Scoring system to facilitate diagnosis of Gaucher disease

Background: Gaucher disease (GD) manifests heterogeneously and other conditions are often misdiagnosed in its place, leading to diagnostic delays. The Gaucher Earlier Diagnosis Consensus (GED‐C) initiative proposed a point‐scoring system (PSS) based on the signs and covariables that are most indicative of GD to help clinicians identify which individuals to test for GD. Aims: To validate the PSS retrospectively in a test population including patients with GD and other conditions with overlapping manifestations. Methods: Four cohorts of adults with GD, liver disease (LD), haematological malignancy (HM) or immune thrombocytopenia were identified from hospital records. Clinical data were audited for GED‐C factors identified as potentially indicative of GD and aggregate scores calculated (sum of scores/number of factors) based on published PSS weightings. Threshold discriminatory PSS scores, sensitivity and specificity were determined by receiver‐operating characteristic (ROC) analysis. Results: Among 100 patients (GD, n = 25; non‐GD, n = 75), analyses based on 11 possible factors estimated group mean (standard deviation) PSS scores of: GD (n = 14), 1.08 (0.25); non‐GD (n = 38), 0.58 (0.31). Mean between‐group difference (95% confidence interval (CI)) was (−0.49 (−0.68, −0.31)) and area under the ROC analysis curve (95% CI) was 0.88 (0.78, 0.97). A threshold PSS score of 0.82 identified all 14 patients with GD in the analysis set (100% sensitivity) and 27 of 38 patients in the non‐GD group (71% specificity). Patients with LD and HM were most likely to have manifestations overlapping GD. Conclusions: Preliminary validation of the GED‐C PSS discriminated effectively between patients with GD and those with overlapping signs

Multicolour Single Molecule Imaging in Cells with Near Infra-Red Dyes

Background: The autofluorescence background of biological samples impedes the detection of single molecules when imaging. The most common method of reducing the background is to use evanescent field excitation, which is incompatible with imaging beyond the surface of biological samples. An alternative would be to use probes that can be excited in the near infra-red region of the spectrum, where autofluorescence is low. Such probes could also increase the number of labels that can be imaged in multicolour single molecule microscopes. Despite being widely used in ensemble imaging, there is a currently a shortage of information available for selecting appropriate commercial near infra-red dyes for single molecule work. It is therefore important to characterise available near infra-red dyes relevant to multicolour single molecule imaging. Methodology/Principal Findings: A range of commercially available near infra-red dyes compatible with multi-colour imaging was screened to find the brightest and most photostable candidates. Image series of immobilised samples of the brightest dyes (Alexa 700, IRDye 700DX, Alexa 790 and IRDye 800CW) were analysed to obtain the mean intensity of single dye molecules, their photobleaching rates and long period blinking kinetics. Using the optimum dye pair, we have demonstrated for the first time widefield, multi-colour, near infra-red single molecule imaging using a supercontinuum light source in MCF-7 cells

A forced titration study of the antioxidant and immunomodulatory effects of Ambrotose AO supplement

Background Oxidative stress plays a role in acute and chronic inflammatory disease and antioxidant supplementation has demonstrated beneficial effects in the treatment of these conditions. This study was designed to determine the optimal dose of an antioxidant supplement in healthy volunteers to inform a Phase 3 clinical trial. Methods The study was designed as a combined Phase 1 and 2 open label, forced titration dose response study in healthy volunteers (n = 21) to determine both acute safety and efficacy. Participants received a dietary supplement in a forced titration over five weeks commencing with a no treatment baseline through 1, 2, 4 and 8 capsules. The primary outcome measurement was ex vivo changes in serum oxygen radical absorbance capacity (ORAC). The secondary outcome measures were undertaken as an exploratory investigation of immune function. Results A significant increase in antioxidant activity (serum ORAC) was observed between baseline (no capsules) and the highest dose of 8 capsules per day (p = 0.040) representing a change of 36.6%. A quadratic function for dose levels was fitted in order to estimate a dose response curve for estimating the optimal dose. The quadratic component of the curve was significant (p = 0.047), with predicted serum ORAC scores increasing from the zero dose to a maximum at a predicted dose of 4.7 capsules per day and decreasing for higher doses. Among the secondary outcome measures, a significant dose effect was observed on phagocytosis of granulocytes, and a significant increase was also observed on Cox 2 expression. Conclusion This study suggests that Ambrotose AO® capsules appear to be safe and most effective at a dosage of 4 capsules/day. It is important that this study is not over interpreted; it aimed to find an optimal dose to assess the dietary supplement using a more rigorous clinical trial design. The study achieved this aim and demonstrated that the dietary supplement has the potential to increase antioxidant activity. The most significant limitation of this study was that it was open label Phase 1/Phase 2 trial and is subject to potential bias that is reduced with the use of randomization and blinding. To confirm the benefits of this dietary supplement these effects now need to be demonstrated in a Phase 3 randomised controlled trial (RCT)

Experimental Design

AbstractThis chapter covers various issues related to the experimental design, a statistical technique at the core of a discrete choice experiment. Specifically, it focuses on the dimensionality of a choice experiment and the statistical techniques used to allocate attribute levels to choice tasks. Among others, the pros and cons of orthogonal designs, optimal orthogonal in the differences designs as well as efficient designs are addressed. The last section shows how a simulation exercise can help to test the appropriateness of the experimental design

The Obesity and Fatty Liver Are Reduced by Plant-Derived Pediococcus pentosaceus LP28 in High Fat Diet-Induced Obese Mice

We evaluated the effect of an oral administration of a plant-derived lactic acid bacterium, Pediococcus pentosaceus LP28 (LP28), on metabolic syndrome by using high fat diet-induced obese mice. The obese mice were divided into 2 groups and fed either a high fat or regular diet for 8 weeks. Each group was further divided into 3 groups, which took LP28, another plant-derived Lactobacillus plantarum SN13T (SN13T) or no lactic acid bacteria (LAB). The lean control mice were fed a regular diet without inducing obesity prior to the experiment. LP28 reduced body weight gain and liver lipid contents (triglyceride and cholesterol), in mice fed a high fat diet for 8 weeks (40%, 54%, and 70% less than those of the control group without LAB, and P = 0.018, P<0.001, and P = 0.021, respectively), whereas SN13T and the heat treated LP28 at 121°C for 15 min were ineffective. Abdominal visceral fat in the high fat diet mice fed with LP28 was also lower than that without LAB by 44%, although it was not significant but borderline (P = 0.076). The sizes of the adipocytes and the lipid droplets in the livers were obviously decreased. A real-time PCR analyses showed that lipid metabolism-related genes, such as CD36 (P = 0.013), SCD1 encoding stearoyl-CoA desaturase 1 (not significant but borderline, P = 0.066), and PPARγ encoding peroxisome proliferator-activated receptor gamma (P = 0.039), were down-regulated by taking LP28 continuously, when compared with those of the control group. In conclusion, LP28 may be a useful LAB strain for the prevention and reduction of the metabolic syndrome

Plant growth environments with programmable relative humidity and homogeneous nutrient availability

We describe the design, characterization, and use of “programmable”, sterile growth environments for individual (or small sets of) plants. The specific relative humidities and nutrient availability experienced by the plant is established (RH between 15% and 95%; nutrient concentration as desired) during the setup of the growth environment, which takes about 5 minutes and <1$ in disposable cost. These systems maintain these environmental parameters constant for at least 14 days with minimal intervention (one minute every two days). The design is composed entirely of off-the-shelf components (e.g., LEGO® bricks) and is characterized by (i) a separation of root and shoot environment (which is physiologically relevant and facilitates imposing specific conditions on the root system, e.g., darkness), (ii) the development of the root system on a flat surface, where the root enjoys constant contact with nutrient solution and air, (iii) a compatibility with root phenotyping. We demonstrate phenotyping by characterizing root systems of Brassica rapa plants growing in different relative humidities (55%, 75%, and 95%). While most phenotypes were found to be sensitive to these environmental changes, a phenotype tightly associated with root system topology – the size distribution of the areas encircled by roots – appeared to be remarkably and counterintuitively insensitive to humidity changes. These setups combine many of the advantages of hydroponics conditions (e.g., root phenotyping, complete control over nutrient composition, scalability) and soil conditions (e.g., aeration of roots, shading of roots), while being comparable in cost and setup time to Magenta® boxes

The implications of a Silurian and other thylacocephalan crustaceans for the functional morphology and systematic affinities of the group

Background: Thylacocephala is a group of enigmatic extinct arthropods. Here we provide a full description of the oldest unequivocal thylacocephalan, a new genus and species Thylacares brandonensis, which is present in the Silurian Waukesha fauna from Wisconsin, USA. We also present details of younger, Jurassic specimens, from the Solnhofen lithographic limestones, which are crucial to our interpretation of the systematic position of Thylacocephala. In the past, Thylacocephala has been interpreted as a crustacean ingroup and as closely related to various groups such as cirripeds, decapods or remipeds. Results: The Waukesha thylacocephalan, Thylacares brandonensis n. gen. n. sp., bears compound eyes and raptorial appendages that are relatively small compared to those of other representatives of the group. As in other thylacocephalans the large bivalved shield encloses much of the entire body. The shield lacks a marked optical notch. The eyes, which project just beyond the shield margin, appear to be stalked. Head appendages, which may represent antennulae, antennae and mandibles, appear to be present. The trunk is comprised of up to 22 segments. New details observed on thylacocephalans from the Jurassic Solnhofen lithographic limestones include antennulae and antennae of Mayrocaris bucculata, and endites on the raptorial appendages and an elongate last trunk appendage in Clausocaris lithographica. Preserved features of the internal morphology in C. lithographica include the muscles of the raptorial appendage and trunk. Conclusions: Our results indicate that some `typical' thylacocephalan characters are unique to the group; these autapomorphies contribute to the difficulty of determining thylacocephalan affinities. While the new features reported here are consistent with a eucrustacean affinity, most previous hypotheses for the position of Thylacocephala within Eucrustacea (as Stomatopoda, Thecostraca or Decapoda) are shown to be unlikely. A sister group relationship to Remipedia appears compatible with the observed features of Thylacocephala but more fossil evidence is required to test this assertion. The raptorial appendages of Thylacocephala most likely projected 45 degrees abaxially instead of directly forward as previously reconstructed. The overall morphology of thylacocephalans supports a predatory mode of life

Achieving Education for Sustainable Development (ESD) in Early Childhood Education Through Critical Reflection in Transformative Learning

The central role of education in creating a more sustainable future has been already recognized by educators and policy-makers alike. This chapter argues that this can only be truly achieved through the efforts of teachers in implementing an “education of a different kind,” a general educational shift that seeks to encompass a converging transformation of the priorities and mindsets of education professionals. In this regard, the professional preparation of teachers, as the leading actors in shaping children’s learning processes, and their continuous professional development are vital considerations for Education for Sustainable Development (ESD) to be successfully achieved. Linking transformative learning and ESD has emerged as a distinct and useful pedagogy because they both support the process of critically examining habits of mind, then revising these habits and acting upon the revised point of view. This study aims to describe and evaluate the potential of transformative learning in innovating mainstream education toward sustainability by focusing on the role of critical reflection in a capacity building research project realized in Turkey. The data was gathered from 24 early childhood educators using a mixed-method research design involving learning diaries, a learning activities survey, and follow-up interviews. This chapter identified content, context, and application method of the in-service training as factors that have contributed to the reflective practices of the participants. In addition, presenting the implications regarding the individual differences in how learners engage in critical reflection practices, this research offers a framework for a content- and process-based approach derived from Mezirow’s conception of critical reflection

The architecture of EGFR's basal complexes reveals autoinhibition mechanisms in dimers and oligomers

Our current understanding of epidermal growth factor receptor (EGFR) autoinhibition is based on X-ray structural data of monomer and dimer receptor fragments and does not explain how mutations achieve ligand-independent phosphorylation. Using a repertoire of imaging technologies and simulations we reveal an extracellular head-to-head interaction through which ligand-free receptor polymer chains of various lengths assemble. The architecture of the head-to-head interaction prevents kinase-mediated dimerisation. The latter, afforded by mutation or intracellular treatments, splits the autoinhibited head-to-head polymers to form stalk-to-stalk flexible non-extended dimers structurally coupled across the plasma membrane to active asymmetric tyrosine kinase dimers, and extended dimers coupled to inactive symmetric kinase dimers. Contrary to the previously proposed main autoinhibitory function of the inactive symmetric kinase dimer, our data suggest that only dysregulated species bear populations of symmetric and asymmetric kinase dimers that coexist in equilibrium at the plasma membrane under the modulation of the C-terminal domain

Metabolic Deficiences Revealed in the Biotechnologically Important Model Bacterium Escherichia coli BL21(DE3)

The Escherichia coli B strain BL21(DE3) has had a profound impact on biotechnology through its use in the production of recombinant proteins. Little is understood, however, regarding the physiology of this important E. coli strain. We show here that BL21(DE3) totally lacks activity of the four [NiFe]-hydrogenases, the three molybdenum- and selenium-containing formate dehydrogenases and molybdenum-dependent nitrate reductase. Nevertheless, all of the structural genes necessary for the synthesis of the respective anaerobic metalloenzymes are present in the genome. However, the genes encoding the high-affinity molybdate transport system and the molybdenum-responsive transcriptional regulator ModE are absent from the genome. Moreover, BL21(DE3) has a nonsense mutation in the gene encoding the global oxygen-responsive transcriptional regulator FNR. The activities of the two hydrogen-oxidizing hydrogenases, therefore, could be restored to BL21(DE3) by supplementing the growth medium with high concentrations of Ni2+ (Ni2+-transport is FNR-dependent) or by introducing a wild-type copy of the fnr gene. Only combined addition of plasmid-encoded fnr and high concentrations of MoO42− ions could restore hydrogen production to BL21(DE3); however, to only 25–30% of a K-12 wildtype. We could show that limited hydrogen production from the enzyme complex responsible for formate-dependent hydrogen evolution was due solely to reduced activity of the formate dehydrogenase (FDH-H), not the hydrogenase component. The activity of the FNR-dependent formate dehydrogenase, FDH-N, could not be restored, even when the fnr gene and MoO42− were supplied; however, nitrate reductase activity could be recovered by combined addition of MoO42− and the fnr gene. This suggested that a further component specific for biosynthesis or activity of formate dehydrogenases H and N was missing. Re-introduction of the gene encoding ModE could only partially restore the activities of both enzymes. Taken together these results demonstrate that BL21(DE3) has major defects in anaerobic metabolism, metal ion transport and metalloprotein biosynthesis