16,631 research outputs found

    Epilepsy Mortality: Leading Causes of Death, Co-morbidities, Cardiovascular Risk and Prevention

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    a reuptake inhibitor selectively prevents seizure-induced sudden death in the DBA/1 mouse model of sudden unexpected ... Bilateral lesions of the fastigial nucleus prevent the recovery of blood pressure following hypotension induced by ..

    The Role of English and Welsh INGOs: A Field Theory-Based Exploration of the Sector

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    This thesis takes a field theory-based approach to exploring the role of English and Welsh international non-governmental organisations (INGOs), using the lens of income source form. First, the thesis presents new income source data drawn from 933 Annual Accounts published by 316 INGOs over three years (2015-2018). The research then draws on qualitative data from 90 Leaders' letters include within the Annual Reports published by 39 INGOS, as well as supplementary quantitative and qualitative data, to explore the ways in which INGOs represent their role. Analysis of this income source data demonstrates that government funding is less important to most INGOs than has previously been assumed, while income from individuals is more important than has been recognised in the extant development studies literature. Funding from other organisations within the voluntary sector is the third most important source of income for these INGOs, while income from fees and trading is substantially less important than the other income source forms. Using this income source data in concert with other quantitative data on INGO characteristics as well as qualitative data drawn from the Leaders' letters, I then show that the English and Welsh INGO sector is a heterogenous space, divided into multiple fields. The set of fields identified by this thesis is arranged primarily around income source form, which is also associated with size, religious affiliation, and activities of focus and ways of working. As Bourdieusian field theory suggests, within these fields individual INGOs are engaged in an ongoing struggle for position: competing to demonstrate their maximal possession of the symbolic capitals they perceive to be valued by (potential) donors to that field. Further analysis of these Leaders' letters, alongside additional Annual Reports and Accounts data, also reveals a dissonance in the way in which INGOs describe their relationship with local partners in these different communication types. While these Leaders' letters and narrative reports tell stories of collaborative associations with locally-based partners, this obscures the nature of these relationships as competitive and hierarchical. The thesis draws on the above findings to reflect on the role of INGOs as suggested in the extant literature. This discussion highlights how the various potential INGO fields identified are associated with differing theoretical roles for INGOs. Finally, the thesis considers how INGO role representations continue to contribute to unequal power relations between INGOs and their partners

    The mechanisms of antibody generation in the llama

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    The llama is able to generate a unique class of antibody. The heavy chain immunoglobulins consist only of two heavy chain polypeptides and bind antigen specifically through single protein domains. Although the mechanisms by which such an antibody interacts with antigen has been studied at some length the manner in which the heavy chain antibody is generated within the llama is unknown. In this study a number of components of the llama immune system have been characterised. The isolation of genes encoding the variable domain of the heavy chain antibody indicates that specific genetic elements within the llama genome are responsible for the generation of the heavy chain antibody. The discovery of constant region genes that encode the heavy chain antibody provides an explanation for the absence of a major immunoglobulin domain from the final, secreted gene product. The lack of this domain within the expressed antibody is believed to be the result of a single nucleotide splice site mutation. In order to investigate the process of llama antibody generation further additional components of the llama immune system, the recombination activating genes (rag) were isolated. One such llama rag gene (rag-i) was cloned, expressed and utilised in an in vitro assay system to investigate recombination events taking place during antibody generation. This assay involved the use of specific signal sequences derived from variable domain gene sequence data and represents, to our knowledge, the first examination of non-murine RAG activity. Through the use of this system distinct differences between llama and mouse recombination signal sequences (RSSs) were uncovered. These differences, located within a specific region of the RSS known as the coding flank, may play an important role in llama antibody generation. These results have led to the proposal of a number of models for the mechanisms involved in llama antibody generation

    Machine learning and large scale cancer omic data: decoding the biological mechanisms underpinning cancer

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    Many of the mechanisms underpinning cancer risk and tumorigenesis are still not fully understood. However, the next-generation sequencing revolution and the rapid advances in big data analytics allow us to study cells and complex phenotypes at unprecedented depth and breadth. While experimental and clinical data are still fundamental to validate findings and confirm hypotheses, computational biology is key for the analysis of system- and population-level data for detection of hidden patterns and the generation of testable hypotheses. In this work, I tackle two main questions regarding cancer risk and tumorigenesis that require novel computational methods for the analysis of system-level omic data. First, I focused on how frequent, low-penetrance inherited variants modulate cancer risk in the broader population. Genome-Wide Association Studies (GWAS) have shown that Single Nucleotide Polymorphisms (SNP) contribute to cancer risk with multiple subtle effects, but they are still failing to give further insight into their synergistic effects. I developed a novel hierarchical Bayesian regression model, BAGHERA, to estimate heritability at the gene-level from GWAS summary statistics. I then used BAGHERA to analyse data from 38 malignancies in the UK Biobank. I showed that genes with high heritable risk are involved in key processes associated with cancer and are often localised in genes that are somatically mutated drivers. Heritability, like many other omics analysis methods, study the effects of DNA variants on single genes in isolation. However, we know that most biological processes require the interplay of multiple genes and we often lack a broad perspective on them. For the second part of this thesis, I then worked on the integration of Protein-Protein Interaction (PPI) graphs and omics data, which bridges this gap and recapitulates these interactions at a system level. First, I developed a modular and scalable Python package, PyGNA, that enables robust statistical testing of genesets' topological properties. PyGNA complements the literature with a tool that can be routinely introduced in bioinformatics automated pipelines. With PyGNA I processed multiple genesets obtained from genomics and transcriptomics data. However, topological properties alone have proven to be insufficient to fully characterise complex phenotypes. Therefore, I focused on a model that allows to combine topological and functional data to detect multiple communities associated with a phenotype. Detecting cancer-specific submodules is still an open problem, but it has the potential to elucidate mechanisms detectable only by integrating multi-omics data. Building on the recent advances in Graph Neural Networks (GNN), I present a supervised geometric deep learning model that combines GNNs and Stochastic Block Models (SBM). The model is able to learn multiple graph-aware representations, as multiple joint SBMs, of the attributed network, accounting for nodes participating in multiple processes. The simultaneous estimation of structure and function provides an interpretable picture of how genes interact in specific conditions and it allows to detect novel putative pathways associated with cancer

    Examining the Potential for Isotope Analyses of Carbon, Nitrogen, and Sulphur in Burned Bone from Experimental and Archaeological Contexts.

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    The aim of this project was to determine whether isotope analyses of carbon, nitrogen and sulphur can be conducted on collagen extracted from burned bone. This project was conducted in two phases: a controlled heating experiment and an archaeological application. The controlled heating experiment used cow (Bos taurus) bone to test the temperature thresholds for the conservation of δ13C, δ15N, and δ34S values. These samples were also used to test the efficacy of Fourier Transform Infrared spectroscopy (FTIR) and colour analysis, for determining the burning intensities experienced by bone burned in unknown conditions. The experiment showed that δ13C values were relatively unchanged up to 400°C (<2‰ variation), while δ15N values were relatively stable up to 200°C (0.5‰ variation). Values of δ34S were also relatively stable up to 200°C (1.4‰ variation). Colour change and FTIR data were well correlated with the change in isotope ratios. Models estimating burning intensities were created from the FTIR data. For the archaeological application, samples were selected from two early Anglo-Saxon cemetery sites: Elsham and Cleatham. Samples were selected from both inhumed and cremated individuals. Among the inhumed individuals δ13C values suggested a C3 terrestrial diet and δ15N values suggested protein derived largely from terrestrial herbivores, as expected for the early Anglo-Saxon period. However, δ34S values suggested the consumption of freshwater resources and that this consumption was related to both the age and sex of the individual. The experimental data shows that there is potential for isotope analyses of cremated remains, as during the cremation process heat exposures are not uniform across the body. The samples selected for the archaeological application, however, were not successful. Bone samples heated in controlled conditions produced viable collagen for isotope analysis; however, there are several differences between experiments conducted in a muffle furnace and open-air pyre cremation that need to be investigated further. Additionally, the influence of taphonomy on collagen survival in burned bone needs to be quantified. Finally, methods of sample selection need to be improved to find bone samples from archaeologically cremated remains that are most likely to retain viable collagen. While there is significant research that must be conducted before this research can be widely applied there are a multitude of cultures that practised cremation throughout history and around the world that could be investigated through the analyses proposed in this project

    Breaking Ub with Leishmania mexicana: a ubiquitin activating enzyme as a novel therapeutic target for leishmaniasis

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    Leishmaniasis is a neglected tropical disease, which inflicts a variety of gruesome pathologies on humans. The number of individuals afflicted with leishmaniasis is thought to vary between 0.7 and 1.2 million annually, of whom it is estimated that 20 to 40 thousand die. This problem is exemplary of inequality in healthcare – current leishmaniasis treatments are inadequate due to toxicity, cost, and ineffectiveness, so there is an urgent need for improved chemotherapies. Ubiquitination is a biochemical pathway that has received attention in cancer research. It is the process of adding the ubiquitin protein as a post-translational modification to substrate proteins, using an enzymatic cascade comprised of enzymes termed E1s, E2s, and E3s. Ubiquitination can lead to degradation of substrate proteins, or otherwise modulate their function. As the name suggests, this modification can be found across eukaryotic cell biology. As such, interfering with ubiquitination may interfere with essential biological processes, which means ubiquitination may present a new therapeutic target for leishmaniasis. Before ubiquitination inhibitors can be designed, components of the ubiquitination system must be identified. To this end, a bioinformatic screening campaign employed BLASTs and hidden Markov models, using characterised orthologs from model organisms as bait, to screen publicly-available Leishmania mexicana genome sequence databases, searching for genes encoding putative E1s, E2s, and E3s. To confirm some of these identifications on a protein level, activity-based probes, protein pulldowns, and mass spectrometry were used. Using an activity-based probe that emulates the structure of adenylated ubiquitin, E1s were identified, and their relative abundance quantified. A chemical crosslinker extended the reach of this probe, allowing the identification of an E2 (LmxM.33.0900). It is noted that L. mexicana has two E1s – unusual for a single celled organism. Of these E1s, LmxM.34.3060 was considerably more abundant than LmxM.23.0550 in both major life cycle stages of the in vitro Leishmania cultures. It is important to describe the wider context of these enzymes – what is their interactome, what are their substrates? To study this, CRISPR was used to fuse a proximity-based labelling system, BioID, on genes of interest – LmxM.34.3060 and LmxM.33.0900. The E2 (LmxM.33.0900) was shown to interact with the E1 (LmxM.34.3060), validating the results from the activity-based probe and crosslinker experiments. Due to sequence homology with characterised orthologs, the E2 was hypothesised to function in the endoplasmic reticulum degradation pathway. Immunoprecipitations of a ubiquitin motif, diglycine, were conducted with a view to gathering information on the substrates of ubiquitin. Anti-diglycine peptides included some of those identified by BioID. Experiments examining ubiquitin’s role in the DNA damage response were also initiated, as were improvements to the proximity-based labelling system, however these were not followed to completion due to a lack of time and resources. To examine the possibility of finding novel drug targets in the ubiquitination cascade, recombinant proteins were expressed. LmxM.34.3060 was expressed in a functional form, while a putative SUMO E2 (LmxM.02.0390) was functional after refolding. Expressed LmxM.33.0900 was not functional and could not be refolded into a functional form. Drug assays were conducted on LmxM.34.3060, which found an inhibitor of the human ortholog, TAK-243, to be 20-fold less effective against the Leishmania enzyme. Additional assays found an inhibitor that was 50-fold more effective at inhibiting the Leishmania enzyme as opposed to its human equivalent - 5'O-sulfamoyl adenosine. Furthermore, a new mechanism of action, inhibiting the E1, for was identified for drugs previously characterised to inhibit protein synthesis. LmxM.34.3060 underwent biophysical characterisation, with structural information obtained using SAXS and protein crystallography. A crystal structure was solved to 3.1 Å, with the in-solution SAXS structure complementary to this. TAK-243 was modelled into the LmxM.34.3060 structure and clashes were predicted, concurring with TAK-243’s reduced efficacy against the Leishmania enzyme in the drug assays. This project aimed to characterise the potential of an understudied biochemical system to provide novel therapeutic targets for a neglected tropical pathogen. To achieve this aim it presents the identifications of two E1s, an interactome, a structure, and a potent, selective inhibitor of a Leishmania ubiquitin activating enzyme

    Can transposon directed insertion-site sequencing be used to predict possible outcomes of evolution?

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    Laboratory-based evolution has become a tool that is widely used to understand an organism’s response to stressful environments through linking the genotype to the phenotype. Within laboratory evolution, the role that loss of function mutations play in adaptation is a topic of debate, with recent observations suggesting that adaptive loss of function mutations are a common adaptive strategy. One limiting factor of this technique is that the time taken to conduct a single experiment can be extensive. With these points in mind, we proposed to see if a short term selection experiment on a high density transposon library, using Transposon Directed Insertion-site Sequencing (TraDIS) to analyse the data, would produce results which correlate with those from long-term evolution experiments. Since TraDIS provides a measure of relative contributions to fitness of each gene, in principle it should be possible to use TraDIS to identify genes whose loss of function provides a fitness benefit on a significantly shorter timescale. Previously in our laboratory, five populations of E. coli K-12 MG1655 were evolved in a dynamic pH environment by daily passaging over five months in unbuffered LB, starting at pH 4.5. Whole genome resequencing of the final populations and clones revealed many striking similarities in the evolutionary trajectories of these populations. Therefore, to explore the hypothesis that short term selection of a high density transposon library could identify genes that were also found in the five month evolution experiment, an E. coli K-12 MG1655 transposon library was constructed and passaged for 10 days under similar conditions as the evolution experiment at both pH 4.5 and pH 7. TraDIS analysis showed that, within these populations, insertions in a few genes had accumulated, suggesting there was a fitness advantage for a strain carrying these insertions. These genes showed a significant overlap with the ones identified in the evolution experiment. These results highlight a possible alternative approach to laboratory evolution when attempting to understand an organism’s response to stress, providing a foundation for future work to explore different conditions. Research data supporting this thesis can be found on the University of Birmingham eData repository at: https://doi.org/10.25500/edata.bham.0000075

    Probing the charge generation and recombination in thin-film, optoelectronic devices

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    Sustainably and environment-friendly manufactured semiconductors are at-tractive candidates for next generation electronic and optoelectronic appli-cations ranging from memory storage and computation, to power manage-ment and energy generation. In this regard, organic semiconductors, i.e., semiconductors based on conjugated carbon-based molecules and polymers derived from earth abundant elements, are the subject of intense basic re-search and technological development efforts. Understanding the funda-mental processes governing these low-mobility and disordered semiconduct-ing materials is therefore key to establish next generation applications based upon flexible and solution-processible organic semiconductors as global com-mercial technologies.The work presented in this thesis focuses on the investigation of charge generation and recombination processes on thin film optoelectronic devices based upon organic semiconductors. A suite of experimental techniques, im-proved measurement setups, and expanded approaches are presented, and form the basis of comprehensive studies on state-of-the-art, high-efficiency organic photovoltaic systems. Specifically, an external quantum efficiency measurement technique with unprecedented dynamic range will be detailed. Using this enhanced apparatus, an approach allowing one to accurately de-termine charge generation quantum yields is introduced. After this, an extended technique to probe photogenerated charge carrier densities is out-lined and applied to thin-film solar cells. Having emphasized the importance of studying charge generation, a combined theoretical and experimental ex-ploration of the light intensity dependence of photocurrent and charge col-lection efficiency under the influence of various loss mechanisms is described. These insights provide the basis of a comprehensive study on organic so-lar cells, where recombination caused by localized trap states is found to be universally present under operational conditions limiting photocurrent and power-conversion efficiency. Overall, the work presented in this thesis expands on existing techniques and approaches, and yields important new understanding as to the device physics of thin-film, optoelectronic applica-tions
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