R2/ERK1/2 phosphorylation. -- Abstract: Vascular endothelial growth factor (VEGF-A) stimulating angiogenesis is required for tumor growth and progression. The conventional VEGF-A isoforms have been considered as pro-angiogenic factors. Another family of VEGF-A isoforms generated by alternative splicing, termed VEGFxxxb isoforms, has anti-angiogenic property, exemplified by VEGF165b. Here, we identify a new number of VEGFxxx family-VEGF111b induced by mitomycin C, although not detected in mitomycin C-unexposed ovarian cancer cells. SKOV3 cells were transfected with pcDNA{sub 3.1} empty vector, pcDNA{sub 3.1}-VEGF111b or pcDNA{sub 3.1}-VEGF165b to collect conditioned mediums respectively. VEGF111b overexpression inhibits proliferation, migration and tube formation of endothelial cell by inhibiting VEGF-R2 phosphorylation and its downstream signaling, similar to VEGF165b but slightly lower than VEGF165b. The anti-angiogenic property depends on the six amino acids of exon 8b of the VEGFxxxb isoforms. Our results show that VEGF111b is a novel potent anti-angiogenic agent that can target the VEGF-R2 and its signaling pathway to inhibit ovarian tumor growth.

  11. VEGF111b, a new member of VEGFxxxb isoforms and induced by mitomycin C, inhibits angiogenesis

    International Nuclear Information System (INIS)

    Gu, Fang; Li, Xiuli; Kong, Jian; Pan, Bing; Sun, Min; Zheng, Lemin; Yao, Yuanqing

    2013-01-01

    Highlights: •We discovered a new member of VEGFxxxb family-VEGF111b. •We found VEGF111b mRNA and protein can be induced by mitomycin C. •We confirmed VEGF111b over-expression inhibits angiogenesis. •VEGF111b inhibits angiogenesis through inhibiting VEGF-R2/PI3K/Akt and VEGF-R2/ERK1/2 phosphorylation. -- Abstract: Vascular endothelial growth factor (VEGF-A) stimulating angiogenesis is required for tumor growth and progression. The conventional VEGF-A isoforms have been considered as pro-angiogenic factors. Another family of VEGF-A isoforms generated by alternative splicing, termed VEGFxxxb isoforms, has anti-angiogenic property, exemplified by VEGF165b. Here, we identify a new number of VEGFxxx family-VEGF111b induced by mitomycin C, although not detected in mitomycin C-unexposed ovarian cancer cells. SKOV3 cells were transfected with pcDNA 3.1 empty vector, pcDNA 3.1 -VEGF111b or pcDNA 3.1 -VEGF165b to collect conditioned mediums respectively. VEGF111b overexpression inhibits proliferation, migration and tube formation of endothelial cell by inhibiting VEGF-R2 phosphorylation and its downstream signaling, similar to VEGF165b but slightly lower than VEGF165b. The anti-angiogenic property depends on the six amino acids of exon 8b of the VEGFxxxb isoforms. Our results show that VEGF111b is a novel potent anti-angiogenic agent that can target the VEGF-R2 and its signaling pathway to inhibit ovarian tumor growth

  12. RTE - Reliability report 2016

    International Nuclear Information System (INIS)

    2017-06-01

    Every year, RTE produces a reliability report for the past year. This document lays out the main factors that affected the electrical power system's operational reliability in 2016 and the initiatives currently under way intended to ensure its reliability in the future. Within a context of the energy transition, changes to the European interconnected network mean that RTE has to adapt on an on-going basis. These changes include the increase in the share of renewables injecting an intermittent power supply into networks, resulting in a need for flexibility, and a diversification in the numbers of stakeholders operating in the energy sector and changes in the ways in which they behave. These changes are dramatically changing the structure of the power system of tomorrow and the way in which it will operate - particularly the way in which voltage and frequency are controlled, as well as the distribution of flows, the power system's stability, the level of reserves needed to ensure supply-demand balance, network studies, assets' operating and control rules, the tools used and the expertise of operators. The results obtained in 2016 are evidence of a globally satisfactory level of reliability for RTE's operations in somewhat demanding circumstances: more complex supply-demand balance management, cross-border schedules at interconnections indicating operation that is closer to its limits and - most noteworthy - having to manage a cold spell just as several nuclear power plants had been shut down. In a drive to keep pace with the changes expected to occur in these circumstances, RTE implemented numerous initiatives to ensure high levels of reliability: - maintaining investment levels of euro 1.5 billion per year; - increasing cross-zonal capacity at borders with our neighbouring countries, thus bolstering the security of our electricity supply; - implementing new mechanisms (demand response, capacity mechanism, interruptibility, etc.); - involvement in tests or projects

  13. Novel isoforms of the TFIID subunit TAF4 modulate nuclear receptor-mediated transcriptional activity

    International Nuclear Information System (INIS)

    Brunkhorst, Adrian; Neuman, Toomas; Hall, Anita; Arenas, Ernest; Bartfai, Tamas; Hermanson, Ola; Metsis, Madis

    2004-01-01

    The transcription factor TFIID consists of TATA-binding protein (TBP) and TBP-associated factors (TAFs). TAFs are essential for modulation of transcriptional activity but the regulation of TAFs is complex and many important aspects remain unclear. In this study, we have identified and characterized five novel truncated forms of the TFIID subunit TAF4 (TAF II 135). Analysis of the mouse gene structure revealed that all truncations were the results of alternative splicing and resulted in the loss of domains or parts of domains implicated in TAF4 functional interactions. Results from transcriptional assays showed that several of the TAF4 isoforms exerted dominant negative effects on TAF4 activity in nuclear receptor-mediated transcriptional activation. In addition, alternative TAF4 isoforms could be detected in specific cell types. Our results indicate an additional level of complexity in TAF4-mediated regulation of transcription and suggest context-specific roles for these new TAF4 isoforms in transcriptional regulation in vivo

  14. Peroxiredoxin isoforms are associated with cardiovascular risk factors in type 2 diabetes mellitus

    Energy Technology Data Exchange (ETDEWEB)

    El Eter, E. [Physiology Department, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Cardiovascular Research Group, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Physiology Department, Faculty of Medicine, Alexandria University, Alexandria (Egypt); Al-Masri, A.A. [Physiology Department, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Cardiovascular Research Group, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia)

    2015-03-03

    The production of oxygen free radicals in type 2 diabetes mellitus contributes to the development of complications, especially the cardiovascular-related ones. Peroxiredoxins (PRDXs) are antioxidant enzymes that combat oxidative stress. The aim of this study was to investigate the associations between the levels of PRDX isoforms (1, 2, 4, and 6) and cardiovascular risk factors in type 2 diabetes mellitus. Fifty-three patients with type 2 diabetes mellitus (28F/25M) and 25 healthy control subjects (7F/18M) were enrolled. We measured the plasma levels of each PRDX isoform and analyzed their correlations with cardiovascular risk factors. The plasma PRDX1, -2, -4, and -6 levels were higher in the diabetic patients than in the healthy control subjects. PRDX2 and -6 levels were negatively correlated with diastolic blood pressure, fasting blood sugar, and hemoglobin A1c. In contrast, PRDX1 levels were positively correlated with low-density lipoprotein and C-reactive protein levels. PRDX4 levels were negatively correlated with triglycerides. In conclusion, PRDX1, -2, -4, and -6 showed differential correlations with a variety of traditional cardiovascular risk factors. These results should encourage further research into the crosstalk between PRDX isoforms and cardiovascular risk factors.

  15. Differential expression of a new isoform of DLG2 in renal oncocytoma

    Directory of Open Access Journals (Sweden)

    Kovacs Gyula

    2006-04-01

    Full Text Available Abstract Background Renal oncocytoma, a benign tumour of the kidney, may pose a differential diagnostic problem due to overlapping phenotype with chromophobe renal cell carcinoma or other types of renal cell tumours. Therefore, identification of molecular markers would be of great value for molecular diagnostics of this tumour type. Methods In the current study we applied various techniques, including Affymetrix microarray hybridization and semiquantitative RT-PCR, to identify genes expressed differentially in renal oncocytomas. Subsequently, we used RACE and Northern blot hybridization to characterize the potential candidates for molecular diagnosis. Results We have identified new isoform of DLG2 gene, which contains 3'-end exons of the known DLG2 gene along with the hypothetical gene FLJ37266. The new isoform is specifically upregulated in renal oncocytoma, whereas the known DLG2 gene is downregulated in this type of kidney tumour. Conclusion The new isoform of DLG2 is the promising candidate gene for molecular differential diagnostics of renal oncocytoma.

  16. Adaptive evolution and elucidating the potential inhibitor against schizophrenia to target DAOA (G72) isoforms.

    Science.gov (United States)

    Sehgal, Sheikh Arslan; Mannan, Shazia; Kanwal, Sumaira; Naveed, Ishrat; Mir, Asif

    2015-01-01

    Schizophrenia (SZ), a chronic mental and heritable disorder characterized by neurophysiological impairment and neuropsychological abnormalities, is strongly associated with D-amino acid oxidase activator (DAOA, G72). Research studies emphasized that overexpression of DAOA may be responsible for improper functioning of neurotransmitters, resulting in neurological disorders like SZ. In the present study, a hybrid approach of comparative modeling and molecular docking followed by inhibitor identification and structure modeling was employed. Screening was performed by two-dimensional similarity search against selected inhibitor, keeping in view the physiochemical properties of the inhibitor. Here, we report an inhibitor compound which showed maximum binding affinity against four selected isoforms of DAOA. Docking studies revealed that Glu-53, Thr-54, Lys-58, Val-85, Ser-86, Tyr-87, Leu-88, Glu-90, Leu-95, Val-98, Ser-100, Glu-112, Tyr-116, Lys-120, Asp-121, and Arg-122 are critical residues for receptor-ligand interaction. The C-terminal of selected isoforms is conserved, and binding was observed on the conserved region of isoforms. We propose that selected inhibitor might be more potent on the basis of binding energy values. Further analysis of this inhibitor through site-directed mutagenesis could be helpful for exploring the details of ligand-binding pockets. Overall, the findings of this study may be helpful in designing novel therapeutic targets to cure SZ.

  17. 55K isoform of CDK9 associates with Ku70 and is involved in DNA repair

    International Nuclear Information System (INIS)

    Liu, Hongbing; Herrmann, Christine H.; Chiang, Karen; Sung, Tzu-Ling; Moon, Sung-Hwan; Donehower, Lawrence A.; Rice, Andrew P.

    2010-01-01

    Positive elongation factor b (P-TEFb) is a cellular protein kinase that is required for RNA polymerase II (RNAP II) transcriptional elongation of protein coding genes. P-TEFb is a set of different molecular complexes, each containing CDK9 as the catalytic subunit. There are two isoforms of the CDK9 protein - the major 42 KDa CDK9 isoform and the minor 55KDa isoform that is translated from an in-frame mRNA that arises from an upstream transcriptional start site. We found that shRNA depletion of the 55K CDK9 protein in HeLa cells induces apoptosis and double-strand DNA breaks (DSBs). The levels of apoptosis and DSBs induced by the depletion were reduced by expression of a 55K CDK9 protein variant resistant to the shRNA, indicating that these phenotypes are the consequence of depletion of the 55K protein and not off-target effects. We also found that the 55K CDK9 protein, but not the 42K CDK9 protein, specifically associates with Ku70, a protein involved in DSB repair. Our findings suggest that the 55K CDK9 protein may function in repair of DNA through an association with Ku70.

  18. Dual roles for coactivator activator and its counterbalancing isoform coactivator modulator in human kidney cell tumorigenesis.

    Science.gov (United States)

    Kang, Yun Kyoung; Schiff, Rachel; Ko, Lan; Wang, Tao; Tsai, Sophia Y; Tsai, Ming-Jer; O'Malley, Bert W

    2008-10-01

    Coactivator activator (CoAA) has been reported to be a coactivator that regulates steroid receptor-mediated transcription and alternative RNA splicing. Herein, we show that CoAA is a dual-function coregulator that inhibits G(1)-S transition in human kidney cells and suppresses anchorage-independent growth and xenograft tumor formation. Suppression occurs in part by down-regulating c-myc and its downstream effectors ccnd1 and skp2 and causing accumulation of p27/Kip1 protein. In this cellular setting, CoAA directly represses the proto-oncogene c-myc by recruiting HDAC3 protein and decreasing both the acetylation of histone H3 and the presence of RNA polymerase II on the c-myc promoter. Interestingly, a splicing isoform of CoAA, coactivator modulator (CoAM), antagonizes CoAA-induced G(1)-S transition and growth inhibition by negatively regulating the mRNA levels of the endogenous CoAA isoform. In addition, we found that expression of CoAA protein is significantly decreased in human renal cell carcinoma compared with normal kidney. Our study presents evidence that CoAA is a potential tumor suppressor in renal carcinoma and that CoAM is a counterbalancing splice isoform. This is, thus far, the only example of a nuclear receptor coregulator involved in suppression of kidney cancer and suggests potentially significant new roles for coregulators in renal cancer biology.

  19. Multiple sodium channel isoforms mediate the pathological effects of Pacific ciguatoxin-1

    Science.gov (United States)

    Inserra, Marco C.; Israel, Mathilde R.; Caldwell, Ashlee; Castro, Joel; Deuis, Jennifer R.; Harrington, Andrea M.; Keramidas, Angelo; Garcia-Caraballo, Sonia; Maddern, Jessica; Erickson, Andelain; Grundy, Luke; Rychkov, Grigori Y.; Zimmermann, Katharina; Lewis, Richard J.; Brierley, Stuart M.; Vetter, Irina

    2017-01-01

    Human intoxication with the seafood poison ciguatoxin, a dinoflagellate polyether that activates voltage-gated sodium channels (NaV), causes ciguatera, a disease characterised by gastrointestinal and neurological disturbances. We assessed the activity of the most potent congener, Pacific ciguatoxin-1 (P-CTX-1), on NaV1.1–1.9 using imaging and electrophysiological approaches. Although P-CTX-1 is essentially a non-selective NaV toxin and shifted the voltage-dependence of activation to more hyperpolarising potentials at all NaV subtypes, an increase in the inactivation time constant was observed only at NaV1.8, while the slope factor of the conductance-voltage curves was significantly increased for NaV1.7 and peak current was significantly increased for NaV1.6. Accordingly, P-CTX-1-induced visceral and cutaneous pain behaviours were significantly decreased after pharmacological inhibition of NaV1.8 and the tetrodotoxin-sensitive isoforms NaV1.7 and NaV1.6, respectively. The contribution of these isoforms to excitability of peripheral C- and A-fibre sensory neurons, confirmed using murine skin and visceral single-fibre recordings, reflects the expression pattern of NaV isoforms in peripheral sensory neurons and their contribution to membrane depolarisation, action potential initiation and propagation. PMID:28225079

  20. Peroxiredoxin isoforms are associated with cardiovascular risk factors in type 2 diabetes mellitus

    International Nuclear Information System (INIS)

    El Eter, E.; Al-Masri, A.A.

    2015-01-01

    The production of oxygen free radicals in type 2 diabetes mellitus contributes to the development of complications, especially the cardiovascular-related ones. Peroxiredoxins (PRDXs) are antioxidant enzymes that combat oxidative stress. The aim of this study was to investigate the associations between the levels of PRDX isoforms (1, 2, 4, and 6) and cardiovascular risk factors in type 2 diabetes mellitus. Fifty-three patients with type 2 diabetes mellitus (28F/25M) and 25 healthy control subjects (7F/18M) were enrolled. We measured the plasma levels of each PRDX isoform and analyzed their correlations with cardiovascular risk factors. The plasma PRDX1, -2, -4, and -6 levels were higher in the diabetic patients than in the healthy control subjects. PRDX2 and -6 levels were negatively correlated with diastolic blood pressure, fasting blood sugar, and hemoglobin A1c. In contrast, PRDX1 levels were positively correlated with low-density lipoprotein and C-reactive protein levels. PRDX4 levels were negatively correlated with triglycerides. In conclusion, PRDX1, -2, -4, and -6 showed differential correlations with a variety of traditional cardiovascular risk factors. These results should encourage further research into the crosstalk between PRDX isoforms and cardiovascular risk factors

  1. Response of slow and fast muscle to hypothyroidism: maximal shortening velocity and myosin isoforms

    Science.gov (United States)

    Caiozzo, V. J.; Herrick, R. E.; Baldwin, K. M.

    1992-01-01

    This study examined both the shortening velocity and myosin isoform distribution of slow- (soleus) and fast-twitch (plantaris) skeletal muscles under hypothyroid conditions. Adult female Sprague-Dawley rats were randomly assigned to one of two groups: control (n = 7) or hypothyroid (n = 7). In both muscles, the relative contents of native slow myosin (SM) and type I myosin heavy chain (MHC) increased in response to the hypothyroid treatment. The effects were such that the hypothyroid soleus muscle expressed only the native SM and type I MHC isoforms while repressing native intermediate myosin and type IIA MHC. In the plantaris, the relative content of native SM and type I MHC isoforms increased from 5 to 13% and from 4 to 10% of the total myosin pool, respectively. Maximal shortening velocity of the soleus and plantaris as measured by the slack test decreased by 32 and 19%, respectively, in response to hypothyroidism. In contrast, maximal shortening velocity as estimated by force-velocity data decreased only in the soleus (-19%). No significant change was observed for the plantaris.

  2. Circadian rhythmicity of active GSK3 isoforms modulates molecular clock gene rhythms in the suprachiasmatic nucleus.

    Science.gov (United States)

    Besing, Rachel C; Paul, Jodi R; Hablitz, Lauren M; Rogers, Courtney O; Johnson, Russell L; Young, Martin E; Gamble, Karen L

    2015-04-01

    The suprachiasmatic nucleus (SCN) drives and synchronizes daily rhythms at the cellular level via transcriptional-translational feedback loops comprising clock genes such as Bmal1 and Period (Per). Glycogen synthase kinase 3 (GSK3), a serine/threonine kinase, phosphorylates at least 5 core clock proteins and shows diurnal variation in phosphorylation state (inactivation) of the GSK3β isoform. Whether phosphorylation of the other primary isoform (GSK3α) varies across the subjective day-night cycle is unknown. The purpose of this study was to determine if the endogenous rhythm of GSK3 (α and β) phosphorylation is critical for rhythmic BMAL1 expression and normal amplitude and periodicity of the molecular clock in the SCN. Significant circadian rhythmicity of phosphorylated GSK3 (α and β) was observed in the SCN from wild-type mice housed in constant darkness for 2 weeks. Importantly, chronic activation of both GSK3 isoforms impaired rhythmicity of the GSK3 target BMAL1. Furthermore, chronic pharmacological inhibition of GSK3 with 20 µM CHIR-99021 enhanced the amplitude and shortened the period of PER2::luciferase rhythms in organotypic SCN slice cultures. These results support the model that GSK3 activity status is regulated by the circadian clock and that GSK3 feeds back to regulate the molecular clock amplitude in the SCN. © 2015 The Author(s).

  3. Role of p73 Dinucleotide Polymorphism in Prostate Cancer and p73 Protein Isoform Balance

    Directory of Open Access Journals (Sweden)

    L. Michael Carastro

    2014-01-01

    Full Text Available Background. Molecular markers for prostate cancer (PCa risks are currently lacking. Here we address the potential association of a dinucleotide polymorphism (DNP in exon 2 of the p73 gene with PCa risk/progression and discern any disruption of p73 protein isoforms levels in cells harboring a p73 DNP allele. Methods. We investigated the association between p73 DNP genotype and PCa risk/aggressiveness and survival by fitting logistic regression models in 1,292 incident cases and 682 controls. Results. Although we detected no association between p73 DNP and PCa risk, a significant inverse relationship between p73 DNP and PCa aggressiveness (AT/AT + GC/AT versus GC/GC, OR = 0.55, 95%Cl = 0.31–0.99 was detected. Also, p73 DNP is marginally associated with overall death (dominant model, HR = 0.76, 95%Cl = 0.57–1.00, P=0.053 as well as PCa specific death (HR = 0.69, 95%Cl = 0.45–1.06, P=0.09. Western blot analyses for p73 protein isoforms indicate that cells heterozygous for the p73 DNP have lower levels of ∆Np73 relative to TAp73 (P<0.001. Conclusions. Our findings are consistent with an association between p73 DNP and low risk for PCa aggressiveness by increasing the expressed TAp73/∆Np73 protein isoform ratio.

  4. Analysis of human articular chondrocyte CD44 isoform expression and function in health and disease.

    Science.gov (United States)

    Salter, D M; Godolphin, J L; Gourlay, M S; Lawson, M F; Hughes, D E; Dunne, E

    1996-08-01

    Interactions between articular chondrocytes and components of the extracellular matrix are of potential importance in the normal function of cartilage and in the pathophysiology of arthritis. Little is known of the basis of these interactions, but cell adhesive molecules such as CD44 are likely to be involved. Immunohistology using six well-characterized anti-CD44 monoclonal antibodies demonstrated standard CD44 isoform (CD44H) expression by all chondrocytes in normal and osteoarthrotic (OA) cartilage but absence of the CD44E variant. Polymerase chain reaction (PCR) of reverse transcribed mRNA from monolayer cultures of normal and OA chondrocytes using primer sequences which span the region containing variably spliced exons produced a predominant band representing the standard form of CD44, which lacks the variable exons 6-15 (v1-v10). No product was seen at the expected size of the epithelial variant of CD44 (CD44v8-10). Use of exon-specific primers, however, showed expression of variant exons resulting in multiple minor isoforms. Standard CD44 was also shown to be the predominantly expressed isoform identified by immunoprecipitation, but human articular chondrocytes did not adhere to hyaluronan in vitro. Chondrocyte CD44 may function as an adhesion receptor for other matrix molecules such as fibronectin or collagen.

  5. The NASA Langley Multidisciplinary Uncertainty Quantification Challenge

    Science.gov (United States)

    Crespo, Luis G.; Kenny, Sean P.; Giesy, Daniel P.

    2014-01-01

    This paper presents the formulation of an uncertainty quantification challenge problem consisting of five subproblems. These problems focus on key aspects of uncertainty characterization, sensitivity analysis, uncertainty propagation, extreme-case analysis, and robust design.

  6. Uncertainty quantification theory, implementation, and applications

    CERN Document Server

    Smith, Ralph C

    2014-01-01

    The field of uncertainty quantification is evolving rapidly because of increasing emphasis on models that require quantified uncertainties for large-scale applications, novel algorithm development, and new computational architectures that facilitate implementation of these algorithms. Uncertainty Quantification: Theory, Implementation, and Applications provides readers with the basic concepts, theory, and algorithms necessary to quantify input and response uncertainties for simulation models arising in a broad range of disciplines. The book begins with a detailed discussion of applications where uncertainty quantification is critical for both scientific understanding and policy. It then covers concepts from probability and statistics, parameter selection techniques, frequentist and Bayesian model calibration, propagation of uncertainties, quantification of model discrepancy, surrogate model construction, and local and global sensitivity analysis. The author maintains a complementary web page where readers ca...

  7. Uncertainty Quantification in Aerodynamics Simulations, Phase I

    Data.gov (United States)

    National Aeronautics and Space Administration — The objective of the proposed work (Phases I and II) is to develop uncertainty quantification methodologies and software suitable for use in CFD simulations of...

  8. Quantification of virus syndrome in chili peppers

    African Journals Online (AJOL)

    Jane

    2011-06-15

    Jun 15, 2011 ... alternative for the quantification of the disease' syndromes in regards to this crop. The result of these ..... parison of treatments such as cultivars or control measures and ..... Vascular discoloration and stem necrosis. 2.

  9. Low cost high performance uncertainty quantification

    KAUST Repository

    Bekas, C.; Curioni, A.; Fedulova, I.

    2009-01-01

    Uncertainty quantification in risk analysis has become a key application. In this context, computing the diagonal of inverse covariance matrices is of paramount importance. Standard techniques, that employ matrix factorizations, incur a cubic cost

  10. Raman spectroscopy for DNA quantification in cell nucleus.

    Science.gov (United States)

    Okotrub, K A; Surovtsev, N V; Semeshin, V F; Omelyanchuk, L V

    2015-01-01

    Here we demonstrate the feasibility of a novel approach to quantify DNA in cell nuclei. This approach is based on spectroscopy analysis of Raman light scattering, and avoids the problem of nonstoichiometric binding of dyes to DNA, as it directly measures the signal from DNA. Quantitative analysis of nuclear DNA contribution to Raman spectrum could be reliably performed using intensity of a phosphate mode at 1096 cm(-1) . When compared to the known DNA standards from cells of different animals, our results matched those values at error of 10%. We therefore suggest that this approach will be useful to expand the list of DNA standards, to properly adjust the duration of hydrolysis in Feulgen staining, to assay the applicability of fuchsines for DNA quantification, as well as to measure DNA content in cells with complex hydrolysis patterns, when Feulgen densitometry is inappropriate. © 2014 International Society for Advancement of Cytometry.

  11. Mammary Gland Tumor Development in Transgenic Mice Overexpressing Different Isoforms of the CDP/Cux Transcription Factor

    National Research Council Canada - National Science Library

    Cadieux, Chantal

    2008-01-01

    Short CUX1 isoforms were found to be overexpressed in breast cancer cell lines, in human breast tumors and in uterine leiomyomas, suggesting that these proteins play a key role in tumor development and progression...

  12. Mammary Gland Tumor Development in Transgenic Mice Overexpressing Different Isoforms of the CDP/Cux Transcription Factor

    National Research Council Canada - National Science Library

    Cadieux, Chantal

    2007-01-01

    Short CDP/Cux isoforms were found to be overexpressed in breast cancer cell lines, in human breast tumors and in uterine leiomyomas, suggesting that these proteins play a key role in tumor development and progression...

  13. Relative Expression Levels Rather Than Specific Activity Plays the Major Role in Determining In Vivo AKT Isoform Substrate Specificity

    Directory of Open Access Journals (Sweden)

    Rachel S. Lee

    2011-01-01

    Full Text Available The AKT protooncogene mediates many cellular processes involved in normal development and disease states such as cancer. The three structurally similar isoforms: AKT1, AKT2, and AKT3 exhibit both functional redundancy and isoform-specific functions; however the basis for their differential signalling remains unclear. Here we show that in vitro, purified AKT3 is ∼47-fold more active than AKT1 at phosphorylating peptide and protein substrates. Despite these marked variations in specific activity between the individual isoforms, a comprehensive analysis of phosphorylation of validated AKT substrates indicated only subtle differences in signalling via individual isoforms in vivo. Therefore, we hypothesise, at least in this model system, that relative tissue/cellular abundance, rather than specific activity, plays the dominant role in determining AKT substrate specificity in situ.

  14. Serum amyloid A isoforms in serum and synovial fluid from spontaneously diseased dogs with joint diseases or other conditions

    DEFF Research Database (Denmark)

    Kjelgaard-Hansen, Mads Jens; Christensen, Michelle B.; Lee, Marcel Huisung

    2007-01-01

    Serum amyloid A (SAA) is a major acute phase protein in dogs. However, knowledge of qualitative properties of canine SAA and extent of its synthesis in extrahepatic tissues is limited. The aim of the study was to investigate expression of different SAA isoforms in serum and synovial fluid...... in samples obtained from dogs (n = 16) suffering from different inflammatory or non-inflammatory conditions, which were either related or unrelated to joints. Expression of SAA isoforms was visualized by denaturing isoelectric focusing and Western blotting. Serum amyloid A was present in serum from all dogs...... with systemic inflammatory activity, and up to four major isoforms with apparent isoelectric points between 6.1 and 7.9 were identified. In synovial fluid from inflamed joints one or more highly alkaline SAA isoforms (with apparent isoelectric points above 9.3) were identified, with data suggesting local...

  15. EXAFS analysis of a human Cu,Zn SOD isoform focused using non-denaturing gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Chevreux, Sylviane; Roudeau, Stephane; Deves, Guillaume; Ortega, Richard [Laboratoire de Chimie Nucleaire Analytique et Bioenvironnementale, CNRS UMR5084, Universite Bordeaux 1, Chemin du Solarium, F-33175 Gradignan cedex (France); Solari, Pier Lorenzo [Synchrotron SOLEIL, L' Orme des Merisiers, BP 48, F-91192 Gif-sur-Yvette cedex, Saint-Aubin (France); Alliot, Isabelle; Testemale, Denis; Hazemann, Jean Louis, E-mail: ortega@cenbg.in2p3.f [FAME, ESRF, 6 rue Jules Horowitz, BP220, F-38043 Grenoble cedex (France)

    2009-11-15

    Isoelectric point isoforms of a metalloprotein, copper-zinc superoxide dismutase (CuZnSOD), separated on electrophoresis gels were analyzed using X-ray Absorption Spectroscopy. Mutations of this protein are involved in familial cases of amyotrophic lateral sclerosis. The toxicity of mutants could be relied to defects in the metallation state. Our purpose is to establish analytical protocols to study metallation state of protein isoforms such as those from CuZnSOD. We previously highlighted differences in the copper oxidation state between CuZnSOD isoforms using XANES. Here, we present the first results for EXAFS analyses performed at Cu and Zn K-edge on the majoritary expressed isoform of human CuZnSOD separated on electrophoresis gels.

  16. EXAFS analysis of a human Cu,Zn SOD isoform focused using non-denaturing gel electrophoresis

    Science.gov (United States)

    Chevreux, Sylviane; Solari, Pier Lorenzo; Roudeau, Stéphane; Deves, Guillaume; Alliot, Isabelle; Testemale, Denis; Hazemann, Jean Louis; Ortega, Richard

    2009-11-01

    Isoelectric point isoforms of a metalloprotein, copper-zinc superoxide dismutase (CuZnSOD), separated on electrophoresis gels were analyzed using X-ray Absorption Spectroscopy. Mutations of this protein are involved in familial cases of amyotrophic lateral sclerosis. The toxicity of mutants could be relied to defects in the metallation state. Our purpose is to establish analytical protocols to study metallation state of protein isoforms such as those from CuZnSOD. We previously highlighted differences in the copper oxidation state between CuZnSOD isoforms using XANES. Here, we present the first results for EXAFS analyses performed at Cu and Zn K-edge on the majoritary expressed isoform of human CuZnSOD separated on electrophoresis gels.

  17. EXAFS analysis of a human Cu,Zn SOD isoform focused using non-denaturing gel electrophoresis

    International Nuclear Information System (INIS)

    Chevreux, Sylviane; Roudeau, Stephane; Deves, Guillaume; Ortega, Richard; Solari, Pier Lorenzo; Alliot, Isabelle; Testemale, Denis; Hazemann, Jean Louis

    2009-01-01

    Isoelectric point isoforms of a metalloprotein, copper-zinc superoxide dismutase (CuZnSOD), separated on electrophoresis gels were analyzed using X-ray Absorption Spectroscopy. Mutations of this protein are involved in familial cases of amyotrophic lateral sclerosis. The toxicity of mutants could be relied to defects in the metallation state. Our purpose is to establish analytical protocols to study metallation state of protein isoforms such as those from CuZnSOD. We previously highlighted differences in the copper oxidation state between CuZnSOD isoforms using XANES. Here, we present the first results for EXAFS analyses performed at Cu and Zn K-edge on the majoritary expressed isoform of human CuZnSOD separated on electrophoresis gels.

  18. Does Increased Expression of the Plasma Membrane Calcium-ATPase Isoform 2 Confer Resistance to Apoptosis on Breast Cancer Cells?

    National Research Council Canada - National Science Library

    VanHouten, Joshua N

    2008-01-01

    The plasma membrane calcium ATPase isoform 2 (PMCA2) is highly expressed on the apical membrane of mammary epithelial cells during lactation, and is the predominant pump responsible for calcium transport into milk...

  19. Waste package reliability analysis

    International Nuclear Information System (INIS)

    Pescatore, C.; Sastre, C.

    1983-01-01

    Proof of future performance of a complex system such as a high-level nuclear waste package over a period of hundreds to thousands of years cannot be had in the ordinary sense of the word. The general method of probabilistic reliability analysis could provide an acceptable framework to identify, organize, and convey the information necessary to satisfy the criterion of reasonable assurance of waste package performance according to the regulatory requirements set forth in 10 CFR 60. General principles which may be used to evaluate the qualitative and quantitative reliability of a waste package design are indicated and illustrated with a sample calculation of a repository concept in basalt. 8 references, 1 table

  20. Accelerator reliability workshop

    Energy Technology Data Exchange (ETDEWEB)

    Hardy, L; Duru, Ph; Koch, J M; Revol, J L; Van Vaerenbergh, P; Volpe, A M; Clugnet, K; Dely, A; Goodhew, D

    2002-07-01

    About 80 experts attended this workshop, which brought together all accelerator communities: accelerator driven systems, X-ray sources, medical and industrial accelerators, spallation sources projects (American and European), nuclear physics, etc. With newly proposed accelerator applications such as nuclear waste transmutation, replacement of nuclear power plants and others. Reliability has now become a number one priority for accelerator designers. Every part of an accelerator facility from cryogenic systems to data storage via RF systems are concerned by reliability. This aspect is now taken into account in the design/budget phase, especially for projects whose goal is to reach no more than 10 interruptions per year. This document gathers the slides but not the proceedings of the workshop.

  1. Human Reliability Program Workshop

    Energy Technology Data Exchange (ETDEWEB)

    Landers, John; Rogers, Erin; Gerke, Gretchen

    2014-05-18

    A Human Reliability Program (HRP) is designed to protect national security as well as worker and public safety by continuously evaluating the reliability of those who have access to sensitive materials, facilities, and programs. Some elements of a site HRP include systematic (1) supervisory reviews, (2) medical and psychological assessments, (3) management evaluations, (4) personnel security reviews, and (4) training of HRP staff and critical positions. Over the years of implementing an HRP, the Department of Energy (DOE) has faced various challenges and overcome obstacles. During this 4-day activity, participants will examine programs that mitigate threats to nuclear security and the insider threat to include HRP, Nuclear Security Culture (NSC) Enhancement, and Employee Assistance Programs. The focus will be to develop an understanding of the need for a systematic HRP and to discuss challenges and best practices associated with mitigating the insider threat.

  2. Reliability and construction control

    Directory of Open Access Journals (Sweden)

    Sherif S. AbdelSalam

    2016-06-01

    Full Text Available The goal of this study was to determine the most reliable and efficient combination of design and construction methods required for vibro piles. For a wide range of static and dynamic formulas, the reliability-based resistance factors were calculated using EGYPT database, which houses load test results for 318 piles. The analysis was extended to introduce a construction control factor that determines the variation between the pile nominal capacities calculated using static versus dynamic formulae. From the major outcomes, the lowest coefficient of variation is associated with Davisson’s criterion, and the resistance factors calculated for the AASHTO method are relatively high compared with other methods. Additionally, the CPT-Nottingham and Schmertmann method provided the most economic design. Recommendations related to a pile construction control factor were also presented, and it was found that utilizing the factor can significantly reduce variations between calculated and actual capacities.

  3. Scyllac equipment reliability analysis

    International Nuclear Information System (INIS)

    Gutscher, W.D.; Johnson, K.J.

    1975-01-01

    Most of the failures in Scyllac can be related to crowbar trigger cable faults. A new cable has been designed, procured, and is currently undergoing evaluation. When the new cable has been proven, it will be worked into the system as quickly as possible without causing too much additional down time. The cable-tip problem may not be easy or even desirable to solve. A tightly fastened permanent connection that maximizes contact area would be more reliable than the plug-in type of connection in use now, but it would make system changes and repairs much more difficult. The balance of the failures have such a low occurrence rate that they do not cause much down time and no major effort is underway to eliminate them. Even though Scyllac was built as an experimental system and has many thousands of components, its reliability is very good. Because of this the experiment has been able to progress at a reasonable pace

  4. Improving Power Converter Reliability

    DEFF Research Database (Denmark)

    Ghimire, Pramod; de Vega, Angel Ruiz; Beczkowski, Szymon

    2014-01-01

    of a high-power IGBT module during converter operation, which may play a vital role in improving the reliability of the power converters. The measured voltage is used to estimate the module average junction temperature of the high and low-voltage side of a half-bridge IGBT separately in every fundamental......The real-time junction temperature monitoring of a high-power insulated-gate bipolar transistor (IGBT) module is important to increase the overall reliability of power converters for industrial applications. This article proposes a new method to measure the on-state collector?emitter voltage...... is measured in a wind power converter at a low fundamental frequency. To illustrate more, the test method as well as the performance of the measurement circuit are also presented. This measurement is also useful to indicate failure mechanisms such as bond wire lift-off and solder layer degradation...

  5. Accelerator reliability workshop

    International Nuclear Information System (INIS)

    Hardy, L.; Duru, Ph.; Koch, J.M.; Revol, J.L.; Van Vaerenbergh, P.; Volpe, A.M.; Clugnet, K.; Dely, A.; Goodhew, D.

    2002-01-01

    About 80 experts attended this workshop, which brought together all accelerator communities: accelerator driven systems, X-ray sources, medical and industrial accelerators, spallation sources projects (American and European), nuclear physics, etc. With newly proposed accelerator applications such as nuclear waste transmutation, replacement of nuclear power plants and others. Reliability has now become a number one priority for accelerator designers. Every part of an accelerator facility from cryogenic systems to data storage via RF systems are concerned by reliability. This aspect is now taken into account in the design/budget phase, especially for projects whose goal is to reach no more than 10 interruptions per year. This document gathers the slides but not the proceedings of the workshop

  6. Uncertainty Quantification in Numerical Aerodynamics

    KAUST Repository

    Litvinenko, Alexander

    2017-05-16

    We consider uncertainty quantification problem in aerodynamic simulations. We identify input uncertainties, classify them, suggest an appropriate statistical model and, finally, estimate propagation of these uncertainties into the solution (pressure, velocity and density fields as well as the lift and drag coefficients). The deterministic problem under consideration is a compressible transonic Reynolds-averaged Navier-Strokes flow around an airfoil with random/uncertain data. Input uncertainties include: uncertain angle of attack, the Mach number, random perturbations in the airfoil geometry, mesh, shock location, turbulence model and parameters of this turbulence model. This problem requires efficient numerical/statistical methods since it is computationally expensive, especially for the uncertainties caused by random geometry variations which involve a large number of variables. In numerical section we compares five methods, including quasi-Monte Carlo quadrature, polynomial chaos with coefficients determined by sparse quadrature and gradient-enhanced version of Kriging, radial basis functions and point collocation polynomial chaos, in their efficiency in estimating statistics of aerodynamic performance upon random perturbation to the airfoil geometry [D.Liu et al \\'17]. For modeling we used the TAU code, developed in DLR, Germany.

  7. Stochastic approach for radionuclides quantification

    Science.gov (United States)

    Clement, A.; Saurel, N.; Perrin, G.

    2018-01-01

    Gamma spectrometry is a passive non-destructive assay used to quantify radionuclides present in more or less complex objects. Basic methods using empirical calibration with a standard in order to quantify the activity of nuclear materials by determining the calibration coefficient are useless on non-reproducible, complex and single nuclear objects such as waste packages. Package specifications as composition or geometry change from one package to another and involve a high variability of objects. Current quantification process uses numerical modelling of the measured scene with few available data such as geometry or composition. These data are density, material, screen, geometric shape, matrix composition, matrix and source distribution. Some of them are strongly dependent on package data knowledge and operator backgrounds. The French Commissariat à l'Energie Atomique (CEA) is developing a new methodology to quantify nuclear materials in waste packages and waste drums without operator adjustment and internal package configuration knowledge. This method suggests combining a global stochastic approach which uses, among others, surrogate models available to simulate the gamma attenuation behaviour, a Bayesian approach which considers conditional probability densities of problem inputs, and Markov Chains Monte Carlo algorithms (MCMC) which solve inverse problems, with gamma ray emission radionuclide spectrum, and outside dimensions of interest objects. The methodology is testing to quantify actinide activity in different kind of matrix, composition, and configuration of sources standard in terms of actinide masses, locations and distributions. Activity uncertainties are taken into account by this adjustment methodology.

  8. Inverse Problems and Uncertainty Quantification

    KAUST Repository

    Litvinenko, Alexander

    2014-01-06

    In a Bayesian setting, inverse problems and uncertainty quantification (UQ) - the propagation of uncertainty through a computational (forward) modelare strongly connected. In the form of conditional expectation the Bayesian update becomes computationally attractive. This is especially the case as together with a functional or spectral approach for the forward UQ there is no need for time- consuming and slowly convergent Monte Carlo sampling. The developed sampling- free non-linear Bayesian update is derived from the variational problem associated with conditional expectation. This formulation in general calls for further discretisa- tion to make the computation possible, and we choose a polynomial approximation. After giving details on the actual computation in the framework of functional or spectral approximations, we demonstrate the workings of the algorithm on a number of examples of increasing complexity. At last, we compare the linear and quadratic Bayesian update on the small but taxing example of the chaotic Lorenz 84 model, where we experiment with the influence of different observation or measurement operators on the update.

  9. Inverse Problems and Uncertainty Quantification

    KAUST Repository

    Litvinenko, Alexander; Matthies, Hermann G.

    2014-01-01

    In a Bayesian setting, inverse problems and uncertainty quantification (UQ) - the propagation of uncertainty through a computational (forward) modelare strongly connected. In the form of conditional expectation the Bayesian update becomes computationally attractive. This is especially the case as together with a functional or spectral approach for the forward UQ there is no need for time- consuming and slowly convergent Monte Carlo sampling. The developed sampling- free non-linear Bayesian update is derived from the variational problem associated with conditional expectation. This formulation in general calls for further discretisa- tion to make the computation possible, and we choose a polynomial approximation. After giving details on the actual computation in the framework of functional or spectral approximations, we demonstrate the workings of the algorithm on a number of examples of increasing complexity. At last, we compare the linear and quadratic Bayesian update on the small but taxing example of the chaotic Lorenz 84 model, where we experiment with the influence of different observation or measurement operators on the update.

  10. Inverse problems and uncertainty quantification

    KAUST Repository

    Litvinenko, Alexander

    2013-12-18

    In a Bayesian setting, inverse problems and uncertainty quantification (UQ)— the propagation of uncertainty through a computational (forward) model—are strongly connected. In the form of conditional expectation the Bayesian update becomes computationally attractive. This is especially the case as together with a functional or spectral approach for the forward UQ there is no need for time- consuming and slowly convergent Monte Carlo sampling. The developed sampling- free non-linear Bayesian update is derived from the variational problem associated with conditional expectation. This formulation in general calls for further discretisa- tion to make the computation possible, and we choose a polynomial approximation. After giving details on the actual computation in the framework of functional or spectral approximations, we demonstrate the workings of the algorithm on a number of examples of increasing complexity. At last, we compare the linear and quadratic Bayesian update on the small but taxing example of the chaotic Lorenz 84 model, where we experiment with the influence of different observation or measurement operators on the update.

  11. Accurate and precise DNA quantification in the presence of different amplification efficiencies using an improved Cy0 method.

    Science.gov (United States)

    Guescini, Michele; Sisti, Davide; Rocchi, Marco B L; Panebianco, Renato; Tibollo, Pasquale; Stocchi, Vilberto

    2013-01-01

    Quantitative real-time PCR represents a highly sensitive and powerful technology for the quantification of DNA. Although real-time PCR is well accepted as the gold standard in nucleic acid quantification, there is a largely unexplored area of experimental conditions that limit the application of the Ct method. As an alternative, our research team has recently proposed the Cy0 method, which can compensate for small amplification variations among the samples being compared. However, when there is a marked decrease in amplification efficiency, the Cy0 is impaired, hence determining reaction efficiency is essential to achieve a reliable quantification. The proposed improvement in Cy0 is based on the use of the kinetic parameters calculated in the curve inflection point to compensate for efficiency variations. Three experimental models were used: inhibition of primer extension, non-optimal primer annealing and a very small biological sample. In all these models, the improved Cy0 method increased quantification accuracy up to about 500% without affecting precision. Furthermore, the stability of this procedure was enhanced integrating it with the SOD method. In short, the improved Cy0 method represents a simple yet powerful approach for reliable DNA quantification even in the presence of marked efficiency variations.

  12. Development of web-based reliability data base platform

    International Nuclear Information System (INIS)

    Hwang, Seok Won; Lee, Chang Ju; Sung, Key Yong

    2004-01-01

    Probabilistic safety assessment (PSA) is a systematic technique which estimates the degree of risk impacts to the public due to an accident scenario. Estimating the occurrence frequencies and consequences of potential scenarios requires a thorough analysis of the accident details and all fundamental parameters. The robustness of PSA to check weaknesses in a design and operation will allow a better informed and balanced decision to be reached. The fundamental parameters for PSA, such as the component failure rates, should be estimated under the condition of steady collection of the evidence throughout the operational period. However, since any single plant data does not sufficiently enough to provide an adequate PSA result, in actual, the whole operating data was commonly used to estimate the reliability parameters for the same type of components. The reliability data of any component type consists of two categories; the generic that is based on the operating experiences of whole plants, and the plant-specific that is based on the operation of a specific plant of interest. The generic data is highly essential for new or recently-built nuclear power plants (NPPs). Generally, the reliability data base may be categorized into the component reliability, initiating event frequencies, human performance, and so on. Among these data, the component reliability seems a key element because it has the most abundant population. Therefore, the component reliability data is essential for taking a part in the quantification of accident sequences because it becomes an input of various basic events which consists of the fault tree

  13. Reliability of Circumplex Axes

    Directory of Open Access Journals (Sweden)

    Micha Strack

    2013-06-01

    Full Text Available We present a confirmatory factor analysis (CFA procedure for computing the reliability of circumplex axes. The tau-equivalent CFA variance decomposition model estimates five variance components: general factor, axes, scale-specificity, block-specificity, and item-specificity. Only the axes variance component is used for reliability estimation. We apply the model to six circumplex types and 13 instruments assessing interpersonal and motivational constructs—Interpersonal Adjective List (IAL, Interpersonal Adjective Scales (revised; IAS-R, Inventory of Interpersonal Problems (IIP, Impact Messages Inventory (IMI, Circumplex Scales of Interpersonal Values (CSIV, Support Action Scale Circumplex (SAS-C, Interaction Problems With Animals (IPI-A, Team Role Circle (TRC, Competing Values Leadership Instrument (CV-LI, Love Styles, Organizational Culture Assessment Instrument (OCAI, Customer Orientation Circle (COC, and System for Multi-Level Observation of Groups (behavioral adjectives; SYMLOG—in 17 German-speaking samples (29 subsamples, grouped by self-report, other report, and metaperception assessments. The general factor accounted for a proportion ranging from 1% to 48% of the item variance, the axes component for 2% to 30%; and scale specificity for 1% to 28%, respectively. Reliability estimates varied considerably from .13 to .92. An application of the Nunnally and Bernstein formula proposed by Markey, Markey, and Tinsley overestimated axes reliabilities in cases of large-scale specificities but otherwise works effectively. Contemporary circumplex evaluations such as Tracey’s RANDALL are sensitive to the ratio of the axes and scale-specificity components. In contrast, the proposed model isolates both components.

  14. The cost of reliability

    International Nuclear Information System (INIS)

    Ilic, M.

    1998-01-01

    In this article the restructuring process under way in the US power industry is being revisited from the point of view of transmission system provision and reliability was rolled into the average cost of electricity to all, it is not so obvious how is this cost managed in the new industry. A new MIT approach to transmission pricing is here suggested as a possible solution [it

  15. Analysis of human bone alkaline phosphatase isoforms: comparison of isoelectric focusing and ion-exchange high-performance liquid chromatography.

    Science.gov (United States)

    Sharp, Christopher A; Linder, Cecilia; Magnusson, Per

    2007-04-01

    Several isoforms of alkaline phosphatase (ALP) can be identified in human tissues and serum after separation by anion-exchange HPLC and isoelectric focusing (IEF). We purified four soluble bone ALP (BALP) isoforms (B/I, B1x, B1 and B2) from human SaOS-2 cells, determined their specific pI values by broad range IEF (pH 3.5-9.5), compared these with commercial preparations of bone, intestinal and liver ALPs and established the effects of neuraminidase and wheat germ lectin (WGA) on enzyme activity. Whilst the isoforms B1x (pI=4.48), B1 (pI=4.32) and B2 (pI=4.12) resolved as well-defined bands, B/I resolved as a complex (pI=4.85-6.84). Neuraminidase altered the migration of all BALP isoforms to pI=6.84 and abolished their binding to the anion-exchange matrix, but increased their enzymatic activities by 11-20%. WGA precipitated the BALP isoforms in IEF gels and the HPLC column and attenuated their enzymatic activities by 54-73%. IEF resolved the commercial BALP into 2 major bands (pI=4.41 and 4.55). Migration of BALP isoforms is similar in IEF and anion-exchange HPLC and dependent on sialic acid content. HPLC is preferable in smaller scale research applications where samples containing mixtures of BALP isoforms are analysed. Circulating liver ALP (pI=3.85) can be resolved from BALP by either method. IEF represents a simpler approach for routine purposes even though some overlapping of the isoforms may occur.

  16. Human HDAC isoform selectivity achieved via exploitation of the acetate release channel with structurally unique small molecule inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Whitehead, Lewis; Dobler, Markus R.; Radetich, Branko; Zhu, Yanyi; Atadja, Peter W.; Claiborne, Tavina; Grob, Jonathan E.; McRiner, Andrew; Pancost, Margaret R.; Patnaik, Anup; Shao, Wenlin; Shultz, Michael; Tichkule, Ritesh; Tommasi, Ruben A.; Vash, Brian; Wang, Ping; Stams, Travis (Novartis)

    2013-11-20

    Herein we report the discovery of a family of novel yet simple, amino-acid derived class I HDAC inhibitors that demonstrate isoform selectivity via access to the internal acetate release channel. Isoform selectivity criteria is discussed on the basis of X-ray crystallography and molecular modeling of these novel inhibitors bound to HDAC8, potentially revealing insights into the mechanism of enzymatic function through novel structural features revealed at the atomic level.

  17. Lung involvement quantification in chest radiographs

    International Nuclear Information System (INIS)

    Giacomini, Guilherme; Alvarez, Matheus; Oliveira, Marcela de; Miranda, Jose Ricardo A.; Pina, Diana R.; Pereira, Paulo C.M.; Ribeiro, Sergio M.

    2014-01-01

    Tuberculosis (TB) caused by Mycobacterium tuberculosis, is an infectious disease which remains a global health problem. The chest radiography is the commonly method employed to assess the TB's evolution. The methods for quantification of abnormalities of chest are usually performed on CT scans (CT). This quantification is important to assess the TB evolution and treatment and comparing different treatments. However, precise quantification is not feasible for the amount of CT scans required. The purpose of this work is to develop a methodology for quantification of lung damage caused by TB through chest radiographs. It was developed an algorithm for computational processing of exams in Matlab, which creates a lungs' 3D representation, with compromised dilated regions inside. The quantification of lung lesions was also made for the same patients through CT scans. The measurements from the two methods were compared and resulting in strong correlation. Applying statistical Bland and Altman, all samples were within the limits of agreement, with a confidence interval of 95%. The results showed an average variation of around 13% between the two quantification methods. The results suggest the effectiveness and applicability of the method developed, providing better risk-benefit to the patient and cost-benefit ratio for the institution. (author)

  18. Software reliability studies

    Science.gov (United States)

    Hoppa, Mary Ann; Wilson, Larry W.

    1994-01-01

    There are many software reliability models which try to predict future performance of software based on data generated by the debugging process. Our research has shown that by improving the quality of the data one can greatly improve the predictions. We are working on methodologies which control some of the randomness inherent in the standard data generation processes in order to improve the accuracy of predictions. Our contribution is twofold in that we describe an experimental methodology using a data structure called the debugging graph and apply this methodology to assess the robustness of existing models. The debugging graph is used to analyze the effects of various fault recovery orders on the predictive accuracy of several well-known software reliability algorithms. We found that, along a particular debugging path in the graph, the predictive performance of different models can vary greatly. Similarly, just because a model 'fits' a given path's data well does not guarantee that the model would perform well on a different path. Further we observed bug interactions and noted their potential effects on the predictive process. We saw that not only do different faults fail at different rates, but that those rates can be affected by the particular debugging stage at which the rates are evaluated. Based on our experiment, we conjecture that the accuracy of a reliability prediction is affected by the fault recovery order as well as by fault interaction.

  19. Investment in new product reliability

    International Nuclear Information System (INIS)

    Murthy, D.N.P.; Rausand, M.; Virtanen, S.

    2009-01-01

    Product reliability is of great importance to both manufacturers and customers. Building reliability into a new product is costly, but the consequences of inadequate product reliability can be costlier. This implies that manufacturers need to decide on the optimal investment in new product reliability by achieving a suitable trade-off between the two costs. This paper develops a framework and proposes an approach to help manufacturers decide on the investment in new product reliability.

  20. A real-time PCR assay for detection and quantification of Verticillium dahliae in spinach seed.

    Science.gov (United States)

    Duressa, Dechassa; Rauscher, Gilda; Koike, Steven T; Mou, Beiquan; Hayes, Ryan J; Maruthachalam, Karunakaran; Subbarao, Krishna V; Klosterman, Steven J

    2012-04-01

    Verticillium dahliae is a soilborne fungus that causes Verticillium wilt on multiple crops in central coastal California. Although spinach crops grown in this region for fresh and processing commercial production do not display Verticillium wilt symptoms, spinach seeds produced in the United States or Europe are commonly infected with V. dahliae. Planting of the infected seed increases the soil inoculum density and may introduce exotic strains that contribute to Verticillium wilt epidemics on lettuce and other crops grown in rotation with spinach. A sensitive, rapid, and reliable method for quantification of V. dahliae in spinach seed may help identify highly infected lots, curtail their planting, and minimize the spread of exotic strains via spinach seed. In this study, a quantitative real-time polymerase chain reaction (qPCR) assay was optimized and employed for detection and quantification of V. dahliae in spinach germplasm and 15 commercial spinach seed lots. The assay used a previously reported V. dahliae-specific primer pair (VertBt-F and VertBt-R) and an analytical mill for grinding tough spinach seed for DNA extraction. The assay enabled reliable quantification of V. dahliae in spinach seed, with a sensitivity limit of ≈1 infected seed per 100 (1.3% infection in a seed lot). The quantification was highly reproducible between replicate samples of a seed lot and in different real-time PCR instruments. When tested on commercial seed lots, a pathogen DNA content corresponding to a quantification cycle value of ≥31 corresponded with a percent seed infection of ≤1.3%. The assay is useful in qualitatively assessing seed lots for V. dahliae infection levels, and the results of the assay can be helpful to guide decisions on whether to apply seed treatments.

  1. Immunopositivity for histone macroH2A1 isoforms marks steatosis-associated hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Francesca Rappa

    Full Text Available Hepatocellular carcinoma (HCC is one of the most common cancers worldwide. Prevention and risk reduction are important and the identification of specific biomarkers for early diagnosis of HCC represents an active field of research. Increasing evidence indicates that fat accumulation in the liver, defined as hepatosteatosis, is an independent and strong risk factor for developing an HCC. MacroH2A1, a histone protein generally associated with the repressed regions of chromosomes, is involved in hepatic lipid metabolism and is present in two alternative spliced isoforms, macroH2A1.1 and macroH2A1.2. These isoforms have been shown to predict lung and colon cancer recurrence but to our knowledge, their role in fatty-liver associated HCC has not been investigated previously.We examined macroH2A1.1 and macroH2A1.2 protein expression levels in the liver of two murine models of fat-associated HCC, the high fat diet/diethylnistrosamine (DEN and the phosphatase and tensin homolog (PTEN liver specific knock-out (KO mouse, and in human liver samples of subjects with steatosis or HCC, using immunoblotting and immunohistochemistry.Protein levels for both macroH2A1 isoforms were massively upregulated in HCC, whereas macroH2A1.2 was specifically upregulated in steatosis. In addition, examination of human liver samples showed a significant difference (p<0.01 in number of positive nuclei in HCC (100% of tumor cells positive for either macroH2A1.1 or macroH2A1.2, when compared to steatosis (<2% of hepatocytes positive for either isoform. The steatotic areas flanking the tumors were highly immunopositive for macroH2A1.1 and macroH2A1.2.These data obtained in mice and humans suggest that both macroH2A1 isoforms may play a role in HCC pathogenesis and moreover may be considered as novel diagnostic markers for human HCC.

  2. The effect of a period of intensive exercise on the isoform test to detect growth hormone doping in sports.

    Science.gov (United States)

    Voss, S C; Giraud, S; Alsayrafi, M; Bourdon, P C; Schumacher, Y O; Saugy, M; Robinson, N

    2013-08-01

    The major objective of this study was to investigate the effects of several days of intense exercise on growth hormone (hGH) testing using the World Anti-Doping Agencies hGH isoform differential immunoassays. Additionally the effects of circadian variation and exercise type on the isoform ratios were also investigated. 15 male athletes performed a simulated nine day cycling stage race. Blood samples were collected twice daily over a period of 15 days (stage race+three days before and after). hGH isoforms were analysed by the official WADA immunoassays (CMZ Assay GmbH). All measured isoform ratios were far below the WADA decision limits for an adverse analytical finding. Changes in the isoform ratios could not be clearly connected to circadian variation, exercise duration or intensity. The present study demonstrates that the hGH isoform ratios are not significantly affected by exercise or circadian variation. We demonstrated that heavy, long term exercise does not interfere with the decision limits for an adverse analytical finding. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Two-dimensional zymography differentiates gelatinase isoforms in stimulated microglial cells and in brain tissues of acute brain injuries.

    Science.gov (United States)

    Chen, Shanyan; Meng, Fanjun; Chen, Zhenzhou; Tomlinson, Brittany N; Wesley, Jennifer M; Sun, Grace Y; Whaley-Connell, Adam T; Sowers, James R; Cui, Jiankun; Gu, Zezong

    2015-01-01

    Excessive activation of gelatinases (MMP-2/-9) is a key cause of detrimental outcomes in neurodegenerative diseases. A single-dimension zymography has been widely used to determine gelatinase expression and activity, but this method is inadequate in resolving complex enzyme isoforms, because gelatinase expression and activity could be modified at transcriptional and posttranslational levels. In this study, we investigated gelatinase isoforms under in vitro and in vivo conditions using two-dimensional (2D) gelatin zymography electrophoresis, a protocol allowing separation of proteins based on isoelectric points (pI) and molecular weights. We observed organomercuric chemical 4-aminophenylmercuric acetate-induced activation of MMP-2 isoforms with variant pI values in the conditioned medium of human fibrosarcoma HT1080 cells. Studies with murine BV-2 microglial cells indicated a series of proform MMP-9 spots separated by variant pI values due to stimulation with lipopolysaccharide (LPS). The MMP-9 pI values were shifted after treatment with alkaline phosphatase, suggesting presence of phosphorylated isoforms due to the proinflammatory stimulation. Similar MMP-9 isoforms with variant pI values in the same molecular weight were also found in mouse brains after ischemic and traumatic brain injuries. In contrast, there was no detectable pI differentiation of MMP-9 in the brains of chronic Zucker obese rats. These results demonstrated effective use of 2D zymography to separate modified MMP isoforms with variant pI values and to detect posttranslational modifications under different pathological conditions.

  4. The Regulation of Nitric Oxide Synthase Isoform Expression in Mouse and Human Fallopian Tubes: Potential Insights for Ectopic Pregnancy

    Directory of Open Access Journals (Sweden)

    Junting Hu

    2014-12-01

    Full Text Available Nitric oxide (NO is highly unstable and has a half-life of seconds in buffer solutions. It is synthesized by NO-synthase (NOS, which has been found to exist in the following three isoforms: neuro nitric oxide synthase (nNOS, inducible nitric oxide synthase (iNOS, and endothelial nitric oxide synthase (eNOS. NOS activity is localized in the reproductive tracts of many species, although direct evidence for NOS isoforms in the Fallopian tubes of mice is still lacking. In the present study, we investigated the expression and regulation of NOS isoforms in the mouse and human Fallopian tubes during the estrous and menstrual cycles, respectively. We also measured isoform expression in humans with ectopic pregnancy and in mice treated with lipopolysaccharide (LPS. Our results confirmed the presence of different NOS isoforms in the mouse and human Fallopian tubes during different stages of the estrous and menstrual cycles and showed that iNOS expression increased in the Fallopian tubes of women with ectopic pregnancy and in LPS-treated mice. Elevated iNOS activity might influence ovulation, cilia beats, contractility, and embryo transportation in such a manner as to increase the risk of ectopic pregnancy. This study has provided morphological and molecular evidence that NOS isoforms are present and active in the human and mouse Fallopian tubes and suggests that iNOS might play an important role in both the reproductive cycle and infection-induced ectopic pregnancies.

  5. The schizophrenia-associated Kv11.1-3.1 isoform results in reduced current accumulation during repetitive brief depolarizations.

    Directory of Open Access Journals (Sweden)

    Juliane Heide

    Full Text Available Recent genome wide association studies identified a brain and primate specific isoform of a voltage-gated potassium channel, referred to as Kv11.1-3.1, which is significantly associated with schizophrenia. The 3.1 isoform replaces the first 102 amino acids of the most abundant isoform (referred to as Kv11.1-1A with six unique amino acids. Here we show that the Kv11.1-3.1 isoform has faster rates of channel deactivation but a slowing of the rates of inactivation compared to the Kv11.1-1A isoform. The Kv11.1-3.1 isoform also has a significant depolarizing shift in the voltage-dependence of steady-state inactivation. The consequence of the altered gating kinetics is that there is lower current accumulation for Kv11.1-3.1 expressing cells during repetitive action potential firing compared to Kv11.1-1A expressing cells, which in turn will result in longer lasting trains of action potentials. Increased expression of Kv11.1-3.1 channels in the brain of schizophrenia patients might therefore contribute to disorganized neuronal firing.

  6. Plasmodium falciparum chloroquine resistance transporter (PfCRT) isoforms PH1 and PH2 perturb vacuolar physiology.

    Science.gov (United States)

    Callaghan, Paul S; Siriwardana, Amila; Hassett, Matthew R; Roepe, Paul D

    2016-03-31

    Recent work has perfected yeast-based methods for measuring drug transport by the Plasmodium falciparum chloroquine (CQ) resistance transporter (PfCRT). The approach relies on inducible heterologous expression of PfCRT in Saccharomyces cerevisiae yeast. In these experiments selecting drug concentrations are not toxic to the yeast, nor is expression of PfCRT alone toxic. Only when PfCRT is expressed in the presence of CQ is the growth of yeast impaired, due to inward transport of chloroquine (CQ) via the transporter. During analysis of all 53 known naturally occurring PfCRT isoforms, two isoforms (PH1 and PH2 PfCRT) were found to be intrinsically toxic to yeast, even in the absence of CQ. Additional analysis of six very recently identified PfCRT isoforms from Malaysia also showed some toxicity. In this paper the nature of this yeast toxicity is examined. Data also show that PH1 and PH2 isoforms of PfCRT transport CQ with an efficiency intermediate to that catalyzed by previously studied CQR conferring isoforms. Mutation of PfCRT at position 160 is found to perturb vacuolar physiology, suggesting a fitness cost to position 160 amino acid substitutions. These data further define the wide range of activities that exist for PfCRT isoforms found in P. falciparum isolates from around the globe.

  7. VEGF-A isoform-specific regulation of calcium ion flux, transcriptional activation and endothelial cell migration.

    Science.gov (United States)

    Fearnley, Gareth W; Bruns, Alexander F; Wheatcroft, Stephen B; Ponnambalam, Sreenivasan

    2015-04-24

    Vascular endothelial growth factor A (VEGF-A) regulates many aspects of vascular physiology such as cell migration, proliferation, tubulogenesis and cell-cell interactions. Numerous isoforms of VEGF-A exist but their physiological significance is unclear. Here we evaluated two different VEGF-A isoforms and discovered differential regulation of cytosolic calcium ion flux, transcription factor localisation and endothelial cell response. Analysis of VEGF-A isoform-specific stimulation of VEGFR2-dependent signal transduction revealed differential capabilities for isoform activation of multiple signal transduction pathways. VEGF-A165 treatment promoted increased phospholipase Cγ1 phosphorylation, which was proportional to the subsequent rise in cytosolic calcium ions, in comparison to cells treated with VEGF-A121. A major consequence of this VEGF-A isoform-specific calcium ion flux in endothelial cells is differential dephosphorylation and subsequent nuclear translocation of the transcription factor NFATc2. Using reverse genetics, we discovered that NFATc2 is functionally required for VEGF-A-stimulated endothelial cell migration but not tubulogenesis. This work presents a new mechanism for understanding how VEGF-A isoforms program complex cellular outputs by converting signal transduction pathways into transcription factor redistribution to the nucleus, as well as defining a novel role for NFATc2 in regulating the endothelial cell response. © 2015. Published by The Company of Biologists Ltd.

  8. VEGF-A isoform-specific regulation of calcium ion flux, transcriptional activation and endothelial cell migration

    Directory of Open Access Journals (Sweden)

    Gareth W. Fearnley

    2015-07-01

    Full Text Available Vascular endothelial growth factor A (VEGF-A regulates many aspects of vascular physiology such as cell migration, proliferation, tubulogenesis and cell-cell interactions. Numerous isoforms of VEGF-A exist but their physiological significance is unclear. Here we evaluated two different VEGF-A isoforms and discovered differential regulation of cytosolic calcium ion flux, transcription factor localisation and endothelial cell response. Analysis of VEGF-A isoform-specific stimulation of VEGFR2-dependent signal transduction revealed differential capabilities for isoform activation of multiple signal transduction pathways. VEGF-A165 treatment promoted increased phospholipase Cγ1 phosphorylation, which was proportional to the subsequent rise in cytosolic calcium ions, in comparison to cells treated with VEGF-A121. A major consequence of this VEGF-A isoform-specific calcium ion flux in endothelial cells is differential dephosphorylation and subsequent nuclear translocation of the transcription factor NFATc2. Using reverse genetics, we discovered that NFATc2 is functionally required for VEGF-A-stimulated endothelial cell migration but not tubulogenesis. This work presents a new mechanism for understanding how VEGF-A isoforms program complex cellular outputs by converting signal transduction pathways into transcription factor redistribution to the nucleus, as well as defining a novel role for NFATc2 in regulating the endothelial cell response.

  9. 'Motion frozen' quantification and display of myocardial perfusion gated SPECT

    International Nuclear Information System (INIS)

    Slomka, P.J.; Hurwitz, G.A.; Baddredine, M.; Baranowski, J.; Aladl, U.E.

    2002-01-01

    Aim: Gated SPECT imaging incorporates both functional and perfusion information of the left ventricle (LV). However perfusion data is confounded by the effect of ventricular motion. Most existing quantification paradigms simply add all gated frames and then proceed to extract the perfusion information from static images, discarding the effects of cardiac motion. In an attempt to improve the reliability and accuracy of cardiac SPECT quantification we propose to eliminate the LV motion prior to the perfusion quantification via automated image warping algorithm. Methods: A pilot series of 14 male and 11 female gated stress SPECT images acquired with 8 time bins have been co-registered to the coordinates of the 3D normal templates. Subsequently the LV endo and epi-cardial 3D points (300-500) were identified on end-systolic (ES) and end-diastolic (ED) frames, defining the ES-ED motion vectors. The nonlinear image warping algorithm (thin-plate-spline) was then applied to warp end-systolic frame was onto the end-diastolic frames using the corresponding ES-ED motion vectors. The remaining 6 intermediate frames were also transformed to the ED coordinates using fractions of the motion vectors. Such warped images were then summed to provide the LV perfusion image in the ED phase but with counts from the full cycle. Results: The identification of the ED/ES corresponding points was successful in all cases. The corrected displacement between ED and ES images was up to 25 mm. The summed images had the appearance of the ED frames but have been much less noisy since all the counts have been used. The spatial resolution of such images appeared higher than that of summed gated images, especially in the female scans. These 'motion frozen' images could be displayed and quantified as regular non-gated tomograms including polar map paradigm. Conclusions: This image processing technique may improve the effective image resolution of summed gated myocardial perfusion images used for

  10. Hepatic Iron Quantification on 3 Tesla (3 T Magnetic Resonance (MR: Technical Challenges and Solutions

    Directory of Open Access Journals (Sweden)

    Muhammad Anwar

    2013-01-01

    Full Text Available MR has become a reliable and noninvasive method of hepatic iron quantification. Currently, most of the hepatic iron quantification is performed on 1.5 T MR, and the biopsy measurements have been paired with R2 and R2* values for 1.5 T MR. As the use of 3 T MR scanners is steadily increasing in clinical practice, it has become important to evaluate the practicality of calculating iron burden at 3 T MR. Hepatic iron quantification on 3 T MR requires a better understanding of the process and more stringent technical considerations. The purpose of this work is to focus on the technical challenges in establishing a relationship between T2* values at 1.5 T MR and 3 T MR for hepatic iron concentration (HIC and to develop an appropriately optimized MR protocol for the evaluation of T2* values in the liver at 3 T magnetic field strength. We studied 22 sickle cell patients using multiecho fast gradient-echo sequence (MFGRE 3 T MR and compared the results with serum ferritin and liver biopsy results. Our study showed that the quantification of hepatic iron on 3 T MRI in sickle cell disease patients correlates well with clinical blood test results and biopsy results. 3 T MR liver iron quantification based on MFGRE can be used for hepatic iron quantification in transfused patients.

  11. Quantification of endogenous and exogenous protein expressions of Na,K-ATPase with super-resolution PALM/STORM imaging.

    Science.gov (United States)

    Bernhem, Kristoffer; Blom, Hans; Brismar, Hjalmar

    2018-01-01

    Transient transfection of fluorescent fusion proteins is a key enabling technology in fluorescent microscopy to spatio-temporally map cellular protein distributions. Transient transfection of proteins may however bypass normal regulation of expression, leading to overexpression artefacts like misallocations and excess amounts. In this study we investigate the use of STORM and PALM microscopy to quantitatively monitor endogenous and exogenous protein expression. Through incorporation of an N-terminal hemagglutinin epitope to a mMaple3 fused Na,K-ATPase (α1 isoform), we analyze the spatial and quantitative changes of plasma membrane Na,K-ATPase localization during competitive transient expression. Quantification of plasma membrane protein density revealed a time dependent increase of Na,K-ATPase, but no increase in size of protein clusters. Results show that after 41h transfection, the total plasma membrane density of Na,K-ATPase increased by 63% while the endogenous contribution was reduced by 16%.

  12. A phase quantification method based on EBSD data for a continuously cooled microalloyed steel

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, H.; Wynne, B.P.; Palmiere, E.J., E-mail: e.j.palmiere@sheffield.ac.uk

    2017-01-15

    Mechanical properties of steels depend on the phase constitutions of the final microstructures which can be related to the processing parameters. Therefore, accurate quantification of different phases is necessary to investigate the relationships between processing parameters, final microstructures and mechanical properties. Point counting on micrographs observed by optical or scanning electron microscopy is widely used as a phase quantification method, and different phases are discriminated according to their morphological characteristics. However, it is difficult to differentiate some of the phase constituents with similar morphology. Differently, for EBSD based phase quantification methods, besides morphological characteristics, other parameters derived from the orientation information can also be used for discrimination. In this research, a phase quantification method based on EBSD data in the unit of grains was proposed to identify and quantify the complex phase constitutions of a microalloyed steel subjected to accelerated coolings. Characteristics of polygonal ferrite/quasi-polygonal ferrite, acicular ferrite and bainitic ferrite on grain averaged misorientation angles, aspect ratios, high angle grain boundary fractions and grain sizes were analysed and used to develop the identification criteria for each phase. Comparing the results obtained by this EBSD based method and point counting, it was found that this EBSD based method can provide accurate and reliable phase quantification results for microstructures with relatively slow cooling rates. - Highlights: •A phase quantification method based on EBSD data in the unit of grains was proposed. •The critical grain area above which GAM angles are valid parameters was obtained. •Grain size and grain boundary misorientation were used to identify acicular ferrite. •High cooling rates deteriorate the accuracy of this EBSD based method.

  13. Review of Reliability-Based Design Optimization Approach and Its Integration with Bayesian Method

    Science.gov (United States)

    Zhang, Xiangnan

    2018-03-01

    A lot of uncertain factors lie in practical engineering, such as external load environment, material property, geometrical shape, initial condition, boundary condition, etc. Reliability method measures the structural safety condition and determine the optimal design parameter combination based on the probabilistic theory. Reliability-based design optimization (RBDO) is the most commonly used approach to minimize the structural cost or other performance under uncertainty variables which combines the reliability theory and optimization. However, it cannot handle the various incomplete information. The Bayesian approach is utilized to incorporate this kind of incomplete information in its uncertainty quantification. In this paper, the RBDO approach and its integration with Bayesian method are introduced.

  14. The National Centre of Systems Reliability and some aspects of its future activities

    International Nuclear Information System (INIS)

    Bourne, A.J.

    1975-01-01

    The National Centre of Systems Reliability (NCSR) has been set up to enhance the work of the Systems Reliability Service (SRS), which during its four years of operation by the UKAEA has offered to industry expertise in the quantification of reliability of systems in various technological applications. An outline is presented of the background to the establishment of the NCSR, including a brief summary of the work of the SRS. Certain aspects of the future activities of the NCSR particularly in relation to research and collaboration with universities are discussed. (U.K.)

  15. Eff ect of vitamin E isoforms on the primary intention Eff ect of vitamin E isoforms on the primary intention skin wound healing of diabetic rats

    Directory of Open Access Journals (Sweden)

    Bijo Elsy

    2017-10-01

    Full Text Available Introduction: Impaired wound healing events is a common complication in diabetes. One of the effective nutritional antioxidant on skin wound healing is vitamin E which contains saturated tocopherol and unsaturated tocotrienol forms. This present study is designed to explore the effect of different vitamin E isoforms on stitched skin wound in both healthy and diabetic rats. Materials and Methods: Forty eight albino rats were divided into eight groups; healthy control, diabetic control, healthy treated (d-α-tocopherol, d-δ-TRF and co-administrated and diabetic treated (d-αtocopherol, d-δ-TRF and co-administrated. Diabetes was induced through single subcutaneous injection of alloxan at the dose of 100 mg/kg. Treated groups were administered d-a-tocopherol (200 mg/kg, d-δ-TRF (200 mg/kg and co-administration (100 mg/kg of these two compounds each orally and daily for three weeks. A horizontal skin incision was made on right mid-thigh region at 2.95 ± 0.17cm in length and wound was closed with an absorbable suture. Results: Histopathological and histomorphological results at the end of 3rd week revealed that the d-δ-TRF treated groups promote the regeneration and reorganization of epidermal and dermal components in healing of primary intention more effectively than the d-α-tocopherol and co-administrated groups. Conclusion: It is concluded that among different vitamin E isoforms the d-δ-TRF appears to be a more effective nutritional antioxidant on skin wound healing in both healthy and diabetics.

  16. Expression and Immunohistochemical Localisation of the G beta gamma activated and Calcineurin-inhibited Adenylyl Cyclase Isoforms in Rat Articular Chondrocytes

    International Nuclear Information System (INIS)

    Memon, I.; Khan, K.M.; Siddiqui, S.; Perveen, S.; Ishaq, M.

    2016-01-01

    Objective: To determine the expression and localisation of the Gβγ-activated adenylyl cyclase (AC) isoforms 2, 4, and 7 and calcineurin-inhibited AC isoform 9 in rat articular chondrocytes. Study Design: Experimental study. Place and Duration of Study: Jumma Research Laboratory and Histology Laboratory, The Aga Khan University, Karachi, from 2009 to 2011. Methodology: Fresh slices of articular cartilage were taken from various synovial joints of rats of different age groups. The expression of AC isoforms was determined by RT-PCR and immunohistochemistry was performed to localise these isoforms in articular chondrocytes. Tissue sections were processed for immunostaining with respective antibodies. The color was developed by diaminobenzidine. Results: All the studied AC isoforms were found to be differentially expressed in different zones of the rat articular cartilage. Generally, expression of all AC isoforms studied increased with age. The expression of the AC isoforms through PCR was almost consistent with the localisation of these isoforms by immunohistochemistry. Conclusion: These data add to the information about signalling cascades possibly involved in articular chondrocytes. Variable expression of AC isoforms 2, 4, 7, and 9 suggest a role for the signalling cascades regulated by the AC isoforms in articular chondrocytes. (author)

  17. Nuclear performance and reliability

    International Nuclear Information System (INIS)

    Rothwell, G.

    1993-01-01

    If fewer forced outages are a sign of improved safety, nuclear power plants have become safer and more productive. There has been a significant improvement in nuclear power plant performance, due largely to a decline in the forced outage rate and a dramatic drop in the average number of forced outages per fuel cycle. If fewer forced outages are a sign of improved safety, nuclear power plants have become safer and more productive over time. To encourage further increases in performance, regulatory incentive schemes should reward reactor operators for improved reliability and safety, as well as for improved performance

  18. [How Reliable is Neuronavigation?].

    Science.gov (United States)

    Stieglitz, Lennart Henning

    2016-02-17

    Neuronavigation plays a central role in modern neurosurgery. It allows visualizing instruments and three-dimensional image data intraoperatively and supports spatial orientation. Thus it allows to reduce surgical risks and speed up complex surgical procedures. The growing availability and importance of neuronavigation makes clear how relevant it is to know about its reliability and accuracy. Different factors may influence the accuracy during the surgery unnoticed, misleading the surgeon. Besides the best possible optimization of the systems themselves, a good knowledge about its weaknesses is mandatory for every neurosurgeon.

  19. The value of reliability

    DEFF Research Database (Denmark)

    Fosgerau, Mogens; Karlström, Anders

    2010-01-01

    We derive the value of reliability in the scheduling of an activity of random duration, such as travel under congested conditions. Using a simple formulation of scheduling utility, we show that the maximal expected utility is linear in the mean and standard deviation of trip duration, regardless...... of the form of the standardised distribution of trip durations. This insight provides a unification of the scheduling model and models that include the standard deviation of trip duration directly as an argument in the cost or utility function. The results generalise approximately to the case where the mean...

  20. CD44 isoforms are heterogeneously expressed in breast cancer and correlate with tumor subtypes and cancer stem cell markers

    International Nuclear Information System (INIS)

    Olsson, Eleonor; Lövgren, Kristina; Fernö, Mårten; Grabau, Dorthe; Borg, Åke; Hegardt, Cecilia; Honeth, Gabriella; Bendahl, Pär-Ola; Saal, Lao H; Gruvberger-Saal, Sofia; Ringnér, Markus; Vallon-Christersson, Johan; Jönsson, Göran; Holm, Karolina

    2011-01-01

    The CD44 cell adhesion molecule is aberrantly expressed in many breast tumors and has been implicated in the metastatic process as well as in the putative cancer stem cell (CSC) compartment. We aimed to investigate potential associations between alternatively spliced isoforms of CD44 and CSCs as well as to various breast cancer biomarkers and molecular subtypes. We used q-RT-PCR and exon-exon spanning assays to analyze the expression of four alternatively spliced CD44 isoforms as well as the total expression of CD44 in 187 breast tumors and 13 cell lines. ALDH1 protein expression was determined by IHC on TMA. Breast cancer cell lines showed a heterogeneous expression pattern of the CD44 isoforms, which shifted considerably when cells were grown as mammospheres. Tumors characterized as positive for the CD44 + /CD24 - phenotype by immunohistochemistry were associated to all isoforms except the CD44 standard (CD44S) isoform, which lacks all variant exons. Conversely, tumors with strong expression of the CSC marker ALDH1 had elevated expression of CD44S. A high expression of the CD44v2-v10 isoform, which retain all variant exons, was correlated to positive steroid receptor status, low proliferation and luminal A subtype. The CD44v3-v10 isoform showed similar correlations, while high expression of CD44v8-v10 was correlated to positive EGFR, negative/low HER2 status and basal-like subtype. High expression of CD44S was associated with strong HER2 staining and also a subgroup of basal-like tumors. Unsupervised hierarchical cluster analysis of CD44 isoform expression data divided tumors into four main clusters, which showed significant correlations to molecular subtypes and differences in 10-year overall survival. We demonstrate that individual CD44 isoforms can be associated to different breast cancer subtypes and clinical markers such as HER2, ER and PgR, which suggests involvement of CD44 splice variants in specific oncogenic signaling pathways. Efforts to link CD44 to