Extracellular Matrix Mineralization Promotes E11/gp38 Glycoprotein Expression and Drives Osteocytic Differentiation

Osteocytes are terminally differentiated osteoblasts which reside in a mineralized extracellular matrix (ECM). The factors that regulate this differentiation process are unknown. We have investigated whether ECM mineralization could promote osteocyte formation. To do this we have utilised MLO-A5 pre-osteocyte-like cells and western blotting and comparative RT-PCR to examine whether the expression of osteocyte-selective markers is elevated concurrently with the onset of ECM mineralization. Secondly, if mineralization of the ECM is indeed a driver of osteocyte formation, we reasoned that impairment of ECM mineralization would result in a reversible inhibition of osteocyte formation. Supplementation of MLO-A5 cell cultures with ascorbic acid and phosphate promoted progressive ECM mineralization as well as temporally associated increases in expression of the osteocyte-selective markers, E11/gp38 glycoprotein and sclerostin. Consistent with a primary role for ECM mineralization in osteocyte formation, we also found that inhibition of ECM mineralization, by omitting phosphate or adding sodium pyrophosphate, a recognized inhibitor of hydroxyapatite formation, resulted in a 15-fold decrease in mineral deposition that was closely accompanied by lower expression of E11 and other osteocyte markers such as Dmp1, Cd44 and Sost whilst expression of osteoblast markers Ocn and Col1a increased. To rule out the possibility that such restriction of ECM mineralization may produce an irreversible modification in osteoblast behaviour to limit E11 expression and osteocytogenesis, we also measured the capacity of MLO-A5 cells to re-enter the osteocyte differentiation programme. We found that the mineralisation process was re-initiated and closely allied to increased expression of E11 protein after re-administration of phosphate or omission of sodium pyrophosphate, indicating an ECM mineralization-induced restoration in osteocyte formation. These results emphasise the importance of cell-ECM interactions in regulating osteoblast behaviour and, more importantly, suggest that ECM mineralization exerts pivotal control during terminal osteoblast differentiation and acquisition of the osteocyte phenotype

An investigation of clinical and immunological events following repeated aerodigestive tract challenge infections with live Mycobacterium bovis Bacille Calmette Guerin

Bacille Calmette Guérin substrain Moreau Rio de Janeiro is an attenuated strain of Mycobacterium bovis that has been used extensively as an oral tuberculosis vaccine. We assessed its potential as a challenge model to study clinical and immunological events following repeated mycobacterial gut infection. Seven individuals received three oral challenges with approximately 107 viable bacilli. Clinical symptoms, T-cell responses and gene expression patterns in peripheral blood were monitored. Clinical symptoms were relatively mild and declined following each oral challenge. Delayed T-cell responses were observed, and limited differential gene expression detected by microarrays. Oral challenge with BCG Moreau Rio de Janeiro vaccine was immunogenic in healthy volunteers, limiting its potential to explore clinical innate immune responses, but with low reactogenicity

A comprehensive database of quality-rated fossil ages for Sahul’s Quaternary vertebrates

Published: 19 July 2016The study of palaeo-chronologies using fossil data provides evidence for past ecological and evolutionary processes, and is therefore useful for predicting patterns and impacts of future environmental change. However, the robustness of inferences made from fossil ages relies heavily on both the quantity and quality of available data. We compiled Quaternary non-human vertebrate fossil ages from Sahul published up to 2013. This, the FosSahul database, includes 9,302 fossil records from 363 deposits, for a total of 478 species within 215 genera, of which 27 are from extinct and extant megafaunal species (2,559 records). We also provide a rating of reliability of individual absolute age based on the dating protocols and association between the dated materials and the fossil remains. Our proposed rating system identified 2,422 records with high-quality ages (i.e., a reduction of 74%). There are many applications of the database, including disentangling the confounding influences of hypothetical extinction drivers, better spatial distribution estimates of species relative to palaeo-climates, and potentially identifying new areas for fossil discovery.Marta Rodríguez-Rey, y, Salvador Herrando-Pérez, Barry W. Brook, Frédérik Saltré, John Alroy, Nicholas Beeton, Michael I. Bird, Alan Cooper, Richard Gillespie, Zenobia Jacobs, Christopher N. Johnson, Gifford H. Miller, Gavin J. Prideaux, Richard G. Roberts, Chris S.M. Turney and Corey J.A. Bradsha

Evaluation of a learner-designed course for teaching health research skills in Ghana

<p>Abstract</p> <p>Background</p> <p>In developing countries the ability to conduct locally-relevant health research and high quality education are key tools in the fight against poverty. The objective of our study was to evaluate the effectiveness of a novel UK accredited, learner-designed research skills course delivered in a teaching hospital in Ghana.</p> <p>Methods</p> <p>Study participants were 15 mixed speciality health professionals from Komfo Anokye Teaching Hospital, Kumasi, Ghana. Effectiveness measures included process, content and outcome indicators to evaluate changes in learners' confidence and competence in research, and assessment of the impact of the course on changing research-related thinking and behaviour. Results were verified using two independent methods.</p> <p>Results</p> <p>14/15 learners gained research competence assessed against UK Quality Assurance Agency criteria. After the course there was a 36% increase in the groups' positive responses to statements concerning confidence in research-related attitudes, intentions and actions. The greatest improvement (45% increase) was in learners' actions, which focused on strengthening institutional research capacity. 79% of paired before/after responses indicated positive changes in individual learners' research-related attitudes (n = 53), 81% in intention (n = 52) and 85% in action (n = 52). The course had increased learners' confidence to start and manage research, and enhanced life-long skills such as reflective practice and self-confidence. Doing their own research within the work environment, reflecting on personal research experiences and utilising peer support and pooled knowledge were critical elements that promoted learning.</p> <p>Conclusion</p> <p>Learners in Ghana were able to design and undertake a novel course that developed individual and institutional research capacity and met international standards. Learning by doing and a supportive peer community at work were critical elements in promoting learning in this environment where tutors were scarce. Our study provides a model for delivering and evaluating innovative educational interventions in developing countries to assess whether they meet external quality criteria and achieve their objectives.</p

A comprehensive database of quality-rated fossil ages for Sahul's Quaternary vertebrates.

The study of palaeo-chronologies using fossil data provides evidence for past ecological and evolutionary processes, and is therefore useful for predicting patterns and impacts of future environmental change. However, the robustness of inferences made from fossil ages relies heavily on both the quantity and quality of available data. We compiled Quaternary non-human vertebrate fossil ages from Sahul published up to 2013. This, the FosSahul database, includes 9,302 fossil records from 363 deposits, for a total of 478 species within 215 genera, of which 27 are from extinct and extant megafaunal species (2,559 records). We also provide a rating of reliability of individual absolute age based on the dating protocols and association between the dated materials and the fossil remains. Our proposed rating system identified 2,422 records with high-quality ages (i.e., a reduction of 74%). There are many applications of the database, including disentangling the confounding influences of hypothetical extinction drivers, better spatial distribution estimates of species relative to palaeo-climates, and potentially identifying new areas for fossil discovery

The UK clinical aptitude test and clinical course performance at Nottingham: a prospective cohort study

Background The UK Clinical Aptitude Test (UKCAT) was introduced in 2006 as an additional tool for the selection of medical students. It tests mental ability in four distinct domains (Verbal Reasoning, Quantitative Reasoning, Abstract Reasoning, and Decision Analysis), and the results are available to students and admission panels in advance of the selection process. Our first study showed little evidence of any predictive validity for performance in the first two years of the Nottingham undergraduate course. The study objective was to determine whether the UKCAT scores had any predictive value for the later parts of the course, largely delivered via clinical placements. Methods Students entering the course in 2007 and who had taken the UKCAT were asked for permission to use their anonymised data in research. The UKCAT scores were incorporated into a database with routine pre-admission socio-demographics and subsequent course performance data. Correlation analysis was followed by hierarchical multivariate linear regression. Results The original study group comprised 204/254 (80%) of the full entry cohort. With attrition over the five years of the course this fell to 185 (73%) by Year 5. The Verbal Reasoning score and the UKCAT Total score both demonstrated some univariate correlations with clinical knowledge marks, and slightly less with clinical skills. No parts of the UKCAT proved to be an independent predictor of clinical course marks, whereas prior attainment was a highly significant predictor (p <0.001). Conclusions This study of one cohort of Nottingham medical students showed that UKCAT scores at admission did not independently predict subsequent performance on the course. Whilst the test adds another dimension to the selection process, its fairness and validity in selecting promising students remains unproven, and requires wider investigation and debate by other schools

Scrapheap Challenge: A novel bulk-bone metabarcoding method to investigate ancient DNA in faunal assemblages

Highly fragmented and morphologically indistinct fossil bone is common in archaeological and paleontological deposits but unfortunately it is of little use in compiling faunal assemblages. The development of a cost-effective methodology to taxonomically identify bulk bone is therefore a key challenge. Here, an ancient DNA methodology using high-throughput sequencing is developed to survey and analyse thousands of archaeological bones from southwest Australia. Fossils were collectively ground together depending on which of fifteen stratigraphical layers they were excavated from. By generating fifteen synthetic blends of bulk bone powder, each corresponding to a chronologically distinct layer, samples could be collectively analysed in an efficient manner. A diverse range of taxa, including endemic, extirpated and hitherto unrecorded taxa, dating back to c.46,000 years BP was characterized. The method is a novel, cost-effective use for unidentifiable bone fragments and a powerful molecular tool for surveying fossils that otherwise end up on the taxonomic “scrapheap”

Search for Doubly-Charged Higgs Boson Production at HERA

A search for the single production of doubly-charged Higgs bosons H^{\pm \pm} in ep collisions is presented. The signal is searched for via the Higgs decays into a high mass pair of same charge leptons, one of them being an electron. The analysis uses up to 118 pb^{-1} of ep data collected by the H1 experiment at HERA. No evidence for doubly-charged Higgs production is observed and mass dependent upper limits are derived on the Yukawa couplings h_{el} of the Higgs boson to an electron-lepton pair. Assuming that the doubly-charged Higgs only decays into an electron and a muon via a coupling of electromagnetic strength h_{e \mu} = \sqrt{4 \pi \alpha_{em}} = 0.3, a lower limit of 141 GeV on the H^{\pm\pm} mass is obtained at the 95% confidence level. For a doubly-charged Higgs decaying only into an electron and a tau and a coupling h_{e\tau} = 0.3, masses below 112 GeV are ruled out.Comment: 15 pages, 3 figures, 1 tabl

Single-cell analysis: visualizing pharmaceutical and metabolite uptake in cells with label-free 3D mass spectrometry imaging

Detecting metabolites and parent compound within a cell type is now a priority for pharmaceutical development. In this context, three-dimensional secondary ion mass spectrometry (SIMS) imaging was used to investigate the cellular uptake of the antiarrhythmic agent amiodarone, a phospholipidosis-inducing pharmaceutical compound. The high lateral resolution and 3D imaging capabilities of SIMS combined with the multiplex capabilities of ToF mass spectrometric detection allows for the visualization of pharmaceutical compound and metabolites in single cells. The intact, unlabeled drug compound was successfully detected at therapeutic dosages in macrophages (cell line: NR8383). Chemical information from endogenous biomolecules was used to correlate drug distributions with morphological features. From this spatial analysis, amiodarone was detected throughout the cell with the majority of the compound found in the membrane and subsurface regions and absent in the nuclear regions. Similar results were obtained when the macrophages were doped with amiodarone metabolite, desethylamiodarone. The FWHM lateral resolution measured across an intracellular interface in a high lateral resolution ion images was approximately 550 nm. Overall, this approach provides the basis for studying cellular uptake of pharmaceutical compounds and their metabolites on the single cell level