Uncovering key transcriptional and epigenetic changes underpinning the emergence of the glioma stem cell state

Glioblastoma Multiforme (GBM) is the most aggressive and common adult primary brain tumour, with an awful prognosis of 14 months median survival. GBMs can also occur in children, as paediatric high-grade glioma (pHGG), or diffuse intrinsic pontine glioma (DIPG), and like adult GBMs are incurable. GBM tumour growth and relapse after therapy is thought to be driven by the glioma stem cell (GSC). GSCs display similarities to neural stem and progenitor cells and likely arise from these cells. However, we currently have a poor understanding of the transcriptional and epigenetic reconfiguration which follows neural stem cell (NSC) to GSC transformation. In particular, further work is required to understand how common driver mutations contribute to immune evasion in adult GBM, and how the common paediatric mutation H3.3-G34R drives pHGG oncogenesis in a forebrain specific context. In the first instance using a novel set of engineered mouse NSC lines harbouring common adult glioma-associated driver mutations (RTK/PI3K/MAPK/P53 pathways) and transcriptional and epigenetic profiling, we explored mechanisms of adult GBM immune evasion as tumour-initiating GSCs were serially transplanted into C57BL/6J (BL6) immunocompetent mice. Second, to model the paediatric disease, an additional set of cortex and hindbrain human foetal NSC lines were engineered with paediatric glioma-associated mutations (H3.3-G34R mutations and RTK/P53 pathways). We used multi-omics data and bioinformatics approaches to characterise these adult and paediatric models of GBM, alongside adult and paediatric GSCs, to understand the transcriptional and epigenetic alterations imposed by these driver mutations. We found that triple mutants (Nf1 and Pten knock-out (KO); EGFRvIII overexpression), herein termed NPE cells, demonstrated unconstrained proliferation and a differentiation block through transcriptional downregulation and epigenetic silencing of Bmpr1b. Furthermore, we found an acquired capacity for immune evasion in NPE derivatives (in the absence of any genetic selection), suggesting a form of epigenetic immunoediting. In particular we found, upregulation of myeloid associated transcription factors (e.g. Irf8), interferon (IFN) response genes, and myeloid suppressor cell chemo-attractants such as Ccl9 which drove a myeloid-derived suppressive cell (MDSC) weighted tumour micro-environment. In human we demonstrated that GSCs could be defined by a single axis of IFN-gamma, immune response gene activation, associated with the ‘mesenchymal’ (MES) transcriptional subtype. We have therefore uncovered a key IFN driven immune evasion signal present within MES like GBM, imposed by the tumour microenvironment, which likely supports immune evasion. In a parallel analysis of foetal engineered NSC transcriptomes (H3.3-G34R, PDGFRA overexpression, and TP53 KO) obtained from cortex and hindbrain regions, we demonstrated that in a forebrain context the H3.3-G34R mutation reinforces pre-existing forebrain progenitor transcriptional circuits, whilst in the hindbrain it drives a cytostatic response. The G34R mutation does not cause widespread epigenetic changes, but instead maintains highly expressed forebrain genes through reduced binding of transcriptional regulator ZMYND11. A key mechanistic role for the oncohistone mutation H3.3-G34R in pHGG is therefore to reinforce forebrain NSC identity, explaining the mutations incompatibility within the hindbrain

Mining and analysis of audiology data to find significant factors associated with tinnitus masker

Objectives: The objective of this research is to find the factors associated with tinnitus masker from the literature, and by using the large amount of audiology data available from a large NHS (National Health Services, UK) hearing aid clinic. The factors evaluated were hearing impairment, age, gender, hearing aid type, mould and clinical comments. Design: The research includes literature survey for factors associated with tinnitus masker, and performs the analysis of audiology data using statistical and data mining techniques. Setting: This research uses a large audiology data but it also faced the problem of limited data for tinnitus. Participants: It uses 1,316 records for tinnitus and other diagnoses, and 10,437 records of clinical comments from a hearing aid clinic. Primary and secondary outcome measures: The research is looking for variables associated with tinnitus masker, and in future, these variables can be combined into a single model to develop a decision support system to predict about tinnitus masker for a patient. Results: The results demonstrated that tinnitus maskers are more likely to be fit to individuals with milder forms of hearing loss, and the factors age, gender, type of hearing aid and mould were all found significantly associated with tinnitus masker. In particular, those patients having Age<=55 years were more likely to wear a tinnitus masker, as well as those with milder forms of hearing loss. ITE (in the ear) hearing aids were also found associated with tinnitus masker. A feedback on the results of association of mould with tinnitus masker from a professional audiologist of a large NHS (National Health Services, UK) was also taken to better understand them. The results were obtained with different accuracy for different techniques. For example, the chi-squared test results were obtained with 95% accuracy, for Support and Confidence only those results were retained which had more than 1% Support and 80% Confidence. Conclusions: The variables audiograms, age, gender, hearing aid type and mould were found associated with the choice of tinnitus masker in the literature and by using statistical and data mining techniques. The further work in this research would lead to the development of a decision support system for tinnitus masker with an explanation that how that decision was obtained

Longest terrestrial migrations and movements around the world

Long-distance terrestrial migrations are imperiled globally. We determined both round-trip migration distances (straight-line measurements between migratory end points) and total annual movement (sum of the distances between successive relocations over a year) for a suite of large mammals that had potential for long-distance movements to test which species displayed the longest of both. We found that caribou likely do exhibit the longest terrestrial migrations on the planet, but, over the course of a year, gray wolves move the most. Our results were consistent with the trophic-level based hypothesis that predators would move more than their prey. Herbivores in low productivity environments moved more than herbivores in more productive habitats. We also found that larger members of the same guild moved less than smaller members, supporting the 'gastro-centric' hypothesis. A better understanding of migration and movements of large mammals should aid in their conservation by helping delineate conservation area boundaries and determine priority corridors for protection to preserve connectivity. The magnitude of the migrations and movements we documented should also provide guidance on the scale of conservation efforts required and assist conservation planning across agency and even national boundaries

Trapping a translocating protein within the anthrax toxin channel: implications for the secondary structure of permeating proteins

Anthrax toxin consists of three proteins: lethal factor (LF), edema factor (EF), and protective antigen (PA). This last forms a heptameric channel, (PA63)7, in the host cell’s endosomal membrane, allowing the former two (which are enzymes) to be translocated into the cytosol. (PA63)7 incorporated into planar bilayer membranes forms a channel that translocates LF and EF, with the N terminus leading the way. The channel is mushroom-shaped with a cap containing the binding sites for EF and LF, and an ∼100 Å–long, 15 Å–wide stem. For proteins to pass through the stem they clearly must unfold, but is secondary structure preserved? To answer this question, we developed a method of trapping the polypeptide chain of a translocating protein within the channel and determined the minimum number of residues that could traverse it. We attached a biotin to the N terminus of LFN (the 263-residue N-terminal portion of LF) and a molecular stopper elsewhere. If the distance from the N terminus to the stopper was long enough to traverse the channel, streptavidin added to the trans side bound the N-terminal biotin, trapping the protein within the channel; if this distance was not long enough, streptavidin did not bind the N-terminal biotin and the protein was not trapped. The trapping rate was dependent on the driving force (voltage), the length of time it was applied, and the number of residues between the N terminus and the stopper. By varying the position of the stopper, we determined the minimum number of residues required to span the channel. We conclude that LFN adopts an extended-chain configuration as it translocates; i.e., the channel unfolds the secondary structure of the protein. We also show that the channel not only can translocate LFN in the normal direction but also can, at least partially, translocate LFN in the opposite direction

Prospective cohort study of procalcitonin levels in children with cancer presenting with febrile neutropenia

BACKGROUND: Febrile neutropenia (FNP) causes significant morbidity and mortality in children undergoing treatment for cancer. The development of clinical decision rules to help stratify risks in paediatric FNP patients and the use of inflammatory biomarkers to identify high risk patients is an area of recent research. This study aimed to assess if procalcitonin (PCT) levels could be used to help diagnose or exclude severe infection in children with cancer who present with febrile neutropenia, both as a single measurement and in addition to previously developed clinical decision rules. METHODS: This prospective cohort study of a diagnostic test included patients between birth and 18 years old admitted with febrile neutropenia to the Paediatric Oncology and Haematology Ward in Leeds between 1(st) October 2012 and 30(th) September 2013. Each admission with FNP was treated as a separate episode. Blood was taken for a procalcitonin level at admission with routine investigations. 'R' was used for statistical analysis. Likelihood ratios were calculated and multivariable logistic regression. RESULTS: Forty-eight episodes from 27 patients were included. PCT >2 ng/dL was strongly associated with increased risk of severe infection (likelihood ratio of 26 [95% CI 3.5, 190]). The data suggests that the clinical decision rules are largely ineffective at risk stratification, frequently over-stating the risk of individual episodes. High procalcitonin levels on admission are correlated with a greatly increased risk of severe infection. CONCLUSIONS: This study does not show a definitive benefit in using PCT in FNP though it supports further research on its use. The benefit of novel biomarkers has not been proven and before introducing new tests for patients it is important their benefit above existing features is proven, particularly due to the increasing importance of health economics

Novel Colicin F-Y of Yersinia frederiksenii Inhibits Pathogenic Yersinia Strains via YiuR-Mediated Reception, TonB Import, and Cell Membrane Pore Formation

A novel colicin type, designated colicin F-Y, was found to be encoded and produced by the strain Yersinia frederiksenii Y27601. Colicin F-Y was active against both pathogenic and nonpathogenic strains of the genus Yersinia. Plasmid YF27601 (5,574 bp) of Y. frederiksenii Y27601 was completely sequenced. The colicin F-Y activity gene (cfyA) and the colicin F-Y immunity gene (cfyI) were identified. The deduced amino acid sequence of colicin F-Y was very similar in its C-terminal pore-forming domain to colicin Ib (69% identity in the last 178 amino acid residues), indicating pore forming as its lethal mode of action. Transposon mutagenesis of the colicin F-Y-susceptible strain Yersinia kristensenii Y276 revealed the yiuR gene (ykris001_4440), which encodes the YiuR outer membrane protein with unknown function, as the colicin F-Y receptor molecule. Introduction of the yiuR gene into the colicin F-Y-resistant strain Y. kristensenii Y104 restored its susceptibility to colicin F-Y. In contrast, the colicin F-Y-resistant strain Escherichia coli TOP10F' acquired susceptibility to colicin F-Y only when both the yiuR and tonB genes from Y. kristensenii Y276 were introduced. Similarities between colicins F-Y and Ib, similarities between the Cir and YiuR receptors, and the detected partial cross-immunity of colicin F-Y and colicin Ib producers suggest a common evolutionary origin of the colicin F-Y-YiuR and colicin Ib-Cir systems

Tunneling Spectra of Individual Magnetic Endofullerene Molecules

The manipulation of single magnetic molecules may enable new strategies for high-density information storage and quantum-state control. However, progress in these areas depends on developing techniques for addressing individual molecules and controlling their spin. Here we report success in making electrical contact to individual magnetic N@C60 molecules and measuring spin excitations in their electron tunneling spectra. We verify that the molecules remain magnetic by observing a transition as a function of magnetic field which changes the spin quantum number and also the existence of nonequilibrium tunneling originating from low-energy excited states. From the tunneling spectra, we identify the charge and spin states of the molecule. The measured spectra can be reproduced theoretically by accounting for the exchange interaction between the nitrogen spin and electron(s) on the C60 cage.Comment: 7 pages, 4 figures. Typeset in LaTeX, updated text of previous versio

The effect of chair massage on muscular discomfort in cardiac sonographers: a pilot study

<p>Abstract</p> <p>Background</p> <p>Cardiac sonographers frequently have work-related muscular discomfort. We aimed to assess the feasibility of having sonographers receive massages during working hours in an area adjacent to an echocardiography laboratory and to assess relief of discomfort with use of the massages with or without stretching exercises.</p> <p>Methods</p> <p>A group of 45 full-time sonographers was randomly assigned to receive weekly 30-minute massage sessions, massages plus stretching exercises to be performed twice a day, or no intervention. Outcome measures were scores of the <it>Quick</it>DASH instrument and its associated work module at baseline and at 10 weeks of intervention. Data were analyzed with standard descriptive statistics and the separation test for early-phase comparative trials.</p> <p>Results</p> <p>Forty-four participants completed the study: 15 in the control group, 14 in the massage group, and 15 in the massage plus stretches group. Some improvement was seen in work-related discomfort by the <it>Quick</it>DASH scores and work module scores in the 2 intervention groups. The separation test showed separation in favor of the 2 interventions.</p> <p>Conclusion</p> <p>On the basis of the results of this pilot study, larger trials are warranted to evaluate the effect of massages with or without stretching on work-related discomfort in cardiac sonographers.</p> <p>Trial Registration</p> <p>NCT00975026 ClinicalTrials.gov</p

Breast cancer risk factors and a novel measure of volumetric breast density: cross-sectional study

We conducted a cross-sectional study nested within a prospective cohort of breast cancer risk factors and two novel measures of breast density volume among 590 women who had attended Glasgow University (1948–1968), replied to a postal questionnaire (2001) and attended breast screening in Scotland (1989–2002). Volumetric breast density was estimated using a fully automated computer programme applied to digitised film-screen mammograms, from medio-lateral oblique mammograms at the first-screening visit. This measured the proportion of the breast volume composed of dense (non-fatty) tissue (Standard Mammogram Form (SMF)%) and the absolute volume of this tissue (SMF volume, cm3). Median age at first screening was 54.1 years (range: 40.0–71.5), median SMF volume 70.25 cm3 (interquartile range: 51.0–103.0) and mean SMF% 26.3%, s.d.=8.0% (range: 12.7–58.8%). Age-adjusted logistic regression models showed a positive relationship between age at last menstrual period and SMF%, odds ratio (OR) per year later: 1.05 (95% confidence interval: 1.01–1.08, P=0.004). Number of pregnancies was inversely related to SMF volume, OR per extra pregnancy: 0.78 (0.70–0.86, P<0.001). There was a suggestion of a quadratic relationship between birthweight and SMF%, with lowest risks in women born under 2.5 and over 4 kg. Body mass index (BMI) at university (median age 19) and in 2001 (median age 62) were positively related to SMF volume, OR per extra kg m−2 1.21 (1.15–1.28) and 1.17 (1.09–1.26), respectively, and inversely related to SMF%, OR per extra kg m−2 0.83 (0.79–0.88) and 0.82 (0.76–0.88), respectively, P<0.001. Standard Mammogram Form% and absolute SMF volume are related to several, but not all, breast cancer risk factors. In particular, the positive relationship between BMI and SMF volume suggests that volume of dense breast tissue will be a useful marker in breast cancer studies