text:
Genome sequencing projects have ciphered millions of protein sequence, which require knowledge of their structure and function to improve the understanding of their biological role. Although experimental methods can provide detailed information for a small fraction of these proteins, computational modeling is needed for the majority of protein molecules which are experimentally uncharacterized. The I-TASSER server is an on-line workbench for high-resolution modeling of protein structure and function. Given a protein sequence, a typical output from the I-TASSER server includes secondary structure prediction, predicted solvent accessibility of each residue, homologous template proteins detected by threading and structure alignments, up to five full-length tertiary structural models, and structure-based functional annotations for enzyme classification, Gene Ontology terms and protein-ligand binding sites. All the predictions are tagged with a confidence score which tells how accurate the predictions are without knowing the experimental data. To facilitate the special requests of end users, the server provides channels to accept user-specified inter-residue distance and contact maps to interactively change the I-TASSER modeling; it also allows users to specify any proteins as template, or to exclude any template proteins during the structure assembly simulations. The structural information could be collected by the users based on experimental evidences or biological insights with the purpose of improving the quality of I-TASSER predictions. The server was evaluated as the best programs for protein structure and function predictions in the recent community-wide CASP experiments. There are currently >20,000 registered scientists from over 100 countries who are using the on-line I-TASSER server.
21 Related JoVE Articles!
Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules
Institutions: Princeton University.
The aim of de novo
protein design is to find the amino acid sequences that will fold into a desired 3-dimensional structure with improvements in specific properties, such as binding affinity, agonist or antagonist behavior, or stability, relative to the native sequence. Protein design lies at the center of current advances drug design and discovery. Not only does protein design provide predictions for potentially useful drug targets, but it also enhances our understanding of the protein folding process and protein-protein interactions. Experimental methods such as directed evolution have shown success in protein design. However, such methods are restricted by the limited sequence space that can be searched tractably. In contrast, computational design strategies allow for the screening of a much larger set of sequences covering a wide variety of properties and functionality. We have developed a range of computational de novo
protein design methods capable of tackling several important areas of protein design. These include the design of monomeric proteins for increased stability and complexes for increased binding affinity.
To disseminate these methods for broader use we present Protein WISDOM (https://www.proteinwisdom.org), a tool that provides automated methods for a variety of protein design problems. Structural templates are submitted to initialize the design process. The first stage of design is an optimization sequence selection stage that aims at improving stability through minimization of potential energy in the sequence space. Selected sequences are then run through a fold specificity stage and a binding affinity stage. A rank-ordered list of the sequences for each step of the process, along with relevant designed structures, provides the user with a comprehensive quantitative assessment of the design. Here we provide the details of each design method, as well as several notable experimental successes attained through the use of the methods.
Genetics, Issue 77, Molecular Biology, Bioengineering, Biochemistry, Biomedical Engineering, Chemical Engineering, Computational Biology, Genomics, Proteomics, Protein, Protein Binding, Computational Biology, Drug Design, optimization (mathematics), Amino Acids, Peptides, and Proteins, De novo protein and peptide design, Drug design, In silico sequence selection, Optimization, Fold specificity, Binding affinity, sequencing
Chromosomics: Detection of Numerical and Structural Alterations in All 24 Human Chromosomes Simultaneously Using a Novel OctoChrome FISH Assay
Institutions: University of California, Berkeley .
Fluorescence in situ
hybridization (FISH) is a technique that allows specific DNA sequences to be detected on metaphase or interphase chromosomes in cell nuclei1
. The technique uses DNA probes with unique sequences that hybridize to whole chromosomes or specific chromosomal regions, and serves as a powerful adjunct to classic cytogenetics. For instance, many earlier studies reported the frequent detection of increased chromosome aberrations in leukemia patients related with benzene exposure, benzene-poisoning patients, and healthy workers exposed to benzene, using classic cytogenetic analysis2
. Using FISH, leukemia-specific chromosomal alterations have been observed to be elevated in apparently healthy workers exposed to benzene3-6
, indicating the critical roles of cytogentic changes in benzene-induced leukemogenesis.
Generally, a single FISH assay examines only one or a few whole chromosomes or specific loci per slide, so multiple hybridizations need to be conducted on multiple slides to cover all of the human chromosomes. Spectral karyotyping (SKY) allows visualization of the whole genome simultaneously, but the requirement for special software and equipment limits its application7
. Here, we describe a novel FISH assay, OctoChrome-FISH, which can be applied for Chromosomics
, which we define here as the simultaneous analysis of all 24 human chromosomes on one slide in human studies, such as chromosome-wide aneuploidy study (CWAS)8
. The basis of the method, marketed by Cytocell as the Chromoprobe Multiprobe System, is an OctoChrome device that is divided into 8 squares, each of which carries three different whole chromosome painting probes (Figure 1). Each of the three probes is directly labeled with a different colored fluorophore, green (FITC), red (Texas Red), and blue (Coumarin). The arrangement of chromosome combinations on the OctoChrome device has been designed to facilitate the identification of the non-random structural chromosome alterations (translocations) found in the most common leukemias and lymphomas, for instance t(9;22), t(15;17), t(8;21), t(14;18)9
. Moreover, numerical changes (aneuploidy) in chromosomes can be detected concurrently. The corresponding template slide is also divided into 8 squares onto which metaphase spreads are bound (Figure 2), and is positioned over the OctoChrome device. The probes and target DNA are denatured at high-temperature and hybridized in a humid chamber, and then all 24 human chromosomes can be visualized simultaneously.
OctoChrome FISH is a promising technique for the clinical diagnosis of leukemia and lymphoma and for detection of aneuploidies in all chromosomes. We have applied this new Chromosomic
approach in a CWAS study of benzene-exposed Chinese workers8,10
Genetics, Issue 60, Chromosomics, OctoChrome-FISH, fluorescence in situ hybridization (FISH), Chromosome-wide aneuploidy study (CWAS), aneuploidy, chromosomal translocations, leukemia, lymphoma
Crystallizing Membrane Proteins for Structure Determination using Lipidic Mesophases
Institutions: Trinity College Dublin.
A detailed protocol for crystallizing membrane proteins by using lipidic mesophases is described. This method has variously been referred to as the lipidic cubic phase or in meso
method. The method has been shown to be quite versatile in that it has been used to solve X-ray crystallographic structures of prokaryotic and eukaryotic proteins, proteins that are monomeric, homo- and hetero-multimeric, chromophore-containing and chromophore-free, and alpha-helical and beta-barrel proteins. Recent successes using in meso
crystallization are the human engineered beta2-adrenergic and adenosine A2a G protein-coupled receptors. Protocols are presented for reconstituting the membrane protein into the monoolein-based mesophase, and for setting up crystallizations in the manual mode. Additional steps in the overall process, such as crystal harvesting, are to be addressed in future video articles. The time required to prepare the protein-loaded mesophase and to set up a crystallization plate manually is about one hour.
Biochemistry, Issue 45, membrane protein, in meso, membrane, crystallization, lipidic mesophases, manual
High Efficiency Differentiation of Human Pluripotent Stem Cells to Cardiomyocytes and Characterization by Flow Cytometry
Institutions: Medical College of Wisconsin, Stanford University School of Medicine, Medical College of Wisconsin, Hong Kong University, Johns Hopkins University School of Medicine, Medical College of Wisconsin.
There is an urgent need to develop approaches for repairing the damaged heart, discovering new therapeutic drugs that do not have toxic effects on the heart, and improving strategies to accurately model heart disease. The potential of exploiting human induced pluripotent stem cell (hiPSC) technology to generate cardiac muscle “in a dish” for these applications continues to generate high enthusiasm. In recent years, the ability to efficiently generate cardiomyogenic cells from human pluripotent stem cells (hPSCs) has greatly improved, offering us new opportunities to model very early stages of human cardiac development not otherwise accessible. In contrast to many previous methods, the cardiomyocyte differentiation protocol described here does not require cell aggregation or the addition of Activin A or BMP4 and robustly generates cultures of cells that are highly positive for cardiac troponin I and T (TNNI3, TNNT2), iroquois-class homeodomain protein IRX-4 (IRX4), myosin regulatory light chain 2, ventricular/cardiac muscle isoform (MLC2v) and myosin regulatory light chain 2, atrial isoform (MLC2a) by day 10 across all human embryonic stem cell (hESC) and hiPSC lines tested to date. Cells can be passaged and maintained for more than 90 days in culture. The strategy is technically simple to implement and cost-effective. Characterization of cardiomyocytes derived from pluripotent cells often includes the analysis of reference markers, both at the mRNA and protein level. For protein analysis, flow cytometry is a powerful analytical tool for assessing quality of cells in culture and determining subpopulation homogeneity. However, technical variation in sample preparation can significantly affect quality of flow cytometry data. Thus, standardization of staining protocols should facilitate comparisons among various differentiation strategies. Accordingly, optimized staining protocols for the analysis of IRX4, MLC2v, MLC2a, TNNI3, and TNNT2 by flow cytometry are described.
Cellular Biology, Issue 91, human induced pluripotent stem cell, flow cytometry, directed differentiation, cardiomyocyte, IRX4, TNNI3, TNNT2, MCL2v, MLC2a
A Research Method For Detecting Transient Myocardial Ischemia In Patients With Suspected Acute Coronary Syndrome Using Continuous ST-segment Analysis
Institutions: University of Nevada, Reno, St. Joseph's Medical Center, University of Rochester Medical Center .
Each year, an estimated 785,000 Americans will have a new coronary attack, or acute coronary syndrome (ACS). The pathophysiology of ACS involves rupture of an atherosclerotic plaque; hence, treatment is aimed at plaque stabilization in order to prevent cellular death. However, there is considerable debate among clinicians, about which treatment pathway is best: early invasive using percutaneous coronary intervention (PCI/stent) when indicated or a conservative approach (i.e.
, medication only with PCI/stent if recurrent symptoms occur).
There are three types of ACS: ST elevation myocardial infarction (STEMI), non-ST elevation MI (NSTEMI), and unstable angina (UA). Among the three types, NSTEMI/UA is nearly four times as common as STEMI. Treatment decisions for NSTEMI/UA are based largely on symptoms and resting or exercise electrocardiograms (ECG). However, because of the dynamic and unpredictable nature of the atherosclerotic plaque, these methods often under detect myocardial ischemia because symptoms are unreliable, and/or continuous ECG monitoring was not utilized.
Continuous 12-lead ECG monitoring, which is both inexpensive and non-invasive, can identify transient episodes of myocardial ischemia, a precursor to MI, even when asymptomatic. However, continuous 12-lead ECG monitoring is not usual hospital practice; rather, only two leads are typically monitored. Information obtained with 12-lead ECG monitoring might provide useful information for deciding the best ACS treatment.
Therefore, using 12-lead ECG monitoring, the COMPARE Study (electroC
n of ischeM
sive to phaR
atment) was designed to assess the frequency and clinical consequences of transient myocardial ischemia, in patients with NSTEMI/UA treated with either early invasive PCI/stent or those managed conservatively (medications or PCI/stent following recurrent symptoms). The purpose of this manuscript is to describe the methodology used in the COMPARE Study.
Permission to proceed with this study was obtained from the Institutional Review Board of the hospital and the university. Research nurses identify hospitalized patients from the emergency department and telemetry unit with suspected ACS. Once consented, a 12-lead ECG Holter monitor is applied, and remains in place during the patient's entire hospital stay. Patients are also maintained on the routine bedside ECG monitoring system per hospital protocol. Off-line ECG analysis is done using sophisticated software and careful human oversight.
Medicine, Issue 70, Anatomy, Physiology, Cardiology, Myocardial Ischemia, Cardiovascular Diseases, Health Occupations, Health Care, transient myocardial ischemia, Acute Coronary Syndrome, electrocardiogram, ST-segment monitoring, Holter monitoring, research methodology
Measuring Attentional Biases for Threat in Children and Adults
Institutions: Rutgers University.
Investigators have long been interested in the human propensity for the rapid detection of threatening stimuli. However, until recently, research in this domain has focused almost exclusively on adult participants, completely ignoring the topic of threat detection over the course of development. One of the biggest reasons for the lack of developmental work in this area is likely the absence of a reliable paradigm that can measure perceptual biases for threat in children. To address this issue, we recently designed a modified visual search paradigm similar to the standard adult paradigm that is appropriate for studying threat detection in preschool-aged participants. Here we describe this new procedure. In the general paradigm, we present participants with matrices of color photographs, and ask them to find and touch a target on the screen. Latency to touch the target is recorded. Using a touch-screen monitor makes the procedure simple and easy, allowing us to collect data in participants ranging from 3 years of age to adults. Thus far, the paradigm has consistently shown that both adults and children detect threatening stimuli (e.g.,
snakes, spiders, angry/fearful faces) more quickly than neutral stimuli (e.g.,
flowers, mushrooms, happy/neutral faces). Altogether, this procedure provides an important new tool for researchers interested in studying the development of attentional biases for threat.
Behavior, Issue 92, Detection, threat, attention, attentional bias, anxiety, visual search
Reverse Yeast Two-hybrid System to Identify Mammalian Nuclear Receptor Residues that Interact with Ligands and/or Antagonists
Institutions: Albert Einstein College of Medicine , Shanghai University of Traditional Chinese Medicine.
As a critical regulator of drug metabolism and inflammation, Pregnane X Receptor (PXR), plays an important role in disease pathophysiology linking metabolism and inflammation (e.g.
. There has been much progress in the identification of agonist ligands for PXR, however, there are limited descriptions of drug-like antagonists and their binding sites on PXR3,4,5
. A critical barrier has been the inability to efficiently purify full-length protein for structural studies with antagonists despite the fact that PXR was cloned and characterized in 1998. Our laboratory developed a novel high throughput yeast based two-hybrid assay to define an antagonist, ketoconazole's, binding residues on PXR6
. Our method involves creating mutational libraries that would rescue the effect of single mutations on the AF-2 surface of PXR expected to interact with ketoconazole. Rescue or "gain-of-function" second mutations can be made such that conclusions regarding the genetic interaction of ketoconazole and the surface residue(s) on PXR are feasible. Thus, we developed a high throughput two-hybrid yeast screen of PXR mutants interacting with its coactivator, SRC-1. Using this approach, in which the yeast was modified to accommodate the study of the antifungal drug, ketoconazole, we could demonstrate specific mutations on PXR enriched in clones unable to bind to ketoconazole. By reverse logic, we conclude that the original residues are direct interaction residues with ketoconazole. This assay represents a novel, tractable genetic assay to screen for antagonist binding sites on nuclear receptor surfaces. This assay could be applied to any drug regardless of its cytotoxic potential to yeast as well as to cellular protein(s) that cannot be studied using standard structural biology or proteomic based methods. Potential pitfalls include interpretation of data (complementary methods useful), reliance on single Y2H method, expertise in handling yeast or performing yeast two-hybrid assays, and assay optimization.
Biochemistry, Issue 81, Orphan nuclear receptor, ketoconazole, yeast two-hybrid, Pregnane X Receptor, ligand, antatogist, coactivators SRC-1 (steroid receptor coactivator 1), drug-receptor interaction
Designing Silk-silk Protein Alloy Materials for Biomedical Applications
Institutions: Rowan University, Rowan University, Cooper Medical School of Rowan University, Rowan University.
Fibrous proteins display different sequences and structures that have been used for various applications in biomedical fields such as biosensors, nanomedicine, tissue regeneration, and drug delivery. Designing materials based on the molecular-scale interactions between these proteins will help generate new multifunctional protein alloy biomaterials with tunable properties. Such alloy material systems also provide advantages in comparison to traditional synthetic polymers due to the materials biodegradability, biocompatibility, and tenability in the body. This article used the protein blends of wild tussah silk (Antheraea pernyi
) and domestic mulberry silk (Bombyx mori
) as an example to provide useful protocols regarding these topics, including how to predict protein-protein interactions by computational methods, how to produce protein alloy solutions, how to verify alloy systems by thermal analysis, and how to fabricate variable alloy materials including optical materials with diffraction gratings, electric materials with circuits coatings, and pharmaceutical materials for drug release and delivery. These methods can provide important information for designing the next generation multifunctional biomaterials based on different protein alloys.
Bioengineering, Issue 90, protein alloys, biomaterials, biomedical, silk blends, computational simulation, implantable electronic devices
The Multiple Sclerosis Performance Test (MSPT): An iPad-Based Disability Assessment Tool
Institutions: Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation.
Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report a new test, the Multiple Sclerosis Performance Test (MSPT), which represents a new approach to quantifying MS related disability. The MSPT takes advantage of advances in computer technology, information technology, biomechanics, and clinical measurement science. The resulting MSPT represents a computer-based platform for precise, valid measurement of MS severity. Based on, but extending the Multiple Sclerosis Functional Composite (MSFC), the MSPT provides precise, quantitative data on walking speed, balance, manual dexterity, visual function, and cognitive processing speed. The MSPT was tested by 51 MS patients and 49 healthy controls (HC). MSPT scores were highly reproducible, correlated strongly with technician-administered test scores, discriminated MS from HC and severe from mild MS, and correlated with patient reported outcomes. Measures of reliability, sensitivity, and clinical meaning for MSPT scores were favorable compared with technician-based testing. The MSPT is a potentially transformative approach for collecting MS disability outcome data for patient care and research. Because the testing is computer-based, test performance can be analyzed in traditional or novel ways and data can be directly entered into research or clinical databases. The MSPT could be widely disseminated to clinicians in practice settings who are not connected to clinical trial performance sites or who are practicing in rural settings, drastically improving access to clinical trials for clinicians and patients. The MSPT could be adapted to out of clinic settings, like the patient’s home, thereby providing more meaningful real world data. The MSPT represents a new paradigm for neuroperformance testing. This method could have the same transformative effect on clinical care and research in MS as standardized computer-adapted testing has had in the education field, with clear potential to accelerate progress in clinical care and research.
Medicine, Issue 88, Multiple Sclerosis, Multiple Sclerosis Functional Composite, computer-based testing, 25-foot walk test, 9-hole peg test, Symbol Digit Modalities Test, Low Contrast Visual Acuity, Clinical Outcome Measure
Developing Neuroimaging Phenotypes of the Default Mode Network in PTSD: Integrating the Resting State, Working Memory, and Structural Connectivity
Institutions: Alpert Medical School, Brown University, University of Georgia.
Complementary structural and functional neuroimaging techniques used to examine the Default Mode Network (DMN) could potentially improve assessments of psychiatric illness severity and provide added validity to the clinical diagnostic process. Recent neuroimaging research suggests that DMN processes may be disrupted in a number of stress-related psychiatric illnesses, such as posttraumatic stress disorder (PTSD).
Although specific DMN functions remain under investigation, it is generally thought to be involved in introspection and self-processing. In healthy individuals it exhibits greatest activity during periods of rest, with less activity, observed as deactivation, during cognitive tasks, e.g.
, working memory. This network consists of the medial prefrontal cortex, posterior cingulate cortex/precuneus, lateral parietal cortices and medial temporal regions.
Multiple functional and structural imaging approaches have been developed to study the DMN. These have unprecedented potential to further the understanding of the function and dysfunction of this network. Functional approaches, such as the evaluation of resting state connectivity and task-induced deactivation, have excellent potential to identify targeted neurocognitive and neuroaffective (functional) diagnostic markers and may indicate illness severity and prognosis with increased accuracy or specificity. Structural approaches, such as evaluation of morphometry and connectivity, may provide unique markers of etiology and long-term outcomes. Combined, functional and structural methods provide strong multimodal, complementary and synergistic approaches to develop valid DMN-based imaging phenotypes in stress-related psychiatric conditions. This protocol aims to integrate these methods to investigate DMN structure and function in PTSD, relating findings to illness severity and relevant clinical factors.
Medicine, Issue 89, default mode network, neuroimaging, functional magnetic resonance imaging, diffusion tensor imaging, structural connectivity, functional connectivity, posttraumatic stress disorder
High Throughput Quantitative Expression Screening and Purification Applied to Recombinant Disulfide-rich Venom Proteins Produced in E. coli
Institutions: Aix-Marseille Université, Commissariat à l'énergie atomique et aux énergies alternatives (CEA) Saclay, France.
Escherichia coli (E. coli)
is the most widely used expression system for the production of recombinant proteins for structural and functional studies. However, purifying proteins is sometimes challenging since many proteins are expressed in an insoluble form. When working with difficult or multiple targets it is therefore recommended to use high throughput (HTP) protein expression screening on a small scale (1-4 ml cultures) to quickly identify conditions for soluble expression. To cope with the various structural genomics programs of the lab, a quantitative (within a range of 0.1-100 mg/L culture of recombinant protein) and HTP protein expression screening protocol was implemented and validated on thousands of proteins. The protocols were automated with the use of a liquid handling robot but can also be performed manually without specialized equipment.
Disulfide-rich venom proteins are gaining increasing recognition for their potential as therapeutic drug leads. They can be highly potent and selective, but their complex disulfide bond networks make them challenging to produce. As a member of the FP7 European Venomics project (www.venomics.eu), our challenge is to develop successful production strategies with the aim of producing thousands of novel venom proteins for functional characterization. Aided by the redox properties of disulfide bond isomerase DsbC, we adapted our HTP production pipeline for the expression of oxidized, functional venom peptides in the E. coli
cytoplasm. The protocols are also applicable to the production of diverse disulfide-rich proteins. Here we demonstrate our pipeline applied to the production of animal venom proteins. With the protocols described herein it is likely that soluble disulfide-rich proteins will be obtained in as little as a week. Even from a small scale, there is the potential to use the purified proteins for validating the oxidation state by mass spectrometry, for characterization in pilot studies, or for sensitive micro-assays.
Bioengineering, Issue 89, E. coli, expression, recombinant, high throughput (HTP), purification, auto-induction, immobilized metal affinity chromatography (IMAC), tobacco etch virus protease (TEV) cleavage, disulfide bond isomerase C (DsbC) fusion, disulfide bonds, animal venom proteins/peptides
Coordinate Mapping of Hyolaryngeal Mechanics in Swallowing
Institutions: Georgia Regents University, New York University, Georgia Regents University, Georgia Regents University.
Characterizing hyolaryngeal movement is important to dysphagia research. Prior methods require multiple measurements to obtain one kinematic measurement whereas coordinate mapping of hyolaryngeal mechanics using Modified Barium Swallow (MBS) uses one set of coordinates to calculate multiple variables of interest. For demonstration purposes, ten kinematic measurements were generated from one set of coordinates to determine differences in swallowing two different bolus types. Calculations of hyoid excursion against the vertebrae and mandible are correlated to determine the importance of axes of reference.
To demonstrate coordinate mapping methodology, 40 MBS studies were randomly selected from a dataset of healthy normal subjects with no known swallowing impairment. A 5 ml thin-liquid bolus and a 5 ml pudding swallows were measured from each subject. Nine coordinates, mapping the cranial base, mandible, vertebrae and elements of the hyolaryngeal complex, were recorded at the frames of minimum and maximum hyolaryngeal excursion. Coordinates were mathematically converted into ten variables of hyolaryngeal mechanics.
Inter-rater reliability was evaluated by Intraclass correlation coefficients (ICC). Two-tailed t-tests were used to evaluate differences in kinematics by bolus viscosity. Hyoid excursion measurements against different axes of reference were correlated. Inter-rater reliability among six raters for the 18 coordinates ranged from ICC = 0.90 - 0.97. A slate of ten kinematic measurements was compared by subject between the six raters. One outlier was rejected, and the mean of the remaining reliability scores was ICC = 0.91, 0.84 - 0.96, 95% CI. Two-tailed t-tests with Bonferroni corrections comparing ten kinematic variables (5 ml thin-liquid vs. 5 ml pudding swallows) showed statistically significant differences in hyoid excursion, superior laryngeal movement, and pharyngeal shortening (p
< 0.005). Pearson correlations of hyoid excursion measurements from two different axes of reference were: r = 0.62, r2
= 0.38, (thin-liquid); r = 0.52, r2
= 0.27, (pudding).
Obtaining landmark coordinates is a reliable method to generate multiple kinematic variables from video fluoroscopic images useful in dysphagia research.
Medicine, Issue 87, videofluoroscopy, modified barium swallow studies, hyolaryngeal kinematics, deglutition, dysphagia, dysphagia research, hyolaryngeal complex
Generation of Comprehensive Thoracic Oncology Database - Tool for Translational Research
Institutions: University of Chicago, University of Chicago, Northshore University Health Systems, University of Chicago, University of Chicago, University of Chicago.
The Thoracic Oncology Program Database Project was created to serve as a comprehensive, verified, and accessible repository for well-annotated cancer specimens and clinical data to be available to researchers within the Thoracic Oncology Research Program. This database also captures a large volume of genomic and proteomic data obtained from various tumor tissue studies. A team of clinical and basic science researchers, a biostatistician, and a bioinformatics expert was convened to design the database. Variables of interest were clearly defined and their descriptions were written within a standard operating manual to ensure consistency of data annotation. Using a protocol for prospective tissue banking and another protocol for retrospective banking, tumor and normal tissue samples from patients consented to these protocols were collected. Clinical information such as demographics, cancer characterization, and treatment plans for these patients were abstracted and entered into an Access database. Proteomic and genomic data have been included in the database and have been linked to clinical information for patients described within the database. The data from each table were linked using the relationships function in Microsoft Access to allow the database manager to connect clinical and laboratory information during a query. The queried data can then be exported for statistical analysis and hypothesis generation.
Medicine, Issue 47, Database, Thoracic oncology, Bioinformatics, Biorepository, Microsoft Access, Proteomics, Genomics
Creating Dynamic Images of Short-lived Dopamine Fluctuations with lp-ntPET: Dopamine Movies of Cigarette Smoking
Institutions: Yale University, Yale University, Yale University, Yale University, Massachusetts General Hospital, University of California, Irvine.
We describe experimental and statistical steps for creating dopamine movies of the brain from dynamic PET data. The movies represent minute-to-minute fluctuations of dopamine induced by smoking a cigarette. The smoker is imaged during a natural smoking experience while other possible confounding effects (such as head motion, expectation, novelty, or aversion to smoking repeatedly) are minimized.
We present the details of our unique analysis. Conventional methods for PET analysis estimate time-invariant kinetic model parameters which cannot capture short-term fluctuations in neurotransmitter release. Our analysis - yielding a dopamine movie - is based on our work with kinetic models and other decomposition techniques that allow for time-varying parameters 1-7
. This aspect of the analysis - temporal-variation - is key to our work. Because our model is also linear in parameters, it is practical, computationally, to apply at the voxel level. The analysis technique is comprised of five main steps: pre-processing, modeling, statistical comparison, masking and visualization. Preprocessing is applied to the PET data with a unique 'HYPR' spatial filter 8
that reduces spatial noise but preserves critical temporal information. Modeling identifies the time-varying function that best describes the dopamine effect on 11
C-raclopride uptake. The statistical step compares the fit of our (lp-ntPET) model 7
to a conventional model 9
. Masking restricts treatment to those voxels best described by the new model. Visualization maps the dopamine function at each voxel to a color scale and produces a dopamine movie. Interim results and sample dopamine movies of cigarette smoking are presented.
Behavior, Issue 78, Neuroscience, Neurobiology, Molecular Biology, Biomedical Engineering, Medicine, Anatomy, Physiology, Image Processing, Computer-Assisted, Receptors, Dopamine, Dopamine, Functional Neuroimaging, Binding, Competitive, mathematical modeling (systems analysis), Neurotransmission, transient, dopamine release, PET, modeling, linear, time-invariant, smoking, F-test, ventral-striatum, clinical techniques
Obtaining Specimens with Slowed, Accelerated and Reversed Aging in the Honey Bee Model
Institutions: Norwegian University of Life Sciences, Arizona State University.
Societies of highly social animals feature vast lifespan differences between closely related individuals. Among social insects, the honey bee is the best established model to study how plasticity in lifespan and aging is explained by social factors.
The worker caste of honey bees includes nurse bees, which tend the brood, and forager bees, which collect nectar and pollen. Previous work has shown that brain functions and flight performance senesce more rapidly in foragers than in nurses. However, brain functions can recover, when foragers revert back to nursing tasks. Such patterns of accelerated and reversed functional senescence are linked to changed metabolic resource levels, to alterations in protein abundance and to immune function. Vitellogenin, a yolk protein with adapted functions in hormonal control and cellular defense, may serve as a major regulatory element in a network that controls the different aging dynamics in workers.
Here we describe how the emergence of nurses and foragers can be monitored, and manipulated, including the reversal from typically short-lived foragers into longer-lived nurses. Our representative results show how individuals with similar chronological age differentiate into foragers and nurse bees under experimental conditions. We exemplify how behavioral reversal from foragers back to nurses can be validated. Last, we show how different cellular senescence can be assessed by measuring the accumulation of lipofuscin, a universal biomarker of senescence.
For studying mechanisms that may link social influences and aging plasticity, this protocol provides a standardized tool set to acquire relevant sample material, and to improve data comparability among future studies.
Developmental Biology, Issue 78, Insects, Microscopy, Confocal, Aging, Gerontology, Neurobiology, Insect, Invertebrate, Brain, Lipofuscin, Confocal Microscopy
Perceptual and Category Processing of the Uncanny Valley Hypothesis' Dimension of Human Likeness: Some Methodological Issues
Institutions: University of Zurich.
Mori's Uncanny Valley Hypothesis1,2
proposes that the perception of humanlike characters such as robots and, by extension, avatars (computer-generated characters) can evoke negative or positive affect (valence) depending on the object's degree of visual and behavioral realism along a dimension of human likeness
) (Figure 1
). But studies of affective valence of subjective responses to variously realistic non-human characters have produced inconsistent findings 3, 4, 5, 6
. One of a number of reasons for this is that human likeness is not perceived as the hypothesis assumes. While the DHL can be defined following Mori's description as a smooth linear change in the degree of physical humanlike similarity, subjective perception of objects along the DHL can be understood in terms of the psychological effects of categorical perception (CP) 7
. Further behavioral and neuroimaging investigations of category processing and CP along the DHL and of the potential influence of the dimension's underlying category structure on affective experience are needed. This protocol therefore focuses on the DHL and allows examination of CP. Based on the protocol presented in the video as an example, issues surrounding the methodology in the protocol and the use in "uncanny" research of stimuli drawn from morph continua to represent the DHL are discussed in the article that accompanies the video. The use of neuroimaging and morph stimuli to represent the DHL in order to disentangle brain regions neurally responsive to physical human-like similarity from those responsive to category change and category processing is briefly illustrated.
Behavior, Issue 76, Neuroscience, Neurobiology, Molecular Biology, Psychology, Neuropsychology, uncanny valley, functional magnetic resonance imaging, fMRI, categorical perception, virtual reality, avatar, human likeness, Mori, uncanny valley hypothesis, perception, magnetic resonance imaging, MRI, imaging, clinical techniques
Cortical Source Analysis of High-Density EEG Recordings in Children
Institutions: UCL Institute of Child Health, University College London.
EEG is traditionally described as a neuroimaging technique with high temporal and low spatial resolution. Recent advances in biophysical modelling and signal processing make it possible to exploit information from other imaging modalities like structural MRI that provide high spatial resolution to overcome this constraint1
. This is especially useful for investigations that require high resolution in the temporal as well as spatial domain. In addition, due to the easy application and low cost of EEG recordings, EEG is often the method of choice when working with populations, such as young children, that do not tolerate functional MRI scans well. However, in order to investigate which neural substrates are involved, anatomical information from structural MRI is still needed. Most EEG analysis packages work with standard head models that are based on adult anatomy. The accuracy of these models when used for children is limited2
, because the composition and spatial configuration of head tissues changes dramatically over development3
In the present paper, we provide an overview of our recent work in utilizing head models based on individual structural MRI scans or age specific head models to reconstruct the cortical generators of high density EEG. This article describes how EEG recordings are acquired, processed, and analyzed with pediatric populations at the London Baby Lab, including laboratory setup, task design, EEG preprocessing, MRI processing, and EEG channel level and source analysis.
Behavior, Issue 88, EEG, electroencephalogram, development, source analysis, pediatric, minimum-norm estimation, cognitive neuroscience, event-related potentials
A Proboscis Extension Response Protocol for Investigating Behavioral Plasticity in Insects: Application to Basic, Biomedical, and Agricultural Research
Institutions: Arizona State University.
Insects modify their responses to stimuli through experience of associating those stimuli with events important for survival (e.g.
, food, mates, threats). There are several behavioral mechanisms through which an insect learns salient associations and relates them to these events. It is important to understand this behavioral plasticity for programs aimed toward assisting insects that are beneficial for agriculture. This understanding can also be used for discovering solutions to biomedical and agricultural problems created by insects that act as disease vectors and pests. The Proboscis Extension Response (PER) conditioning protocol was developed for honey bees (Apis mellifera
) over 50 years ago to study how they perceive and learn about floral odors, which signal the nectar and pollen resources a colony needs for survival. The PER procedure provides a robust and easy-to-employ framework for studying several different ecologically relevant mechanisms of behavioral plasticity. It is easily adaptable for use with several other insect species and other behavioral reflexes. These protocols can be readily employed in conjunction with various means for monitoring neural activity in the CNS via electrophysiology or bioimaging, or for manipulating targeted neuromodulatory pathways. It is a robust assay for rapidly detecting sub-lethal effects on behavior caused by environmental stressors, toxins or pesticides.
We show how the PER protocol is straightforward to implement using two procedures. One is suitable as a laboratory exercise for students or for quick assays of the effect of an experimental treatment. The other provides more thorough control of variables, which is important for studies of behavioral conditioning. We show how several measures for the behavioral response ranging from binary yes/no to more continuous variable like latency and duration of proboscis extension can be used to test hypotheses. And, we discuss some pitfalls that researchers commonly encounter when they use the procedure for the first time.
Neuroscience, Issue 91, PER, conditioning, honey bee, olfaction, olfactory processing, learning, memory, toxin assay
Improving IV Insulin Administration in a Community Hospital
Institutions: Wyoming Medical Center.
Diabetes mellitus is a major independent risk factor for increased morbidity and mortality in the hospitalized patient, and elevated blood glucose concentrations, even in non-diabetic patients, predicts poor outcomes.1-4
The 2008 consensus statement by the American Association of Clinical Endocrinologists (AACE) and the American Diabetes Association (ADA) states that "hyperglycemia in hospitalized patients, irrespective of its cause, is unequivocally associated with adverse outcomes."5
It is important to recognize that hyperglycemia occurs in patients with known or undiagnosed diabetes as well as during acute illness in those with previously normal glucose tolerance.
The Normoglycemia in Intensive Care Evaluation-Survival Using Glucose Algorithm Regulation (NICE-SUGAR) study involved over six thousand adult intensive care unit (ICU) patients who were randomized to intensive glucose control or conventional glucose control.6
Surprisingly, this trial found that intensive glucose control increased the risk of mortality by 14% (odds ratio, 1.14; p=0.02). In addition, there was an increased prevalence of severe hypoglycemia in the intensive control group compared with the conventional control group (6.8% vs. 0.5%, respectively; p<0.001). From this pivotal trial and two others,7,8
Wyoming Medical Center (WMC) realized the importance of controlling hyperglycemia in the hospitalized patient while avoiding the negative impact of resultant hypoglycemia.
Despite multiple revisions of an IV insulin paper protocol, analysis of data from usage of the paper protocol at WMC shows that in terms of achieving normoglycemia while minimizing hypoglycemia, results were suboptimal. Therefore, through a systematical implementation plan, monitoring of patient blood glucose levels was switched from using a paper IV insulin protocol to a computerized glucose management system. By comparing blood glucose levels using the paper protocol to that of the computerized system, it was determined, that overall, the computerized glucose management system resulted in more rapid and tighter glucose control than the traditional paper protocol. Specifically, a substantial increase in the time spent within the target blood glucose concentration range, as well as a decrease in the prevalence of severe hypoglycemia (BG < 40 mg/dL), clinical hypoglycemia (BG < 70 mg/dL), and hyperglycemia (BG > 180 mg/dL), was witnessed in the first five months after implementation of the computerized glucose management system. The computerized system achieved target concentrations in greater than 75% of all readings while minimizing the risk of hypoglycemia. The prevalence of hypoglycemia (BG < 70 mg/dL) with the use of the computer glucose management system was well under 1%.
Medicine, Issue 64, Physiology, Computerized glucose management, Endotool, hypoglycemia, hyperglycemia, diabetes, IV insulin, paper protocol, glucose control
Using Visual and Narrative Methods to Achieve Fair Process in Clinical Care
Institutions: Brandeis University, Brandeis University.
The Institute of Medicine has targeted patient-centeredness as an important area of quality improvement. A major dimension of patient-centeredness is respect for patient's values, preferences, and expressed needs. Yet specific approaches to gaining this understanding and translating it to quality care in the clinical setting are lacking. From a patient perspective quality is not a simple concept but is best understood in terms of five dimensions: technical outcomes; decision-making efficiency; amenities and convenience; information and emotional support; and overall patient satisfaction. Failure to consider quality from this five-pronged perspective results in a focus on medical outcomes, without considering the processes central to quality from the patient's perspective and vital to achieving good outcomes. In this paper, we argue for applying the concept of fair process in clinical settings. Fair process involves using a collaborative approach to exploring diagnostic issues and treatments with patients, explaining the rationale for decisions, setting expectations about roles and responsibilities, and implementing a core plan and ongoing evaluation. Fair process opens the door to bringing patient expertise into the clinical setting and the work of developing health care goals and strategies. This paper provides a step by step illustration of an innovative visual approach, called photovoice or photo-elicitation, to achieve fair process in clinical work with acquired brain injury survivors and others living with chronic health conditions. Applying this visual tool and methodology in the clinical setting will enhance patient-provider communication; engage patients as partners in identifying challenges, strengths, goals, and strategies; and support evaluation of progress over time. Asking patients to bring visuals of their lives into the clinical interaction can help to illuminate gaps in clinical knowledge, forge better therapeutic relationships with patients living with chronic conditions such as brain injury, and identify patient-centered goals and possibilities for healing. The process illustrated here can be used by clinicians, (primary care physicians, rehabilitation therapists, neurologists, neuropsychologists, psychologists, and others) working with people living with chronic conditions such as acquired brain injury, mental illness, physical disabilities, HIV/AIDS, substance abuse, or post-traumatic stress, and by leaders of support groups for the types of patients described above and their family members or caregivers.
Medicine, Issue 48, person-centered care, participatory visual methods, photovoice, photo-elicitation, narrative medicine, acquired brain injury, disability, rehabilitation, palliative care
Interview: Protein Folding and Studies of Neurodegenerative Diseases
Institutions: MIT - Massachusetts Institute of Technology.
In this interview, Dr. Lindquist describes relationships between protein folding, prion diseases and neurodegenerative disorders. The problem of the protein folding is at the core of the modern biology. In addition to their traditional biochemical functions, proteins can mediate transfer of biological information and therefore can be considered a genetic material. This recently discovered function of proteins has important implications for studies of human disorders. Dr. Lindquist also describes current experimental approaches to investigate the mechanism of neurodegenerative diseases based on genetic studies in model organisms.
Neuroscience, issue 17, protein folding, brain, neuron, prion, neurodegenerative disease, yeast, screen, Translational Research

domain: medical


text: High-speed machining is investigated in a new viewpoint, thermodynamic theory. Many phenomena in cutting, such as heat conduction, chemical reaction, diffusion and so on, can be studied with thermodynamic theory. In this paper, entropy generation and entropy flow in cutting system are revealed as well as the interaction of sub processes. As a case study, the oxidation of cemented carbide tool was studied in thermodynamic theory and experiments. The results show that thermodynamic theory is effective.

domain: literature


text:
Speakers include CP Foods’ Robins McIntosh and Hamish Rodger of FishVet Group
Wally Stevens, executive director of the Global Aquaculture Alliance, asks the same question – what is the aquaculture industry’s greatest challenge? – at GAA’s annual GOAL conference, and each year he hears the same answer from the audience: disease.
Major diseases have impacted global farmed shrimp and salmon production, for example, throughout the industries’ relatively brief history. This year at GOAL – held at the White Swan Hotel in Guangzhou, China – leading animal welfare and aquatic animal disease experts will share their knowledge about how producers can gird their operations during the panel discussion titled “Animal Health & Welfare: Latest Developments in Disease & Area Management.”
(Day 1, Sept. 20, 10:30 to 11:30 a.m. in the Grand Ballroom. See full conference schedule here: https://www.aquaculturealliance.org/goal/conference-program/.)
“Disease is the No. 1 constraint limiting aquaculture,” said George Chamberlain, president of the GAA. “It can impact profitability, sustainability, animal welfare and market acceptance.”
We are beginning to roll back the mystery [of EMS]. Effective solutions have been found, but there is still a reluctance on the part of some to embrace these solutions.
Chamberlain will moderate the discussion with other noted disease experts, including Robins McIntosh, senior VP at Charoen Pokphand Foods Public Company, based in Thailand; and Hamish Rodger, global managing director at FishVet Group, a Benchmark Company based in Scotland. The session will discuss advances in the control of the shrimp disease EMS (Early Mortality Syndrome), EHP (Enterocytozoon hepatopenaei) infections in shrimp ponds, streptococcosis in tilapia, and ISA (Infectious Salmon Anemia) and sea lice in farmed salmon.
“Disease control is best achieved by an integrated health management program including diagnostic testing, breeding, vaccines, biosecurity, sanitation and husbandry practices,” added Chamberlain.
EMS, also known as Acute Hepatopancreatic Necrosis Disease (AHPND), had a severe impact on global farmed shrimp supplies beginning in 2009 in China. It soon spread to other areas in Southeast Asia, costing the industry billions of dollars, and remains a very complicated issue, said McIntosh.
“We are beginning to roll back the mystery. Effective solutions have been found, but there is still a reluctance on the part of some to embrace these solutions,” he said. “But for those that have, and many are in Thailand, the outcomes are very rewarding.”
In this battle, McIntosh added, shrimp farmers are not fighting just one disease. EMS is a “concurrent synthesis” of several diseases and bacteria, he noted, that often makes test tube results irrelevant. It is therefore difficult, he said, to determine the biggest problem in a particular pond because each situation can present unique obstacles.
“We tend to give a name to a singular farm issue – such as, that farm has WSSV (White Spot Syndrome Virus), or that pond has EHP, or that pond has AHPND,” said McIntosh. “The fact is, the pond often has more than one and it is the interactions that often makes the interpretation and solutions more difficult.”
While the farmed salmon industry can be viewed as a success story in aquaculture’s war against aquatic animal diseases, Rodger noted that the naturally occurring sea lice (L. salmonis and Caligus sp.) parasite remains a challenge in many regions where salmon is produced.
“Novel methods for the prevention, control and management of these parasites have been developed and utilized in the past few years and details of these will be discussed and presented including the use of cleaner fish, mechanical and environmental management methods, novel monitoring and intervention strategies, use of genomics, alternative chemotherapy and combinations and rotations of the above,” Rodger wrote in an e-mail.
Genetic selection, biosecurity, vaccination, nutritional support and management strategies are all key tools for producers in the fight against aquatic diseases, he added, including streptococcosis in tilapia.
It makes sense for producers to work together to put themselves in the best possible situation to help prevent (or mitigate) outbreaks.
As the industry convenes to discuss disease, Iain Shone, development director at GAA, will present the latest on what is a potentially a key defense strategy known as area management.
Shone calls area management the “final frontier” for aquaculture development, adding that a select number of countries – as well as species sectors – have good examples of advanced area management systems, but it’s “far from commonplace.” Shone will detail for how GAA is approaching an area management standard, with a focus on biosecurity.
“There is always a new disease. It makes sense for producers to work together to put themselves in the best possible situation to help prevent (or mitigate) outbreaks,” said Shone. “The draft standard we are developing lays out best practice to do so. But we know this is an extremely difficult task, with many obstacles to overcome. Encouraging producers to work together can be extremely difficult for lots of reasons, but we have had great support for this development, including from the other certification schemes.”
Now that you've reached the end of the article ...
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Health & Welfare
Piscirickettsia salmonis, or P. sal, is a persistent pathogen in the waters off Chile, causing millions of dollars of damage to the nation’s massive aquaculture industry. A university researcher 6,000 miles to the north hopes to provide insight that leads to the development of an effective vaccine.
Health & Welfare
Early Mortality Syndrome has devastated farmed shrimp in Asia and Latin America. With better understanding of the pathogen and the development and improvement of novel strategies, shrimp farmers are now able to better manage the disease.
Health & Welfare
Authors examined the histological features from shrimp infected with the emerging microsporidian parasite Enterocytozoon hepatopenaei (EHP). A PCR assay method was used to detected in hepatopancreatic tissue, feces and water sampled from infected shrimp tanks, and in some samples of Artemia biomass.
Health & Welfare
A study was conducted to measure the effects of commercial probiotics on Pacific white shrimp in a standardized AHPND challenge model under controlled laboratory conditions. Results show that the probiotics treatments by themselves have beneficial effects, such as higher survival and histological signs of hepatopancreas regeneration.

domain: news


text:
Another great quarter is about to end, the company will be holding a conference call to discuss its earnings release on July 24th as per the company’s investor relations website. The call will be live streamed and the earnings releases will be available shortly before the call on Apple’s site. We are expecting huge from the company side as Apple turning bigger as the day passes on every instance.
Back in April, Apple announced its earnings for second fiscal quarter with revenue was $39.2 billion. With earnings of $13.87 per share, Apple’s net profit in Q2 was $11.6 billion. The company even sold 35.1 million iPhones and 11.8 million iPads last quarter. The company also sold 4 million Macs and 7.7 million iPods.


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8 POST CONTACTORS FOUND IN WARN® ATV WINCH KITS
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Warn Industries is voluntarily recalling 8 post contactors that were part of
winch kits for All Terrain Vehicles and Utility Vehicles (ATV's).*
* RECALL INVOLVES 2500 - 3500 LB. WINCH KITS
POTENTIAL FIRE HAZARD
WARN received one confirmed report of an 8 post contactor failure that resulted in excessive heat within the contactor. Under rare circumstances, the contactor could potentially create a fire hazard.
Consumers who have WARN ATV winches with 8 post contactors should immediately disconnect the contactor wires from their ATV battery, and call us immediately to receive a free replacement contactor and installation instructions.
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text:
The persona method is aimed at talking about the users and representing users in the design process. I (and some of my students9 have experimented with having users present in the initial design process and let users act as personas for innovation. I gave a talk about this in Chicago, Sept 2011. These are my slides from the talk at UX Masterclass in Chicago lene nielsen – codesign with personas
Why co-design with personas
Including users in large participatory innovation projects together with professional innovators such as designers, people from marketing, engineers etc puts a strain on the user that might not like to be the focus of attention. With point of departure in two cases, one from business and a student project, I will illustrate and discuss the use of personas as a mean to get users involved in innovation, address their needs, and at the same time be a platform that gives all participants equal involvement.
I aim to present a novel way of using role-playing, immersion, and personas. Two workshops are described and analysed in an attempt to explain one of the ways in which innovation and user participation are addressed and oriented towards acting out the future.
A persona is a fictitious user described with basis in data. The personas method is recognised in IT development within the private sector, but has spread to other areas such as marketing and product development. An example of how the method is used in marketing is the Japanese beer company Asahi Breweries that used personas to strategise the future of its Super Dry beer brand (Browne, Temkin & Geller, 2008). The most common use of personas is for a design team to use the user description to understand and engage in the user’s world in order to create new interaction forms or products that correlate with the users’ needs and contexts. In this use of personas actual users are present in the data, but not in the design process.
The workshops described here are based on 10 Steps to Personas (Nielsen, 2007). Key to the 10 steps are scenarios, that are stories describing the persona’s interaction with an interface or product. As a story, the scenario has a main character, a setting, a goal, it has actions that lead to the goal, and it has obstacles that hinder the way to the goal (Madsen & Nielsen, 2009).
The two cases show how users 1) are able to act as personas and be as creative as professional designers 2) use their understanding of the area in focus to create scenarios both from the perspective of personas that are similar to them, but also from personas that are different from them, because they are familiar with different behaviours within the given design area.
A professional case
Arla Foods amba wanted to innovate within the, until then unknown area of canteens. For the purpose of creating new products from user knowledge an innovation process was created that consisted of: Scientific data gathering. User data gathering – 4 dynamic focus groups, each video filmed. Data analysis. From the analysis a documentary film lasting 30 minutes and two personas were produced. An innovation workshop lasting two days.
The workshop had the following course of events: Introduction to data. A design game using the documentary and focusing on pain points. Presentation of findings in the game. Participatory innovation from personas and scenarios. Presentation and ranking of best ideas.
The participants that innovated were canteen managers, concept developers, persons from marketing, and engineers. All groups had at least one person from each category. Even though the canteen managers came on the second day of the workshop, they entered the groups without hesitation and got engaged in the creative process. It was easy for them to relate to the persona descriptions and they felt on equal foot with the designers.
A student case
The aim of the innovation session was to develop a tool that could support communication between soccer trainers, kids, and parents. Prior to the session, data was gathered from observations and focus groups. From this two personas that had different behaviour and media use were created as well as a number of scenarios that varied in situation and context. The participant was asked to go through all the scenarios from the point of view of both personas, with the intention of creating novel solutions.
The participant, a mother to a child who played soccer, had no problem in switching between the two personas even though only one resembled herself. She was able to drawn on her knowledge of other parents and their preferences and behaviour, but when she acted as the persona that resembled her-self, she often commented on the likeness, how she herself would react, and her own needs.
Browne, J., Temkin, B. D., Geller, S. 2008. Design Persona Best Practices From Japan. Examining How Four Organizations Successfully Use Design Personas. In Forrester Report, September 16, 2008
Madsen S. & Nielsen L. 2009. Using Storytelling to Improve Scenarios. In Proceedings of the IADIS International Conference Information Systems, Barcelona, Spain, February (pp. 25-27).
Medler, B. & Magerko, B. The Implications of Improvisational Acting and Role-Playing on Design Methodologies. CHI 2010: Dance, Dust, and Drama: Designing Design April 10–15, 2010, Atlanta, GA, USA (pp. 483-492)
Nielsen, L. 2007. Ten Steps to Personas, http://www.hceye.org/UsabilityInsights/?p=73

domain: design


Tour the Kress Lofts at For the Child fundraiser
2009-10-20 · By Editor
Tour the Innovative Kress loft Homes at a benefit event for For The Child on Saturday, October 24. Join owners of a variety of lofts in their living space to see and hear how they have creatively turned these open spaces into homes with style and the building into a fabulous place to live and share a sense of community.
Following the modern home tour, guests are invited to enjoy a glass of wine on the rooftop deck while sampling cuisine from Pine Avenue restaurants. Entertainment will be provided by Jazz and R & B artist Eddie Gip Noble.
The Kress Building, located at 100 West 5th Street in downtown Long Beach, was designed in the Italian Renaissance style in 1923 by Thomas Franklin Power. Architect Power also designed early buildings of Loyola Marymount University, Long Beach’s St. Bartholomew’s Church, Blessed Sacrament Church in Hollywood as well as dozens of historic churches and high schools throughout Los Angeles.
The Lofts are a conversion of this old department store and office building into loft condominiums with brick walls, exposed wooden rafters, massive concrete columns and steel I-beams. The exterior of the building has been carefully restored to its original architecture.
The city recognized the dramatic restoration by designating the building as a Historic Landmark.
For the Child has helped young victims of abuse and neglect heal so that they can lead healthy, productive lives for more than 35 years. Proceeds from The Kress Loft Home Tour will support these services.
The benefit event takes place October 24 from 4:00 p.m. to 8:00 p.m. Tickets for the Kress Loft Tour cost $45 and must be ordered in advance. To purchase tickets, visit www.forthechild.org or call For The Child at (562) 422-8472.


Nepal holds highest Cabinet meeting at Mount Everest
Nepal's top politicians strapped on oxygen tanks Friday and held a Cabinet meeting amid the frigid, thin air of Mount Everest to highlight the danger global warming poses to glaciers ahead of next week's international climate change talks.
Published 11:37 pm, Thursday, December 3, 2009
SYANGBOCHE, Nepal - Nepal's top politicians strapped on oxygen tanks Friday and held a Cabinet meeting amid the frigid, thin air of Mount Everest to highlight the danger global warming poses to glaciers ahead of next week's international climate change talks.
The government billed the event as the world's highest Cabinet meeting, and getting the ministers to the mountain safely required extensive planning.
Prime Minister Madhav Kumar Nepal, his two deputy prime ministers and the 20 Cabinet ministers were examined by doctors before boarding helicopters to Kalapathar, a flat area at an altitude of 17,192 feet next to Everest base camp, the jumping point for climbers seeking to scale the peak.
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The Himalayan Rescue Association's Bikram Neupane said the politicians - bundled in thick jackets, windproof gear and woolen hats - all had adequate oxygen levels in their blood and they were in no immediate danger.
The Cabinet met only briefly next to the mountain in an effort to prevent any of the ministers, unused to the heights of the Himalayas, from getting altitude sickness.
The ministers stayed overnight Thursday in the town of Lukla, about 9,180 feet high, to acclimatize to the higher elevation. They then traveled to Syangboche - 12,800 feet high - where they took the helicopter to the mountain.
They were expected to return to Syangboche later to hold a news conference.
The event came ahead of the international climate change conference next week in Copenhagen, Denmark, and was intended to draw attention to the effects of global warming on the region surrounding the world's highest peak.
Scientists say the Himalayan glaciers are melting at an alarming rate, creating lakes with walls that could burst and flood villages below. Melting ice and snow also make the routes for mountaineers less stable and more difficult to follow.
"The hills and mountains used to be covered with snow even during the summer. But now snow can be seen only on the higher peaks," said Ngyendon, 66, who like many in the region uses only one name.
Local residents welcomed their guests with cream-colored silk scarfs used for auspicious occasions.
"We are glad the government is taking initiatives before it is too late. Usually authorities tend to act only after accidents. We are now hopeful that something might be done," said Mingma, a 47-year-old inn keeper at Syangboche.
There are no climbers during the winter season and most climbing is done during the spring. Kalapathar is considered the best spot to view Everest.


text:
Sizeable Single Level Home
Well located in Isabella Plains and surrounded by excellent amenities this four bedroom ensuite family home presents a spacious floorplan and offers modern finishes throughout. With two separate living areas that include a generous lounge room and additional family plus meals area that incorporates the impressive, fully equipped galley style kitchen this is a home that promotes a family friendly atmosphere with plenty of room to spread out and come together within.
Four bedrooms including the segregated master, which sits towards the front of the property and contains a generous robe and ensuite facilities, is ideally positioned within the home for those who enjoy a little privacy. The additional bedrooms share the main bathroom facilities with sleek modern design and inclusions. Along with the additional wet areas these rooms all flow naturally off an elongated hallway. The bedrooms also include built in robes as well as good natural light.
Internally the home has a warm welcoming feel with timber laminate floors flowing freely throughout, crisp white walls and various window treatments included. Ducted gas will keep you warm through winter and the home also has solar panels.
The clever combination of family, meals and kitchen areas deliver a family friendly open plan style of living as well as direct access out to the rear yard and the elevated timber entertainment deck. Here entertaining will be easy whether you are catering indoors or outdoors and the natural extension to the internal living areas will add valuable space all year round.
Set on a manageable, level block that currently requires minimal maintenance the home offers side access as well as internal access into the home or out to the yard via the double garage which also has an electronic roller door. Secure from the street and yet accessible when you need it.
Combine all this with a good inner Tuggeranong location where schools of all ages and stages are either within walking distance or a short bus ride away. Convenience stores, medical facilities and recreational areas are also all close by and Tuggeranong CBD is just a few minute’s drive away for those who love a little retail therapy or a good night out.
Available now and ready for immediate inspection this property presents well, offers good internal space and inclusions as well as plenty of possibilities to enhance what already exists.
* Four bedroom ensuite home
* Two separate living areas
* Galley style kitchen and meals
* Ducted gas heating, timber laminate flooring
* Large timber entertainment deck
* Neat presentation and modernised bathrooms
* Double garage with internal access
* Rainwater tank, solar panels
* Manageable easy care block
* Good location close to amenities


text:
World Opinion: Suez Canal traffic jam blocks the world's jugular vein
How That Massive Container Ship Stuck in the Suez Canal Is Already Costing the World Billions of Dollars
$3 billion a day — and that's not factoring in the environmental tollAs backhoes and tug boats worked around the Panama-flagged Ever Giving’s 400-meter-long hull on Thursday evening, experts began to tot up the economic and environmental ramifications of a protracted obstruction. Meanwhile, vessel tracking data showed that some container ships had already started redirecting around the African Cape, a route that can add two weeks of journey length.
The Suez Canalon Wednesday. A 59-foot-wide container ship called the Ever Given of the 120-mile-long Suez Canal. Traveling at a speed of 13 knots (15 mph), the 1,300 foot, 200,000-ton ultra large container ship came to a sudden stop with her bow aground in Asia and her astern at rest in Africa, creating a blockage in a waterway that sees 12% of the world commerce pass through it every year. As of Thursday, tugboats are still working to dislodge the Ever Given.
This is not a story you hear every day; maritime logistics do not cross most people's minds often, let alone wind up in the national news. But they are nonetheless pivotal to global trade. This story provides a stark reminder of the tenuous nature of our maritime global supply chain and the dangers to maritime choke points.
Tugs and dredgers try to free megaship blocking Suez Canal
Tugboats and dredgers were working Friday to free a giant container ship blocking Egypt's Suez Canal for a fourth day, forcing companies to re-route services from the vital shipping lane around Africa. The MV Ever Given, which is longer than four football fields, has been wedged diagonally across the entire canal since Tuesday, shutting the waterway in both directions. The blockage has caused a huge traffic jam for more than 200 ships at either end of the 193-kilometre (120-mile) long canal and major delays in the delivery of oil and other products.
The closure of the Suez Canal has massive ramifications. Every day,traverse this waterway. On average, waiting to embark on their day-long passage. As Ever Given remains firmly across the canal, shipping companies will have to fathom the option of sending ships around Africa, adding as much as two weeks to their journey. All of this means a slow down in the delivery of goods, fuel and essential material between Asia and Europe. Factories depending on parts from Asia may have to close, and essential goods and products to battle Covid-19 may be inaccessible. Fuel prices, already high in Europe, could .
A failure of machinery, human error or natural events -- high winds and reduced visibility -- may have caused Ever Given toin the Suez Canal.
Five things to know about the cargo ship blocking the Suez Canal
A massive, 1,300-foot cargo ship remains stuck in the Suez Canal after running aground nearly a week ago and becoming wedged sideways in the waterway, blocking all traffic through the vital shipping lane and causing major traffic jams in the Mediterranean and Red Seas.It soon became clear that major trade delays were on the horizon and would grow with each day the ship wasn't freed.Experts and officials are exploring multiple options to refloat the skyscraper-length vessel, but as of Sunday afternoon its bow section remained wedged in sand and rock.Here are five things to know about the grounded ship and the broader issues it is creating around the world:1.
But its impact will resonate far from its banks as it has blocked the jugular of one of the largest trade routes in human history.
The Suez Canal was opened into great fanfare after 10 years of excavation. A joint endeavor by France and Great Britain, it provided a shorter route to Asia than having to circumnavigate Africa. Connecting the Mediterranean Sea and the Indian Ocean via the Red Sea, the vital waterway was essential to maritime commerce. In the First World War, Turkish forces tried to attack the canal from the east, and in the , General and the Afrika Korps aimed to do the same from the west.
In 1956, after the war and during the height of anti-colonialism, Egyptian President Gamal Abdel Nasser nationalized the canal, precipitating thewith Israel, France and Great Britain. This resulted in its closing until . Then, in June 1967, the the Suez Canal becoming the border between a warring Egypt and Israel . The outbreak of the conflict found more than a dozen ships -- the Yellow Fleet -- caught in the canal and trapped for the duration. The crews of this international fleet formed the "Great Bitter Lake Association," and even competed in their own version of the Olympics. The ships remained until 1975.
How the Giant Boat Blocking the Suez Canal Was Freed: Dredgers, Tugboats, and a Full Moon
How a celestial body contributed to the rescue effortsThe recovery vessels took advantage of high spring tides around the full moon on Sunday to free the Ever Given, which had blocked Egypt’s Suez Canal for almost a week. Its partial refloating just before dawn on Monday drew cheers and foghorn blasts from the bridges of other vessels caught in the bi-directional snarl that had held up hundreds of ships and billions of dollars worth of cargo each day since March 23.
This closure forced a revolution in maritime transportation with the creation of-- very large crude carriers. With the canal closed, ship designers built ships that were too large for the Suez Canal and had to sail around the Cape of Good Hope. This economy of scale in transportation has evolved from supertankers to mega cruise ships and container ships such as Ever Given. In 1956, American developed the concept of intermodalism. Instead of individual goods being loaded into a truck, then offloaded at a port, and then loaded into a ship and the process reversed, he developed the modular shipping container, which allowed for containers filled with goods to be transferred, streamlining the process. When SS Ideal X sailed from Newark, New Jersey, to Houston, Texas, with 58 containers on its deck, McLean could not have envisioned a ship the size and scale of Ever Given.
By 2020, containerized cargo made up nearlyshipped on the world's oceans annually. The movement of goods by container has continually grown except during the economic recession of 2008 and the tariff fight between China and the United States along with the Covid-19 crisis in 2020. One of the main arteries for this trade runs between Eastern Asia and Europe.
Egypt has opened a formal investigation into how the Ever Given got stuck in the Suez Canal
A Suez Canal Authority advisor told Reuters that authorities plan to focus on the state of the ship and the captain's actions before the grounding.Captain Sayed Sheaysha, an advisor to the Suez Canal Authority, said that experts would board the ship on Wednesday to try and gather information, the BBC reported.
The 23 million containers that moved between Asia and Europe in 2020 sailed through some of history's most contested waters, such as the Strait ofand Malacca Strait, , the Straits of and the most relevant today, the Suez Canal.
Great Britain controlled all these maritime choke points during its imperial height. It allowed Britain to influence ocean commerce and trade. Today, many of these are patrolled and regulated by international conventions and patrolled by navies, but the Suez Canal is wholly owned and controlled.
Suez Canal Ship Update as Ever Given Blockage Backlog Set to Finally be Cleared .
The last remaining 61 ships of the 422 that were caught in a backlog after the Ever Given got stuck will finally be able to continue with their journey.Officials at the Suez Canal Authority (SCA) said the last ships that were stranded after the massive vessel would pass through today, according to Reuters.

domain: geography


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Pandemic hurt women workers more than men
Last year, the number of employed women in the country declined by 137,000 on year, according to a Friday government report, far more than the 82,000 male jobs lost.
According to a survey by the Korean Women’s Development Institute published Monday, one out of five women who participated in the survey lost her job after the Covid-19 outbreak. A total of 3,007 female workers aged between 20 and 59 participated in the survey.
Among survey respondents that lost their jobs or were forced to take leave during the pandemic, four out of 10 said female workers, pregnant employees or workers on parental leave were among the first in their workplace to be affected by layoffs or leaves.
“This proves that restructurings were gender-discriminative, as in previous economic recessions,” the report said.
The survey showed that women in their 20s and 30s had a higher percentage of losing jobs during the pandemic. Nearly 30 percent of respondents in their 20s and 19.5 percent in their 30s said they became jobless after the Covid-19 outbreak. The percentages for women in their 40s and 50s age group were smaller at 18.4 and 18.5 percent, respectively.
Both reports pointed out that female workers were more prevalent in service industries like retail, food and education that typically involve face-to-face interaction. That made them more vulnerable in a downturn.
Because these industries are filled with small companies, it was more likely that employers wouldn’t offer paid leaves or pay unemployment benefits—the kind of support common at larger, more established firms.
Cultural norms also played a role.
“As schools and kindergartens shut down, women mainly took on the responsibility of childcare and this led to situations where they voluntarily chose to leave the workplace,” said the government report.
“At a time when digital businesses are growing, the proportion of women that graduate with engineering degrees in this country remains small— and that’s holding back an increase of jobs for female workers.”
The Federation of Korean Industries (FKI), a lobbying group for local companies, stressed the importance of increasing the female workforce, saying it could be a core solution to Korea's shrinking population and aging society. Korea’s fertility rate reached a new low last year of 0.84.
According to FKI’s analysis of OECD statistics, Korea was to have the highest old-age support ratio among OECD countries by 2080. And yet, its female labor participation rate was lower than the OECD average by 5 percentage points at 60 percent.
The female labor participation rate fell drastically from 71.8 percent in the age group of 25 to 34 years, to 62.9 percent in the 35-to-44-year age group. The FKI pointed out that career interruptions due to marriage and having children was the biggest obstacle to female workers keeping their positions at work.
“Cases where female labor participation is currently high -- like England, Germany, France and Sweden -- show that fertility rates dropped in the first few years when the female workforce started going up,” the FKI report said.
“But as working conditions for women improve and the female labor participation rate surpasses 60 percent, it showed that fertility rates started going up again. Statistics show that an increase in economically active women can contribute to birth rates and population.”
In a Monday message marking International Women’s Day, President Moon Jae-in remarked Korea’s reality is “embarrassing” when it comes to female leadership.
“Recovery and growth will be accelerated once women in this country get more jobs without career interruptions,” he said. “The government will lead efforts to be an exemplar case.”
The government has dedicated a portion of an extra budget to create 77,000 jobs for female workers this year. That bring the goal of female job positions newly created this year to 780,000.
BY SONG KYOUNG-SON [email@example.com]

domain: sociology


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An eight-storey apartment block, which will contain 88 flats has been approved despite not offering a single parking space. The new building is due to be built on the site of the old BT office in the Old Town South Conservation Area. It will house 25 studio flats, 26 one-bed flats and 37 two-bed flats.
The site currently has a car park, but the developer plans to remove it as they want a ‘ car-free development’. Residents Karen and Gary Callaway are two of 15 nearby residents who have objected to the proposal, which was discussed in yesterday’s (22nd November) Planning and Right of Way Panel.
They said: “ Car parking is (a) big problem for the residents in 1 Briton street. There’s already a shortage of parking spaces within Gloucester Square and whilst there may be other space available in another surface car park, this really isn’t desirable especially for the young and older families with children having to carry shopping, buggies, (and those with) disabilities.”
Planning officers at the council say that having a ‘car-free development’ is ‘acceptable’. “Car-free developments are deemed to be acceptable in the city centre, and comply with our adopted maximum parking standards due to the wide range of services, employment and local transport connections on offer.
“Having regard to the nature of the proposed uses and the city centre location of the site, a car-free approach is considered to be consistent with other high density residential schemes of this nature in the city centre.” Other concerns on the new block include the height of the building ‘ruining the sea view’ of residents in other flats, ‘breaking an already broken system’ by raising demand for strained nearby doctors surgeries, and historic murals being forced to be taken down and ‘stuck on the back of benches’.
Telereal Trillium, the company developing the apartment block, listed the positives for the local area. These include:
- New tree planting
- Replacing old paving
- New concrete benches
- New feature lighting
They say that the space that is being made available for all of these improvements is only possible because they are making the building taller, and therefore, can ‘take from the sides’. Stuart Brooks, the council’s planning officer, who recommended the development for approval spoke about how the new complex will also help to reduce crime rates in the area by offering ‘natural surveillance’
He said: “People would be more put off committing crimes because there are people in the area.” However, this notion did not sell to all the councillors on the panel, as Councillor John Savage disagreed and said: “You don’t get rid of the problem by moving people in; you just increase the number of victims.”
Despite the numerous concerns raised, councillors, all apart from Cllr Savage, opted to follow the planning officer’s recommendation and the proposal was approved. It has now been delegated to the planning officer so that, providing minor conditions are met, the development will be under way.
- Man charged with murder after woman's body found in Southampton home


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Letter: CV Trail attracts many to region
To the editor:
Goodhue County, Cannon Falls and Red Wing governments, I request that you increase financial support for Cannon Valley Trail.
This trail is a real gem for riders and walkers of all ages. The trail's popularity extends well into Wisconsin and the Twin Cities.
Many of the folks I meet when biking also overnight in this area. They come to enjoy the natural beauty, seclusion and ease of access to a well maintained trail.


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