Expression profiling and cross-species RNA interference (RNAi) of desiccation-induced transcripts in the anhydrobiotic nematode Aphelenchus avenae

Abstract Background Some organisms can survive extreme desiccation by entering a state of suspended animation known as anhydrobiosis. The free-living mycophagous nematode Aphelenchus avenae can be induced to enter anhydrobiosis by pre-exposure to moderate reductions in relative humidity (RH) prior to extreme desiccation. This preconditioning phase is thought to allow modification of the transcriptome by activation of genes required for desiccation tolerance. Results To identify such genes, a panel of expressed sequence tags (ESTs) enriched for sequences upregulated in A. avenae during preconditioning was created. A subset of 30 genes with significant matches in databases, together with a number of apparently novel sequences, were chosen for further study. Several of the recognisable genes are associated with water stress, encoding, for example, two new hydrophilic proteins related to the late embryogenesis abundant (LEA) protein family. Expression studies confirmed EST panel members to be upregulated by evaporative water loss, and the majority of genes was also induced by osmotic stress and cold, but rather fewer by heat. We attempted to use RNA interference (RNAi) to demonstrate the importance of this gene set for anhydrobiosis, but found A. avenae to be recalcitrant with the techniques used. Instead, therefore, we developed a cross-species RNAi procedure using A. avenae sequences in another anhydrobiotic nematode, Panagrolaimus superbus, which is amenable to gene silencing. Of 20 A. avenae ESTs screened, a significant reduction in survival of desiccation in treated P. superbus populations was observed with two sequences, one of which was novel, while the other encoded a glutathione peroxidase. To confirm a role for glutathione peroxidases in anhydrobiosis, RNAi with cognate sequences from P. superbus was performed and was also shown to reduce desiccation tolerance in this species. Conclusions This study has identified and characterised the expression profiles of members of the anhydrobiotic gene set in A. avenae. It also demonstrates the potential of RNAi for the analysis of anhydrobiosis and provides the first genetic data to underline the importance of effective antioxidant systems in metazoan desiccation tolerance.

Transcriptional profiling of Saccharomyces cerevisiae exposed to propolis

Abstract Background Propolis is a natural product of plant resins collected by honeybees (Apis mellifera) from various plant sources. Our previous studies indicated that propolis sensitivity is dependent on the mitochondrial function and that vacuolar acidification and autophagy are important for yeast cell death caused by propolis. Here, we extended our understanding of propolis-mediated cell death in the yeast Saccharomyces cerevisiae by applying systems biology tools to analyze the transcriptional profiling of cells exposed to propolis. Methods We have used transcriptional profiling of S. cerevisiae exposed to propolis. We validated our findings by using real-time PCR of selected genes. Systems biology tools (physical protein-protein interaction [PPPI] network) were applied to analyse the propolis-induced transcriptional bevavior, aiming to identify which pathways are modulated by propolis in S. cerevisiae and potentially influencing cell death. Results We were able to observe 1,339 genes modulated in at least one time point when compared to the reference time (propolis untreated samples) (t-test, p-value 0.01). Enrichment analysis performed by Gene Ontology (GO) Term finder tool showed enrichment for several biological categories among the genes up-regulated in the microarray hybridization such as transport and transmembrane transport and response to stress. Real-time RT-PCR analysis of selected genes showed by our microarray hybridization approach was capable of providing information about S. cerevisiae gene expression modulation with a considerably high level of confidence. Finally, a physical protein-protein (PPPI) network design and global topological analysis stressed the importance of these pathways in response of S. cerevisiae to propolis and were correlated with the transcriptional data obtained thorough the microarray analysis. Conclusions In summary, our data indicate that propolis is largely affecting several pathways in the eukaryotic cell. However, the most prominent pathways are related to oxidative stress, mitochondrial electron transport chain, vacuolar acidification, regulation of macroautophagy associated with protein target to vacuole, cellular response to starvation, and negative regulation of transcription from RNA polymerase II promoter. Our work emphasizes again the importance of S. cerevisiae as a model system to understand at molecular level the mechanism whereby propolis causes cell death in this organism at the concentration herein tested. Our study is the first one that investigates systematically by using functional genomics how propolis influences and modulates the mRNA abundance of an organism and may stimulate further work on the propolis-mediated cell death mechanisms in fungi.

Morphological alterations and gene and protein expression profiling of bladder tumor cells after treatment with gemcitabine.

Chemical agents used in cancer therapy are associated with cell cycle arrest, activation or deactivation of mechanisms associated to DNA repair and apoptosis. However, due to the complexity of biological systems, the molecular mechanisms responsible for these activities are not fully understood. Thus, studies about gene and protein expression have shown promising results for understanding the mechanisms related to cellular responses and regression of cancer after chemotherapy. This study aimed to evaluate the gene and protein expression profiling in bladder transitional cell carcinoma (TCC) with different TP53 status after gemcitabine (1.56 μM) treatment. The RT4 (grade 1, TP53 wild type), 5637 (grade 2, TP53 mutated) and T24 (grade 3, TP53 mutated) cell lines were used. PCR arrays and mass spectrometry were used to analyze gene and protein expression, respectively. Morphological alterations were observed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The results of PCR array showed that gemcitabine activity was mainly related to CDKN1A, GADD45A and SERTDA1 overexpression, and BAX overexpression only in the wild type TP53 cells. Mass spectrometry demonstrated that gemcitabine modulated the protein expression, especially those from genes related to apoptosis, transport of vesicles and stress response. Analyses using SEM and TEM showed changes in cell morphology independently on the cell line studied. The observed decreased number of microvillus suggests low contact among the cells and between cell and extracellular matrix; irregular forms might indicate actin cytoskeleton deregulation; and the reduction in the amount of organelles and core size might indicate reduced cellular metabolism. In conclusion, independently on TP53 status or grade of bladder tumor, gemcitabine modulated genes related to the cell cycle and apoptosis, that reflected in morphological changes indicative of future cell death.

New molecular approaches to control rhabdomyosarcoma onset and metastasis in preclinical systems

Rhabdomyosarcoma is the most common soft tissue sarcoma of childhood. The aim of this study was to identify molecular events involved in rhabdomyosarcoma onset for the development of new therapeutic approaches against specific molecular targets. BALB-p53neu mice develop pelvic rhabdomyosarcoma and combines the activation of HER-2/neu oncogene with the inactivation of an allele of p53 oncosuppressor gene. Gene expression profiling led to the identification of genes potentially involved in rhabdomyosarcoma genesis and therefore of candidate targets. The pattern of expression of p53, HER-2/neu, CDKN2A/p19ARF and IGF-2 suggested that these alterations might be involved in gender-, site- and strain-specific development of rhabdomyosarcoma. Other genes such as CDKN1A/p21 might be involved. The role of IGF-2, CDKN2A/p19ARF and CDKN1A/p21 in tumor growth was investigated with siRNA in murine rhabdomyosarcoma cells. Silencing of p19ARF and p21 induced inhibition of growth and of migration ability, indicating a possible pro-tumor and pro-metastatic role in rhabdomyosarcoma in absence of p53. In addition the autocrine IGF-2/IGF-1R loop found in early phases of cancer progression strengthens its key role in sustaining rhabdomyosarcoma growth. As rhabdomyosarcoma displays defective myogenic differentiation, a therapeutic approach aimed at enhancing myogenic differentiation of rhabdomyosarcoma cells. Forced expression of myogenin was able to restore myogenic differentiation, significantly reduced cell motility and impaired tumor growth and metastatic spread. IL-4 treatment increased rhabdomyosarcoma cell growth, decreased myogenin expression and promoted migration of cells lacking myogenin. Another approach was based on small kinase inhibitors. Agents specifically targeting members of the HER family (Lapatinib), of the IGF system (NVP-AEW541) or downstream signal transducers (NVP-BEZ235) were investigated in vitro in human rhabdomyosarcoma cell lines as therapeutic anti-tumor and anti-metastatic tools. The major effects were obtained with NVP-BEZ235 treatment that was able to strongly inhibit cell growth in vitro and showed anti-metastatic effects in vivo.

Digital Systems for Open Access to Formal and Informal Learning

Today, Digital Systems and Services for Technology Supported Learning and Education are recognized as the key drivers to transform the way that individuals, groups and organizations “learn” and the way to “assess learning” in 21st Century. These transformations influence: Objectives - moving from acquiring new “knowledge” to developing new and relevant “competences”; Methods – moving from “classroom” based teaching to “context-aware” personalized learning; and Assessment – moving from “life-long” degrees and certifications to “on-demand” and “in-context” accreditation of qualifications. Within this context, promoting Open Access to Formal and Informal Learning, is currently a key issue in the public discourse and the global dialogue on Education, including Massive Open Online Courses (MOOCs) and Flipped School Classrooms. This volume on Digital Systems for Open Access to Formal and Informal Learning contributes to the international dialogue between researchers, technologists, practitioners and policy makers in Technology Supported Education and Learning. It addresses emerging issues related with both theory and practice, as well as, methods and technologies that can support Open Access to Formal and Informal Learning. In the twenty chapters contributed by international experts who are actively shaping the future of Educational Technology around the world, topics such as: - The evolution of University Open Courses in Transforming Learning - Supporting Open Access to Teaching and Learning of People with Disabilities - Assessing Student Learning in Online Courses - Digital Game-based Learning for School Education - Open Access to Virtual and Remote Labs for STEM Education - Teachers’ and Schools’ ICT Competence Profiling - Web-Based Education and Innovative Leadership in a K-12 International School Setting are presented. An in-depth blueprint of the promise, potential, and imminent future of the field, Digital Systems for Open Access to Formal and Informal Learning is necessary reading for researchers and practitioners, as well as, undergraduate and postgraduate students, in educational technology.

Profiling invasiveness in head and neck cancer: recent contributions of genomic and transcriptomic approaches.

High-throughput molecular profiling approaches have emerged as precious research tools in the field of head and neck translational oncology. Such approaches have identified and/or confirmed the role of several genes or pathways in the acquisition/maintenance of an invasive phenotype and the execution of cellular programs related to cell invasion. Recently published new-generation sequencing studies in head and neck squamous cell carcinoma (HNSCC) have unveiled prominent roles in carcinogenesis and cell invasion of mutations involving NOTCH1 and PI3K-patwhay components. Gene-expression profiling studies combined with systems biology approaches have allowed identifying and gaining further mechanistic understanding into pathways commonly enriched in invasive HNSCC. These pathways include antigen-presenting and leucocyte adhesion molecules, as well as genes involved in cell-extracellular matrix interactions. Here we review the major insights into invasiveness in head and neck cancer provided by high-throughput molecular profiling approaches.

High-resolution microarray analysis of chromosome 20q in human colon cancer metastasis model systems

Amplification of human chromosome 20q DNA is the most frequently occurring chromosomal abnormality detected in sporadic colorectal carcinomas and shows significant correlation with liver metastases. Through comprehensive high-resolution microarray comparative genomic hybridization and microarray gene expression profiling, we have characterized chromosome 20q amplicon genes associated with human colorectal cancer metastasis in two in vitro metastasis model systems. The results revealed increasing complexity of the 20q genomic profile from the primary tumor-derived cell lines to the lymph node and liver metastasis derived cell lines. Expression analysis of chromosome 20q revealed a subset of over expressed genes residing within the regions of genomic copy number gain in all the tumor cell lines, suggesting these are Chromosome 20q copy number responsive genes. Bases on their preferential expression levels in the model system cell lines and known biological function, four of the over expressed genes mapping to the common intervals of genomic copy gain were considered the most promising candidate colorectal metastasis-associated genes. Validation of genomic copy number and expression array data was carried out on these genes, with one gene, DNMT3B, standing out as expressed at a relatively higher levels in the metastasis-derived cell lines compared with their primary-derived counterparts in both the models systems analyzed. The data provide evidence for the role of chromosome 20q genes with low copy gain and elevated expression in the clonal evolution of metastatic cells and suggests that such genes may serve as early biomarkers of metastatic potential. The data also support the utility of the combined microarray comparative genomic hybridization and expression array analysis for identifying copy number responsive genes in areas of low DNA copy gain in cancer cells. ^

Magnetic susceptibility and electric conductivity of marine surficial sediments by benthic electromagnetic profiling

Distribution, accumulation and diagenesis of surficial sediments in coastal and continental shelf systems follow complex chains of localized processes and form deposits of great spatial variability. Given the environmental and economic relevance of ocean margins, there is growing need for innovative geophysical exploration methods to characterize seafloor sediments by more than acoustic properties. A newly conceptualized benthic profiling and data processing approach based on controlled source electromagnetic (CSEM) imaging permits to coevally quantify the magnetic susceptibility and the electric conductivity of shallow marine deposits. The two physical properties differ fundamentally insofar as magnetic susceptibility mostly assesses solid particle characteristics such as terrigenous or iron mineral content, redox state and contamination level, while electric conductivity primarily relates to the fluid-filled pore space and detects salinity, porosity and grain-size variations. We develop and validate a layered half-space inversion algorithm for submarine multifrequency CSEM with concentric sensor configuration. Guided by results of modeling, we modified a commercial land CSEM sensor for submarine application, which was mounted into a nonconductive and nonmagnetic bottom-towed sled. This benthic EM profiler Neridis II achieves 25 soundings/second at 3-4 knots over continuous profiles of up to hundred kilometers. Magnetic susceptibility is determined from the 75 Hz in-phase response (90% signal originates from the top 50 cm), while electric conductivity is derived from the 5 kHz out-of-phase (quadrature) component (90% signal from the top 92 cm). Exemplary survey data from the north-west Iberian margin underline the excellent sensitivity, functionality and robustness of the system in littoral (~0-50 m) and neritic (~50-300 m) environments. Susceptibility vs. porosity cross-plots successfully identify known lithofacies units and their transitions. All presently available data indicate an eminent potential of CSEM profiling for assessing the complex distribution of shallow marine surficial sediments and for revealing climatic, hydrodynamic, diagenetic and anthropogenic factors governing their formation.

Using quantitative descriptive analysis and temporal dominance of sensations analysis as complementary methods for profiling commercial blackcurrant squashes

Quantitative descriptive analysis (QDA) is used to describe the nature and the intensity of sensory properties from a single evaluation of a product, whereas temporal dominance of sensation (TDS) is primarily used to identify dominant sensory properties over time. Previous studies with TDS have focused on model systems, but this is the first study to use a sequential approach, i.e. QDA then TDS in measuring sensory properties of a commercial product category, using the same set of trained assessors (n = 11). The main objectives of this study were to: (1) investigate the benefits of using a sequential approach of QDA and TDS and (2) to explore the impact of the sample composition on taste and flavour perceptions in blackcurrant squashes. The present study has proposed an alternative way of determining the choice of attributes for TDS measurement based on data obtained from previous QDA studies, where available. Both methods indicated that the flavour profile was primarily influenced by the level of dilution and complexity of sample composition combined with blackcurrant juice content. In addition, artificial sweeteners were found to modify the quality of sweetness and could also contribute to bitter notes. Using QDA and TDS in tandem was shown to be more beneficial than each just on its own enabling a more complete sensory profile of the products.

Feasibility and utility of population-level geospatial injury profiling : prospective, national cohort study

The Health Services Research Unit receives funding from the Chief Scientist Office of the Scottish Government Health and Social Care Directorates. The opinions expressed in this article are those of the authors alone. The GEOS study was funded by the North of Scotland Planning Group.

Profiling in sport using momentum and perturbations

Examining a team’s performance from a physical point of view their momentum might indicate unexpected turning points in defeat or success. Physicists describe this value as to require some effort to be started, but also that it is relatively easy to keep it going once a sufficient level is reached (Reed and Hughes, 2006). Unlike football, rugby, handball and many more sports, a regular volleyball match is not limited by time but by points that need to be gathered. Every minute more than one point is won by either one team or the other. That means a series of successive points enlarges the gap between the teams making it more and more difficult to catch up with the leading one. This concept of gathering momentum, or the reverse in a performance, can give the coaches, athletes and sports scientists further insights into winning and losing performances. Momentum investigations also contain dependencies between performances or questions if future performances are reliant upon past streaks. Squash and volleyball share the characteristic of being played up to a certain amount of points. Squash was examined according to the momentum of players by Hughes et al. (2006). The initial aim was to expand normative profiles of elite squash players using momentum graphs of winners and errors to explore ‘turning points’ in a performance. Dynamic systems theory has enabled the definition of perturbations in sports exhibiting rhythms (Hughes et al., 2000; McGarry et al., 2002; Murray et al., 2008), and how players and teams cause these disruptions of rhythm can inform on the way they play, these techniques also contribute to profiling methods. Together with the analysis of one’s own performance it is essential to have an understanding of your oppositions’ tactical strengths and weaknesses. By modelling the oppositions’ performance it is possible to predict certain outcomes and patterns, and therefore intervene or change tactics before the critical incident occurs. The modelling of competitive sport is an informative analytic technique as it directs the attention of the modeller to the critical aspects of data that delineate successful performance (McGarry & Franks, 1996). Using tactical performance profiles to pull out and visualise these critical aspects of performance, players can build justified and sophisticated tactical plans. The area is discussed and reviewed, critically appraising the research completed in this element of Performance Analysis.

Towards user profiling for web recommendation

Collaborative recommendation is one of widely used recommendation systems, which recommend items to visitor on a basis of referring other's preference that is similar to current user. User profiling technique upon Web transaction data is able to capture such informative knowledge of user task or interest. With the discovered usage pattern information, it is likely to recommend Web users more preferred content or customize the Web presentation to visitors via collaborative recommendation. In addition, it is helpful to identify the underlying relationships among Web users, items as well as latent tasks during Web mining period. In this paper, we propose a Web recommendation framework based on user profiling technique. In this approach, we employ Probabilistic Latent Semantic Analysis (PLSA) to model the co-occurrence activities and develop a modified k-means clustering algorithm to build user profiles as the representatives of usage patterns. Moreover, the hidden task model is derived by characterizing the meaningful latent factor space. With the discovered user profiles, we then choose the most matched profile, which possesses the closely similar preference to current user and make collaborative recommendation based on the corresponding page weights appeared in the selected user profile. The preliminary experimental results performed on real world data sets show that the proposed approach is capable of making recommendation accurately and efficiently.

Load profiling and data mining techniques in electricity deregulated market

This paper presents load profiles of electricity customers, using the knowledge discovery in databases (KDD) procedure, a data mining technique, to determine the load profiles for different types of customers. In this paper, the current load profiling methods are compared using data mining techniques, by analysing and evaluating these classification techniques. The objective of this study is to determine the best load profiling methods and data mining techniques to classify, detect and predict non-technical losses in the distribution sector, due to faulty metering and billing errors, as well as to gather knowledge on customer behaviour and preferences so as to gain a competitive advantage in the deregulated market. This paper focuses mainly on the comparative analysis of the classification techniques selected; a forthcoming paper will focus on the detection and prediction methods.

Optical in-fiber Bragg grating sensor systems for medical applications

Two in-fiber Bragg grating (FBG) temperature sensor systems for medical applications are demonstrated: (1) an FBG flow-directed thermodilution catheter based on interferometric detection of wavelength shift that is used for cardiac monitoring; and (2) an FBG sensor system with a tunable Fabry-Perot filter for in vivo temperature profiling in nuclear magnetic resonance (NMR) machines. Preliminary results show that the FBG sensor is in good agreement with electrical sensors that are widely used in practice. A field test shows that the FBG sensor system is suitable for in situ temperature profiling in NMR machines for medical applications.

Dispersion aspects of periodically amplified soliton transmission systems

This thesis presents improvements to optical transmission systems through the use of optical solitons as a digital transmission format, both theoretically and experimentally. An introduction to the main concepts and impairments of optical fibre on pulse transmission is included before introducing the concept of solitons in optically amplified communications and the problems of soliton system design. The theoretical work studies two fibre dispersion profiling schemes and a soliton launch improvement. The first provides superior pulse transmission by optimally tailoring the fibre dispersion to better follow the power, and hence nonlinearity, decay and thus allow soliton transmission for longer amplifier spacings and shorter pulse widths than normally possible. The second profiling scheme examines the use of dispersion compensating fibre in the context of soliton transmission over existing, standard fibre systems. The limits for solitons in uncompensated standard fibre are assessed, before the potential benefits of dispersion compensating fibre included as part of each amplifier are shown. The third theoretical investigation provides a simple improvement to the propagation of solitons in a highly perturbed system. By introducing a section of fibre of the correct length prior to the first system amplifier span, the soliton shape can be better coupled into the system thus providing an improved "average soliton" propagation model. The experimental work covers two areas. An important issue for soliton systems is pulse sources. Three potential lasers are studied, two ring laser configurations and one semiconductor device with external pulse shaping. The second area studies soliton transmission using a recalculating loop, reviewing the advantages and draw-backs of such an experiment in system testing and design. One particular example of employing the recirculating loop is also examined, using a novel method of pulse shape stabilisation over long distances with low jitter. The future for nonlinear optical communications is considered with the thesis conclusions.