Green Boxes Research Articles

Accurate Fully Automated 4D Segmentation of Osteoarthritic Knee MRI

Purpose: A method for 4D (3D+time) segmentation of bone and cartilage surfaces of the knee joint imaged by magnetic resonance (MR) is reported. Accurate segmentation is necessary to understand osteoarthritis (OA) disease processes and quantify the efficacy of DMOAD (disease-modifying OA drug) drug trials. Manual segmentation is tedious, time-consuming (several hours per joint) and irreproducible. We have previously reported an automated 3D approach based on layered optimal graph segmentation of multiple objects and surfaces (LOGISMOS). This study extends 3D LOGISMOS to 3D+time (4D) LOGISMOS to simultaneously segment four cartilage and bone surfaces (femur, tibia) in longitudinal sequences of follow-up MRI's. The key benefit of using our automated 4D LOGISMOS method is that information from all time points of the temporal sequence contributes to the single optimal solution that utilizes temporal and spatial context between adjacent time points. Methods: Our 4D LOGISMOS approach consists of two major steps: a) Constraining topological relationships between identical pre-segmentation meshes between time-points using rigid registration, and b) enforcing physiologically permissible maximum changes between time-points by introducing inter-time point graph arcs that longitudinally link the temporally corresponding locations of the respective bones and cartilages of the femur and tibia. Robust hierarchical random-forest classifiers were used to derive cartilage cost functions trained using segmentation examples. Once segmented, automated sub-plate analysis was used to identify segmentation accuracy in medial/lateral regions of the femur (cMF/cLF) and tibia (cMT/cLT). To assess segmentation accuracy, 54 patients at baseline and 12-month follow-up visits from the OAI (Osteoarthritis Initiative) progressive cohort were analyzed (108 3D MR DESS datasets) and compared with the manually-traced independent standard (IS). To assess the temporal consistency of the segmentations, a large sub-cohort of 399 patients with five yearly follow-ups (baseline, 12, 24, 36, 48 month; 1,995 DESS MRI’s) were analyzed to show the benefits of 4D over 3D analysis. Since no independent standard was available for this cohort, consistency assessment was based on a hypothesis that cartilage thickness losses over the five-year period would be monotonically decreasing and that this hypothesis can be tested using longitudinal cartilage thickness correlation coefficient R. Higher occurrence of analyses satisfying this hypothesis indicates a larger success rate – histograms of R-values were analyzed to determine whether 4D LOGISMOS outperformed the 3D approach. Results: Tables show signed and unsigned cartilage border positioning errors using the 4D and 3D methods at baseline and 12 months with green boxes representing significant improvement while red boxes indicate a worsened performance. Analyzing histograms of R-values for different sub-plates showed a significantly (p<0.05) better performance of the 4D LOGISMOS when compared to that of 3D.Table 1Baseline signed and unsigned cartilage surface positioning errors between 4D and 3D LOGISMOS on the major femoral and tibial sub-plates.Table 212-month signed and unsigned cartilage surface positioning errors comparison between 4D and 3D LOGISMOS on the major femoral and tibial sub-plates.Table 212-month signed and unsigned cartilage surface positioning errors comparison between 4D and 3D LOGISMOS on the major femoral and tibial sub-plates. Conclusions: The proposed 4D LOGISMOS segmentation algorithm outperformed the 3D approach and demonstrated the benefits of using temporal contextual information in studies with multiple follow-up MRIs. Temporal-context aware image segmentation techniques offer a significant improvement of segmentation performance in hard-to-analyze longitudinal studies of OA progression.

Genome Skimming: A Rapid Approach to Gaining Diverse Biological Insights into Multicellular Pathogens.

Genomic data acquisition is now trivial for biologists. Yet, moving from millions of sequence reads to an assembled and annotated genome continues to pose a daunting challenge. The first animal genome sequenced arose from the free-living model nematode Caenorhabditis elegans [1]. This venture provided an unprecedented foundation for new insights into genome function and ‘omics tool development. However, the C. elegans endeavor has been tough to repeat, even with the advent of new high-throughput DNA sequencing technologies. For example, the first plant-parasitic nematode (PPN) genomes were published ten years after the C. elegans genome [2,3], and only five publication-quality PPN genomes are presently available [4–6]. Fig 1 overviews the course of a typical genome project. Millions of DNA sequences are initially collected in a matter of days, thanks to new DNA sequencing technologies. Early analytical phases (quality control and initial assembly) are also quick and usually straightforward. However, the subsequent computational stages (refining the assembly, gene prediction, and annotation) present significant bioinformatics bottlenecks. These lengthy in silico steps require multiple iterative stages of analysis, finally leading to a finished genome deemed “good enough” for publication. These latter stages often take years. Open in a separate window Fig 1 Genome skimming schematic. Boxes progressing diagonally from top left to bottom right show steps typical of conventional genome projects. Grey boxes show steps shared by genome skimming and conventional genome projects. Red boxes, arrows, and Xs show conventional genome project steps eliminated in the genome skimming approach. Green boxes show analyses specific to our genome skimming strategy.

219 - Effect of Co-Medications and Endotoxins on Mesenchymal Stem Cell Characteristics

The impact of immunosuppressive medications and endotoxemia on mesenchymal stem cell (MSC) therapy have not been established. We examined the effects of calcineurin inhibitors cyclosporin (Cyc) & tacrolimus (Tac), corticosteroids hydrocortisone (Hyd) & dexamethasone (Dex), rapamycin (Rap), mycophenolic acid (MPA) and lipopolysaccharide (LPS) endotoxins on bone marrow MSC secretion of cytokines and growth factors that contribute to MSC activation, anti-inflammatory and regenerative functions. We used Luminex bead platform to assess expression of 41 cytokines (Table 1). We showed secretion of cytokines that activate MSC such as IFNγ and TNFα was increased in response to all compounds tested. However, other cytokines with similar effect such as IL-1α and IL-17 were largely unaffected. Factors known to play a role in MSC-induced immunosuppression and cell differentiation such as IL-10, EGF and TGFα were variably affected. LPS and Dex increased secretion of IL-10, while effects of other medications were concentration dependent. TGFα secretion was increased by LPS and MPA with variable effects induced by other drugs. EGF secretion was largely decreased, yet was increased by MPA and Tac at high concentration. We evaluated changes in secretion of factors known to induce angiogenesis, neurogenesis, hematopoiesis and epithelial proliferation such as VEGF, IL-6, GM-CSF and G-CSF. LPS, Tac, and Cyc exposure increased secretion of VEGF-A. Contrarily, corticosteroids and Rap (except 1 ng/ml) decreased VEGF-A secretion. IL-6 secretion was increased by LPS, Rap, Tac and Cyc and decreased by MPA and corticosteroids. With few exceptions, LPS increased and immunosuppressive drugs decreased GM-CSF secretion. G-CSF was increased by LPS, but unaffected by the drugs evaluated. In summary, MSC characteristics are influenced by immunosuppressive medications and endotoxins. Further studies are necessary to evaluate how these variations might affect the intended therapeutic function of MSC. Table 1(a) Drugs and LPS were added to BM-MSC culture medium in three concentrations. Analyte concentrations after 72 hours in culture were compared with concentrations in the control wells containing 0.2% DMSO only. Increases over the control concentration are denoted in red, decreases are denoted in green and white indicates there was no change in analyte concentration between the control and experiment. (b) Red boxes indicate that all three drug/LPS dilutions tested resulted in an increase in analyte concentration. Green boxes indicate that all dilutions produced a decrease in analyte concentration. White boxes indicate that there is variability among the three concentrations as to whether an increase, decrease or no change was observed.

Policy Space in Agriculture Under the WTO Rules on Domestic Support

A concept of policy space is formulated under the domestic support rules of the WTO Agreement on Agriculture. This policy space is dyadic, comprising two kinds of space: exemption space and limited space. The two kinds are available differently for different countries. Exemption space derives from the entitlement to exempt policies that meet given criteria from those policies under which support counts against the country’s limit(s), i.e., exemption space allows limitless support. The criteria are those of Annex 2, Article 6.2 and Article 6.5 of the Agreement (green, development and blue boxes). Limited space derives from the entitlement to provide support under non-exempt policies up to the country’s limit(s). For most countries the limits are the de minimis levels calculated from the value of production of each product and of agriculture as whole, using the country’s de minimis percentage. For 32 countries (15 developed and 17 developing) the limit is the Bound Total Aggregate Measurement of Support (BTAMS) and the de minimis levels are thresholds above which support counts against the limit. Some acquisition of foodstuffs at administered prices is subject to special rules, now subject to negotiations with implications for both exemption space and limited space and the interaction between these two kinds of space. The Agreement’s criteria are reviewed to interpret the factors determining a country’s exemption space. In determining limited space, the centrality of a country’s values of production is illustrated by tracking limited space from 1995 to 2012 for the EU, the US, India and China under assumptions of perfect management of maximum support. China’s rapidly rising values of production have generated limited space that now greatly exceeds that of the EU. The US limited space has been rising but India’s limited space is growing faster and approaches the US level. In recent years these four countries have used only a small part of their limited space. As values of production rise in individual countries and in the world, the relative importance of BTAMS as a component of limited space declines since it stays fixed in nominal terms. The amount and the share of non-exempt support that can be accommodated within the de minimis provisions are thus increasing for individual countries and in total for the world. Some countries with declining values of production in agriculture are the exceptions, e.g., Japan. The different entitlement of different countries to limitless exemption space, in combination with limited space growing faster in some countries than in others, means that the picture of countries’ policy space for domestic support to agricultural producers is rapidly changing from the picture when the Doha negotiations started.

Potential of Group Incentive Schemes to Encourage Use of Driving Safety Apps

Encouraging young drivers to use driving feedback systems (referred to here as “green boxes”) is a challenge. Prior research and experience have indicated that adoption of green boxes has been minimal, primarily because of insufficient motivation. Research has also demonstrated that providing feedback from green boxes to young drivers or their parents has reduced the frequency of risky behavior. This study investigated the possible benefits of providing group incentives to encourage the use of green boxes among young drivers. The specific green box used in this study was a smartphone app that needed to be initiated at the beginning of each trip. Once initiated, the app monitored driving behavior in terms of g-force events, provided a score and feedback, and shared the driving information with a predefined sponsor. This study built and expanded on a previous study by making the duration of the experiment longer and by allowing participants to recruit their friends to help the group win its rewards. Despite the more demanding scheme, the results obtained repeated the success of the previous study: all eligible participants downloaded the app and used it to win the rewards for the group. Additionally, friends were recruited by participants to use the app to help the group win rewards (without acquiring any personal rewards for themselves). Once all prespecified rewards were achieved within the allotted period, the young drivers stopped using the app. This study confirms again that barriers to adoption of the green box app by young drivers can be overcome by choosing low-cost, suitable group incentive schemes.

DCIs, SEPs, and CCs, Oh My! Understanding the Three Dimensions of the NGSS

The Next Generation Science Standards' three dimensions--disciplinary core ideas (DCIs), science and engineering practices (SEPs), and crosscutting concepts (CCs)--were headliners at NSTA's national conference in Chicago and featured in many of the organization's other professional-development efforts this year (NGSS Lead States 2013). To some, the idea of DCIs, SEPs, and CCs may seem obvious and not all that new (Figure 1, p. 68). Haven't we been doing science inquiry since the release of the National Science Education Standards (NSES) in 1996? What is new and different about the NGSS, and what do these three dimensions mean for curriculum, instruction, and assessment? [ILLUSTRATION OMITTED] There are, in fact, several major differences in how the new standards portray science education. These differences call for substantial shifts in terms of our learning goals, instructional strategies, and assessment. In the next sections, we highlight what is new and different about each dimension. We do not address explicitly intertwining the three dimensions; a forthcoming article will discuss their integration. Much of what we discuss below draws directly from A Framework for K--12 Science Education (NRC 2012), which guided the development of the NGSS and is the foundation for the new standards. While the NGSS do provide some information from the Framework (in the blue, orange, and green boxes; see Figure 1) to elaborate on the performance expectations, these are only snippets of the more elaborate descriptions found in the Framework. We therefore urge readers to turn to the Framework when trying to understand the three dimensions and the standards. DCIs There are DCIs for each of the four major disciplines: physical sciences, life sciences, Earth and space sciences, and engineering (engineering, technology, and applications of science). Each of these disciplines includes no more than four DCIs, reflecting a concerted effort to cull the numerous ideas that all students are expected to know. While there are somewhat fewer ideas to teach, each is complex, with ample depth to delve into over the course of schooling. To rise to the level of a DCI, an idea must meet four criteria. First, it must be a key organizing principle within the discipline or across several disciplines; that is, it should be a core idea in the eyes of scientists. Second, it must have broad explanatory power: It should help learners understand and be able to reason about an array of phenomena and problems in the discipline. In this sense, it needs to be a useful thinking tool that is generative for students, and it should help them think about phenomena and problems they may encounter in and out of the classroom, both now and in their future. Third, a DCI needs to be relevant and meaningful for students. It should relate to phenomena and problems that students find intriguing. Fourth, the idea needs to have depth that allows for continued learning over the course of schooling. There are two complementary implications of this last point. First, the DCI, in some basic form, must be accessible to young learners, and, second, it must have enough complexity that it can be unpacked and deepened in higher grades. Many of the concepts (e.g., ionic bonds, mitochondria) and even topics (e.g., volcanoes, taxonomy) currently taught in school do not meet these criteria. As educators, it is incumbent upon us to closely evaluate what we are teaching and to dramatically prune the unwieldy tree of concepts we try to cover. Taken together, the DCIs create a conceptual toolkit that students can use to reason about and explain phenomena. The focus on explaining phenomena represents an important shift in the goals of instruction. Rather than teaching ideas in the abstract or in isolation, the new aim is to engage students in using these ideas to explain interesting phenomena. For example, instead of having students describe the water cycle and its components, students should be explaining cloud formation or precipitation patterns by using understandings about the water cycle and thermal-energy transfer to describe how weather events come about. …

Identification of small ORFs in vertebrates using ribosome footprinting and evolutionary conservation

Identification of the coding elements in the genome is a fundamental step to understanding the building blocks of living systems. Short peptides (< 100 aa) have emerged as important regulators of development and physiology, but their identification has been limited by their size. We have leveraged the periodicity of ribosome movement on the mRNA to define actively translated ORFs by ribosome footprinting. This approach identifies several hundred translated small ORFs in zebrafish and human. Computational prediction of small ORFs from codon conservation patterns corroborates and extends these findings and identifies conserved sequences in zebrafish and human, suggesting functional peptide products (micropeptides). These results identify micropeptide-encoding genes in vertebrates, providing an entry point to define their function in vivo.

Sovent-Free Synthesis of Lithium Intercalated Graphites Using Thermal or Mechanical Reactors

The intercalation of lithium into graphite is an important process for energy storage devices. The industrial route of intercalation is electrolysis in pre-assembled Li-ion batteries with excess Li loading in the cathode. This has a number of disadvantages: increased battery processing time, mass and volume (from needing excess Li loading), and parallel formation of a solid-electrolyte interphase (SEI). The later is difficult to control and is a major factor in the battery's overall performance and cost. An alternative is to prepare the anode with partially lithiated graphite prior to electrolyte contact. As far as we know this has not been demonstrated. Each step – lithiation of graphite followed by making slurry with binder – presents added complications with the reactive anode powder, but it offers compelling advantages such as, increase in "usable" Li, control of SEI formation, and decreased battery processing time. These advantages may also lead to battery performance and lifetime. Also, it would also allow for investigations into spontaneous formation of SEI, which may provide insight into producing an effective artificial SEI.As reported in the literature, LiC x has been chemically synthesized viacontact with lithium vapor and compression with lithium metal. Both of these has a flaw: graphite reaction with Li vapor is kinetically unfavored due to the low vapor pressure of Li and it is difficult to control the stoichiometry; the compression method has been shown to produce the important stages (I–IV) but it is slow, taking 24 h. Here we present two solvent-free and compression-free methods that can produce stage I or stage II lithiated graphites. The thermal method heats graphite and lithium powders to ca. 300 °C under ultrahigh vacuum (UHV) for 24 h. The mechanical method involves ball-milling the two powders together under ultra-high purity Ar(g) atmosphere for 1 h.Both processes work with different types of graphite sources and we compare the two synthetic methods using a battery-grade mesophase graphite. The products were characterized by x-ray diffraction (XRD) and x-ray photoelectron spectroscopy (XPS). We show that while the mechanical reactor seems to lithiate the graphite more quickly, it may affect the morphology. Hard carbon fibers were also lithiated using the mechanical reactor. We compare the stability of the LiC6made using mesophase graphite and carbon fiber.XRD and NMR are used to follow the delithiated of the products when in contact with a non-zero moisture environment. It was found that a surface passivation layer is formed that protects intercalated lithium. We also report open-circuit potential (OCP) and electrochemical impedance spectroscopy measurements of the products in contact with standard Li-ion battery electrolyte. These results are used to discuss the spontaneous formation SEI at the LiC6interface. Figure 1: SEM images of a) stabilized lithium metal powder; b) carbon fiber, Pyrograf I; c) MGPA; d) ball-milled lithiated-MGPA. e)X-ray diffractogram of mechanically synthesized lithiated MGPA. Inset is log-scaled expansions of regions in green boxes. (o) raw graphite; (*) LiC12; (†) LiC6.Research was supported by the Fluid Interface Reactions, Structures, and Transport (FIRST) Center, an Energy Frontier Research Center funded by the U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences (RLS, NJD). XPS analysis was done with support of the U.S. Department of Energy, Office of Basic Energy Sciences, Materials Sciences and Engineering Division (GMV). The authors thank Applied Science, Inc. for samples of the Pyrograf I carbon fibers and Pred Materials International for the MGPA.

Population Pharmacological Profiles Of AML Treatments In Patient Samples By Automated Flow Cytometry; A Bridge To Individualized Medicine

BackgroundTo aid in the identification of effective treatments for individual patients, ex vivo assays for detecting cell death inducible by drugs for hematological malignancies have been in development for over 20 years. We have developed a novel approach incorporating 4 key innovations; incubating drugs in whole bone marrow sample without isolating leukocytes, using flow cytometry enables identification of the malignant cells selectively, an automated flow cytometry-based platform (ExviTech) decreases errors and enables full pharmacological characterization, and analyzing the data using pharmacodynamic population models. AimThe purpose of this study is to derive the ex vivo pharmacological profiles across the AML patient population of single drugs and combination treatments as a tool for individualized treatment selection. Patients and MethodsBone-marrow samples from 160 patients diagnosed with AML were sent to Vivia from 24 hospitals across Spain within 24 hrs. The plates were incubated for 48-hours prior to analysis with ExviTech, The percentage of leukemic cell death was determined via labeling with monoclonal antibodies and AnnexinV-FITC. A survival index is computed for each drug, the lower the survival index, the more effective the drug. Dose-response curves of cytarabine, idarubicin, daunorubicine, etoposide, mitoxantrone, fludarabine, clofarabine, and 6-thioguanine were measured in 160 patient samples. The added benefit of combining these drugs into 12 combination treatments was assessed by measuring their synergy in each individual patient. In 39 patients treated with CYT IDA we had clinical data of response, and then we performed a blinded interpretation of this in vitro test by an expert hematologist, to predict the clinical response based in this test result. ResultsThere was a large range of interpatient variability in the response to a single drug and even larger in the synergism between drugs. The Population Pharmacological Profiles for an individual patient is shown on the figure below. The relative drug potency in terms of their percentile ranking within the population is shown in the left panel from 0 (weakest) to 100 (most potent). Green lines represent the individual patient potency relative to the population ranking, with confidence intervals. Third column lists when a drug leaves a significant % of leukemic cells alive, potential resistant clones. The panel on the right side shows the synergism of the drug combinations treatments shown as box-plots at 10-25-75-90% to highlight their distribution. The synergism value for an individual patient in each combination is shown in green, with confidence interval as parallel dotted green lines. This representation of the Pharmacological Profile of an individual patient sample quickly identifies extreme values, when a drug or combination is very sensitive (rightward shift green lines, green boxes) or very resistant (leftward shift green lines, red boxes). This patient showed average sensitivities for most drugs though highly resistant to Clofarabine (red box) that leaves 45% alive. However this patient showed lack of synergism in multiple treatments (right, red boxes). CYT and IDA show average potencies but lack of synergism, suggesting CYT-DAU might be a more efficient treatment. These representations lead to clear guidelines in >90% samples, and based on hematologist's interpretation of these guidelines show a clinical correlation with clinical responses to CYT-IDA of 84%. ConclusionWe have developed an improved a methodology to measure the pharmacological activity of drugs and drug combinations in AML patient samples as well as modeling their pharmacological behavior. This information may be useful in selecting the optimal treatment for the individual patient, especially relapse/refractory patients in need of therapeutic alternatives. By testing the drugs used in the treatment protocols for AML directly on patient samples, a pharmacological based model has been developed to infer drug resistance or sensitivity, patient by patient.▪ Disclosures:Ballesteros:Vivia Biotech: Equity Ownership. Primo:Vivia Biotech: Employment. Hernandez-Campo:Vivia Biotech: Employment. Rojas:Vivia Biotech: Employment. Liebana:Vivia Biotech: Employment. Lopez:Vivia Biotech: Employment. Iñaki:Vivia Biotech: Consultancy. Bennett:Vivia Biotech: Employment.

Drug-Induced Amyloidosis: A Proteomic Insight Into 52 Cases

The amyloidoses are heterogeneous diseases associated with the deposition of insoluble proteins or peptides with a characteristic beta-diffraction pattern extracellularly. At the current time, over 27 different extracellular fibril proteins are known to cause disease in humans. Iatrogenic amyloidosis is a rare, often not sought diagnosis. On occasion, patients with drug induced amyloidosis can present with other systemic features reminiscent of systemic immunoglobulin-derived (AL) amyloidosis, and may present a diagnostic challenge. Treatment of the latter often involves chemotherapy and/or stem cell transplantation, while the former tends to remain localized and needs local therapy; thus accurate diagnosis is critical. To this end, proteomic analysis of amyloid tissue has proven to be an invaluable tool in amyloid typing. We have analyzed the biochemical composition of iatrogenic amyloid using laser capture/tandem mass spectrometry (LC-MS/MS)-based proteomic analysis in 52 cases of insulin and enfuvirtide (Fuzeon¨) associated amyloidosis.In brief, 10-μm-thick sections of formalin-fixed paraffin-embedded tissues were stained with Congo red. Congo red positively staining tissue as viewed with a fluorescent light source appeared bright red. Positive areas were dissected using laser microdissection to a volume of at least 60,000 μm2; three microdissections were analyzed for each case. The microdissected material was collected into 0.5-ml microcentrifuge tube caps containing 35 μL Tris/EDTA/0.002% Zwittergent buffer. Microdissected fragments were subjected to a heat-mediated antigen retrieval method (98C for 90minutes) before being denatured via sonication and subsequently digested into tryptic peptides overnight using 0.5ug of trypsin. The resulting digests were then analyzed with nanoflow LC-MS/MS. The MS/MS spectra of each case were matched against a composite protein sequence database using three different search algorithms (Sequest, X!Tandem, and Mascot). The composite database contained the human SwissProt entries but was also augmented with known immunoglobulin variant domains, known amyloidogenic mutations from literature, the enfuvirtide amino acid sequence, and common contaminants. Reversed protein sequences were appended to the database for estimating the false discovery rates of the identifications. The peptide identification results were filtered using Scaffold software (Proteome Software, Portland, OR) and then filtered peptides were assembled into protein identifications. Candidate proteins with at least one high-confident (probability of identification >90%) unique peptide identification and at least four MS/MS spectral matches were considered for clinical interpretation.For each case, we created a personalized proteomic profile that lists all the confident protein identifications in each of the microdissection along with their respective MS/MS spectral counts. The number of MS/MS spectra matching to a protein is considered as a semi-quantitative measure of its abundance. The most abundant amyloidogenic protein detected across all microdissections and as interpreted in the context of the clinical history is considered to be the amyloid subtype.Figure 1 shows the results of insulin amyloidosis. Figure 2 shows the results of enfuvirtide amyloidosis. Amyloid deposits are shown to be composed of the recombinant drug in addition to amyloid precursor proteins such as apolipoprotein A-I, A-IV, E and serum amyloid P (SAP). [Display omitted] [Display omitted] In conclusion, we show the biochemical composition of all known drug-induced iatrogenic amyloidosis and provide the utility of proteomic analysis in elucidating amyloid subtyping for accurate diagnosis and management.Legend: A spectral count number of greater than 4 is significant. The green boxes denote protein identification at a probability of over 95%, and yellow 80-94%. In figure 1, insulin (# 2) and in figure 2, enfuvirtide (#5) are shown in abundance, additionally other amyloid precursors such as apolipoproteins A-IV, E, A-I and SAP are also seen. Disclosures:No relevant conflicts of interest to declare.

Looking for CDKN1C enhancers

Looking for CDKN1C enhancers

Dynamic signaling of facial expressions transmit social information in a hierarchical manner over time

Designed by both biological (Susskind et al., 2008; Andrew, 1963) and social evolutionary pressures (Darwin, 1999/1872), facial expressions transmit specific sequences of information across time to achieve an optimal system of social signaling and decoding (Darwin, 1999/1872; Schyns et al., 2009). Here, we show for the first time that this dynamic signaling transmits social information over time in a hierarchical manner that fits evolutionary pressures. We proceeded in two steps. First, we used 4-dimensional (3-dimensional face over time) reverse correlation to reconstruct the dynamic mental models of the 6 basic facial expressions of emotion, for 60 Western Caucasian observers individually (Yu et al., 2012; Jack et al., 2012. See Figure S1, Panel A). Second, from the distribution of mental models, we constructed a 6-choice Bayesian classifier (see Bayesian Classifiers in Supplementary Material) that accrues Action Unit (AUs; Ekman et al., 1972) signals over time, as a function of their availability. Analyses revealed that early facial signalling leads to systematic confusions (p<.01. See Figure S1, Panel B, pink boxes) between fear and surprise (and also disgust and anger), due to specific shared AUs (eye lid raiser and nose wrinkler, respectively. See Figure S1, Panel C, pink boxes). Accurate discrimination of each one of the six basic emotions is delayed by the later availability of diagnostic AUs (e.g., eye brow raiser and upper lip raiser, respectively. See Figure S1, Panel C, green boxes). We conclude that the face transmits social signals in a hierarchical manner that fits evolutionary pressures: Initially, the eye lid raiser and the nose wrinkler signal two types of danger (Susskind et al., 2008; Lee et al., in press; Chapman et al., 2009), which, with the addition of diagnostic AUs, are distinguished further into the six basic emotions (e.g., surprise) and their subtypes (e.g., positive vs. negative surprise).

Image cytometer method for automated assessment of human spermatozoa concentration

In the basic clinical work-up of infertile couples, a semen analysis is mandatory and the sperm concentration is one of the most essential variables to be determined. Sperm concentration is usually assessed by manual counting using a haemocytometer and is hence labour intensive and may be subjected to investigator bias. Here we show that image cytometry can be used to accurately measure the sperm concentration of human semen samples with great ease and reproducibility. The impact of several factors (pipetting, mixing, round cell content, sperm concentration), which can influence the read-out as well as inter-operator and -cytometer variation on two different image cytometers (NC-3000 and SP-100) were evaluated. Furthermore, 725 semen samples were assessed both by manual assessment (WHO recommended method) and by image cytometry and tight correlations between the measured concentrations were shown. Moreover, by evaluation of repeated measurements it appeared that image cytometry produced more consistent and accurate measurements than manual counting of human spermatozoa concentration. In conclusion, image cytometry provides an appealing substitute of manual counting by providing reliable, robust and easy measurement of human sperm concentration.

Three-Dimensional Computed Tomographic Analysis of a Rare Left Coronary Artery to Left Ventricle Fistula

A 16-month-old child with a known large coronary fistula between the left coronary artery and left ventricle was referred for computed tomographic (CT) coronary angiography to further characterize the anatomy in anticipation of transcatheter device closure. The diagnosis of this rare coronary anomaly was made initially by echocardiography in the newborn period, and over time, the left ventricle had become dilated with evidence of diastolic dysfunction. A CT coronary angiography was performed using a dual-source CT scanner (Somatom Definition; Siemens Healthcare, Forchheim, Germany) with the child under general anesthesia. Postprocessing was performed on an Aquarius 3D Workstation (TeraRecon Inc., San Mateo, CA, USA). The maximum-intensity projection image shown in Fig. 1 demonstrates the course of the fistula (yellow arrow) as it originates from a dilated left main coronary artery and drains into the left ventricle. The straightened planar reformatted image and its accompanying graph shown in Fig. 2 provide precise measurements of the cross-sectional diameter of the fistula along its course. The yellow and green boxes show cross-sections of the fistula in exact orthogonal planes and exhibit the vessel’s circumferential irregularity. Rotation of the curved planar reformatted image, as shown in Fig. 3, demonstrates the origin of the circumflex coronary artery from the dilated left main coronary artery. Several obtuse marginal branches also are appreciated. The left anterior descending coronary artery is not seen arising from either the left main coronary artery or the fistula. This observation was subsequently confirmed at catheterization, in which atresia of the proximal left anterior descending coronary artery was demonstrated. The distal left anterior descending coronary artery was supplied via collateral vessels from the right coronary artery. The volume-rendered image shown in Fig. 4 demonstrates the fistula arising from behind the main pulmonary artery and entering the anterior aspect of the left ventricle. The hollow view shown in Fig. 5 (also seen in the accompanying video) provides the most dramatic view of the three-dimensional anatomy. Using this advanced software algorithm, unique en face views of the fistula’s origin and drainage site are displayed. The internal mural surface of the coronary fistula is also demonstrated. With the aid of information provided by the CT angiogram, the child underwent successful transcatheter closure of the large coronary artery fistula. The use of advanced imaging CT software not only depicted the course of the fistula and its surrounding anatomy from a three-dimensional perspective but also provided precise anatomic measurements for optimal sizing and positioning of the occluder device. Electronic supplementary material The online version of this article (doi:10.1007/s00246-012-0552-9) contains supplementary material, which is available to authorized users.

Malignant transformation: The role of MRS

Magnetic Resonance Spectroscopy (MRS) has been instrumental in elucidating the metabolic changes that arise as a consequence of malignant transformation. The metabolic phenotype of cancers is characterized by high glucose uptake, high glycolytic activity and lactate production, low mitochondrial activity, low bioenergetic status, and abnormal phospholipid metabolism [1,2]. As shown in Fig. 1, noninvasive H MRS is able to detect elevated lactate and increased total choline (tCho) levels in breast (and other) cancers, which primarily arise from oncogenic signaling pathways in cancer cells as indicated by green boxes in Fig. 1A. The increased levels of lactate and tCho provide noninvasive MR biomarkers of transformation, staging, and response to therapy. Enzymes in glucose and choline metabolism present targets for image-guided cancer therapy.

English

Summary of roof characteristics. Plant list is a partial list of the species in sampling plots on each roof. See Table A1 for more complete lists of plants, including those outside of plot areas. Area range for CH indicates sizes of three green roof boxes. Location Coordinates Soil characteristics Soil Depth (cm) Area (m 2 ) Height above ground (m) Distance to nearest green area (m) % Vegetation Cover (Mean ± standard deviation) Plant richness (Mean ± standard deviation) Plant Species Identified Water treatment vault, Wellesley College Botanic Gardens (WT) 42.2942°N-71.3039°W Low organic content, expanded shale and compost 15 55 0-2 0 41±11 4.9±1.2 Carex pensylvanica , Solidago sciaphila , Fragaria virginiana , Dennstaedtia punctilobula , Geum triflorum , Aster spp. , Campanula rotundifolia , Allium sp. Massachusetts College of Art and Design (MA) 42.3369°N-71.0985°W High organic content 30 149 60 16 49±14 7.4±0.9 Arctostaphylos uva-ursi , Prunus maritima Linaria vulgaris

Evaluation of bike boxes at signalized intersections

This paper presents a before–after study of bike boxes at 10 signalized intersections in Portland, Oregon. The bike boxes, also known as advanced stop lines or advanced stop boxes, were installed to increase visibility of cyclists and reduce conflicts between motor vehicles and cyclists, particularly in potential “right-hook” situations. Before and after video were analyzed for seven intersections with green bike boxes, three intersections with uncolored bike boxes, and two control intersections. User perceptions were measured through surveys of cyclists passing through five of the bike box intersections and of motorists working downtown, where the boxes were concentrated. Both the observations and survey of motorists found a high rate of compliance and understanding of the markings. Overall, 73% of the stopping motor vehicles did not encroach at all into the bike box. Both motor vehicle and bicycle encroachment in the pedestrian crosswalk fell significantly at the bike box locations compared to the control intersections. The bike boxes had mixed effects on the motorists’ encroachment in the bicycle lane. The number of observed conflicts at the bike box locations decreased, while the total number of cyclists and motor vehicles turning right increased. Negative-binomial models based upon the data predict fewer conflicts with the boxes, particularly as right-turning motor vehicle volumes increase. Observations of yielding behavior at two bike box and one control intersection found an improvement in motorists yielding to cyclists at the bike box locations. Differences in the traffic volumes and location contexts make firm conclusions about the effects of green coloring of the boxes difficult. Higher shares of surveyed motorists felt that the bike boxes made driving safer rather than more dangerous, even when the sample was narrowed to respondents who were not also cyclists. Over three-quarters of the surveyed cyclists thought that the boxes made the intersection safer.

Dystrophin Delivery to Muscles of mdx Mice Using Lentiviral Vectors Leads to Myogenic Progenitor Targeting and Stable Gene Expression

To explore whether stable transduction of myogenic stem cells using lentiviral vectors could be of benefit for treating dystrophic muscles, we generated vectors expressing a functional microdystrophin/enhanced green fluorescence protein fusion (microDys/eGFP) gene. Lentiviral vector injection into neonatal mdx(4cv) muscles resulted in widespread and stable expression of dystrophin for at least 2 years. This expression resulted in a significant amelioration of muscle pathophysiology as assessed by a variety of histological and functional assays. To assess whether this long-term expression was accompanied by stable transduction of satellite cells, we harvested muscle mononuclear cells 1 year after vector injection. Up to 20% of the cultured myoblast colonies expressed the microDys/eGFP transgene following myotube formation. Furthermore, transplantation of the muscle mononuclear cells into secondary mdx(4cv) recipients showed their ability to regenerate dystrophin-expressing myofibers in vivo. The ability to isolate myogenic cells able to form dystrophin-positive myotubes or myofibers in vitro and in vivo >1 year postinjection indicates that the vectors stably transduced muscle satellite cells, or a progenitor of such cells, in neonatal mdx(4cv) muscles. These studies suggest that integrating lentiviral vectors have potential utility for gene therapy of muscular dystrophy.

The long arm of BRAFV600E gets to mTORC1

The long arm of BRAF^V600E gets to mTORC1

RNA-dependent recruitment of the origin recognition complex

The origin recognition complex (ORC) has an important function in determining the initiation sites of DNA replication. In higher eukaryotes, ORC lacks sequence-specific DNA binding, and the mechanisms of ORC recruitment and origin determination are poorly understood. ORC is recruited with high efficiency to the Epstein-Barr virus origin of plasmid replication (OriP) through a complex mechanism involving interactions with the virus-encoded EBNA1 protein. We present evidence that ORC recruitment to OriP and DNA replication function depends on RGG-like motifs, referred to as LR1 and LR2, in the EBNA1 amino-terminal domain. Moreover, we show that LR1 and LR2 recruitment of ORC is RNA dependent. HMGA1a, which can functionally substitute for LR1 and LR2 domain, can also recruit ORC in an RNA-dependent manner. EBNA1 and HMGA1a RGG motifs bound to structured G-rich RNA, as did ORC1 peptides, which interact with EBNA1. RNase A treatment of cellular chromatin released a fraction of the total ORC, suggesting that ORC association with chromatin, and possibly cellular origins, is stabilized by RNA. We propose that structural RNA molecules mediate ORC recruitment at some cellular and viral origins, similar to OriP.