526 research outputs found

    The development of bioinformatics workflows to explore single-cell multi-omics data from T and B lymphocytes

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    The adaptive immune response is responsible for recognising, containing and eliminating viral infection, and protecting from further reinfection. This antigen-specific response is driven by T and B cells, which recognise antigenic epitopes via highly specific heterodimeric surface receptors, termed T-cell receptors (TCRs) and B cell receptors (BCRs). The theoretical diversity of the receptor repertoire that can be generated via homologous recombination of V, D and J genes is large enough (>1015 unique sequences) that virtually any antigen can be recognised. However, only a subset of these are generated within the human body, and how they succeed in specifically recognising any pathogen(s) and distinguishing these from self-proteins remains largely unresolved. The recent advances in applying single-cell genomics technologies to simultaneously measure the clonality, surface phenotype and transcriptomic signature of pathogen- specific immune cells have significantly improved understanding of these questions. Single-cell multi-omics permits the accurate identification of clonally expanded populations, their differentiation trajectories, the level of immune receptor repertoire diversity involved in the response and the phenotypic and molecular heterogeneity. This thesis aims to develop a bioinformatic workflow utilising single-cell multi-omics data to explore, quantify and predict the clonal and transcriptomic signatures of the human T-cell response during and following viral infection. In the first aim, a web application, VDJView, was developed to facilitate the simultaneous analysis and visualisation of clonal, transcriptomic and clinical metadata of T and B cell multi-omics data. The application permits non-bioinformaticians to perform quality control and common analyses of single-cell genomics data integrated with other metadata, thus permitting the identification of biologically and clinically relevant parameters. The second aim pertains to analysing the functional, molecular and immune receptor profiles of CD8+ T cells in the acute phase of primary hepatitis C virus (HCV) infection. This analysis identified a novel population of progenitors of exhausted T cells, and lineage tracing revealed distinct trajectories with multiple fates and evolutionary plasticity. Furthermore, it was observed that high-magnitude IFN-γ CD8+ T-cell response is associated with the increased probability of viral escape and chronic infection. Finally, in the third aim, a novel analysis is presented based on the topological characteristics of a network generated on pathogen-specific, paired-chain, CD8+ TCRs. This analysis revealed how some cross-reactivity between TCRs can be explained via the sequence similarity between TCRs and that this property is not uniformly distributed across all pathogen-specific TCR repertoires. Strong correlations between the topological properties of the network and the biological properties of the TCR sequences were identified and highlighted. The suite of workflows and methods presented in this thesis are designed to be adaptable to various T and B cell multi-omic datasets. The associated analyses contribute to understanding the role of T and B cells in the adaptive immune response to viral-infection and cancer

    International consensus statement on allergy and rhinology: Allergic rhinitis – 2023

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    Background In the 5 years that have passed since the publication of the 2018 International Consensus Statement on Allergy and Rhinology: Allergic Rhinitis (ICAR-Allergic Rhinitis 2018), the literature has expanded substantially. The ICAR-Allergic Rhinitis 2023 update presents 144 individual topics on allergic rhinitis (AR), expanded by over 40 topics from the 2018 document. Originally presented topics from 2018 have also been reviewed and updated. The executive summary highlights key evidence-based findings and recommendation from the full document. Methods ICAR-Allergic Rhinitis 2023 employed established evidence-based review with recommendation (EBRR) methodology to individually evaluate each topic. Stepwise iterative peer review and consensus was performed for each topic. The final document was then collated and includes the results of this work. Results ICAR-Allergic Rhinitis 2023 includes 10 major content areas and 144 individual topics related to AR. For a substantial proportion of topics included, an aggregate grade of evidence is presented, which is determined by collating the levels of evidence for each available study identified in the literature. For topics in which a diagnostic or therapeutic intervention is considered, a recommendation summary is presented, which considers the aggregate grade of evidence, benefit, harm, and cost. Conclusion The ICAR-Allergic Rhinitis 2023 update provides a comprehensive evaluation of AR and the currently available evidence. It is this evidence that contributes to our current knowledge base and recommendations for patient evaluation and treatment

    Current Insights on Lipid-Based Nanosystems

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    Lipid-based nanosystems, including solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs), cationic lipid nanoparticles, nanoemulsions, and liposomes, have been extensively studied to improve drug delivery through different administration routes. The main advantages of these systems are their ability to protect, transport, and control the release of lipophilic and hydrophilic molecules (either small-molecular-weight molecules or macromolecules); the use of generally recognized as safe (GRAS) excipients that minimize the toxicity of the formulations; and the possibility to modulate pharmacokinetics and enable the site-specific delivery of encapsulated payloads. In addition, the versatility of lipid-based nanosystems has further been demonstrated for the delivery of vaccines, the protection of active cosmetic ingredients, and the improvement of moisturizing properties of cosmetic formulations.Lipid-based nanosystems are well established and there are already different commercially approved formulations for various human disorders. This success has paved the way for the diversification of the pipeline of development, to address unmet medical needs for several indications, such as cancer, neurological disorders, and autoimmune, genetic, and infectious diseases.This Special Issue aims to update readers on the latest research on lipid-based nanosystems, both at the preclinical and clinical levels. A series of 15 articles (six reviews and nine studies) is presented, with authors from 12 different countries, showing the globality of the investigations that are being carried out in this area

    Engineering CAR-T cells to overcome the immunosuppressive microenvironment of solid tumours

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    Chimeric antigen receptor (CAR)-T cell therapies have revolutionised the treatment landscape for cancer patients. However, engineered adoptive lymphocyte-based therapies face significant resistance in solid tumour microenvironments due to amino acid and nutrient scarcity, as well as significant infiltration of suppressive myeloid cells. Here, we investigated the microenvironment of different solid tumours and assessed the detrimental effects of myeloid-derived suppressor cells (MDSCs) on the autologous and engineered anti-cancer immunity. We demonstrated a novel strategy to improve CAR-T cell efficacy and tumour clearance by depleting MDSCs with a repurposed CD33-directed immunoconjugate, Gemtuzumab Ozogamicin. In addition, we designed metabolically enhanced CAR-T cell constructs, to endow the T cells with increased L-arginine catabolic activity. We found that arginase-transduced CAR-T cells were able to recognise and lyse target cells in a comparable fashion to the anti-GD2 control. However, they presented an enhanced bioenergetic flexibility, evidenced by the increased maximal respiration achieved during the MitoStress test (p = 0.010), and a higher intracellular abundance of key metabolites, such as pyruvate (p = 0.028) and glutamine (p = 0.003). Ultimately, the novel CAR-T cells were shown to have a proliferative advantage upon antigen stimulation (p = 0.015) and induce superior tumour reduction in vivo (p = 0.010) compared to the standard CAR-T cells

    Investigating predation by the predatory bacterium Bdellovibrio bacteriovorus at the interface between predatory bacteria, pathogenic prey, and the host immune response

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    Bdellovibrio bacteriovorus is a Gram-negative predatory bacterium that is able to prey upon and kill other Gram-negative bacteria, including bacterial pathogens that are resistant to antibiotic treatment. This suggests that B. bacteriovorus, or components of its predatory lifecycle, may be of great use in the continued treatment of bacterial infection, as novel antimicrobial therapies. However, before B. bacteriovorus can be applied to these scenarios, we must understand how the eukaryotic immune response may alter the efficacy of predation. B. bacteriovorus has been postulated to be less immunogenic than other Gram-negative bacteria, in part due to its outer membrane containing a modified Lipid A head group that reduces detection by host pattern recognition receptors and interactions with bactericidal antimicrobial proteins, two key components of the innate immune response. However, the interactions between B. bacteriovorus and the cells of the immune system have still not been fully characterised. In the first part of my PhD, I aimed to characterise how predation by B. bacteriovorus proceeds, and may be affected, within a host. Building on previous lab findings, I discovered that, in the context of large changes in transcription, transcriptional upregulation of some major surface components by the pathogen Serratia marcescens, whilst in human serum, led to resistance to predation. The main outer membrane component implicated in this resistance period was Lipid A, which had been modified with an L-Ara(4)N sugar to reduce its negative charge. Disruption of this LPS modification pathway, through directed gene deletion, did delay the development of resistance to predation in serum, but did not abrogate it entirely, suggesting that other surface components or factors also contribute to this resistance phenotype. This work demonstrates that host environmental factors can modify Gram-negative pathogen behaviour and therefore impact the efficacy of predation by B. bacteriovorus. In the second part of my PhD, I focus on the interactions between B. bacteriovorus and macrophages, which are a key component of the mammalian innate immune system, asking what molecular factors allow B. bacteriovorus to temporarily survive for approximately 24 hours inside macrophages. This is relevant to understanding and enhancing the efficacy of predation within a human host. Focusing on genes related to oxidative stress tolerance, that are involved in prolonging the survival of pathogens within the phagosomes of macrophages, I asked whether these genes contribute to the temporary tolerance of phagosomal conditions, extending Bdellovibrio survival, and whether these genes also play a role in tolerating the oxidative stresses experienced by Bdellovibrio throughout predation. I discovered that two alkyl hydroperoxide reductase proteins and a “survival associated” chaperone protein were essential for predation, whilst a single superoxide dismutase (SodC) enzyme contributed to both predation and macrophage survival. Finally, I take a wider view on how Bdellovibrio is perceived and processed by macrophage of the host immune response asking, through investigation of the macrophage transcriptional response to engulfed Bdellovibrio. whether Bdellovibrio is recognised, phagocytosed, and destroyed in a similar way to well-characterised Gram-negative bacterial pathogens. I discovered that, although Bdellovibrio does induce a proinflammatory immune response and is subsequently killed by host macrophage after 24-48 hours, no discernible transcriptional response is initiated towards the LPS of Bdellovibrio, in stark contrast to the detection of other Gram-negative bacteria, where LPS detection forms a cornerstone of the initial immune response. This may, in part, explain the relatively low immunogenicity of Bdellovibrio seen in animal studies. This understanding can inform future applications of B. bacteriovorus as a novel antimicrobial therapy within a human host. The work presented in this thesis has further characterised the immune response to Bdellovibrio and informs us on potential considerations of, and barriers to, predation within a human host

    Current Insights on Lipid-Based Nanosystems

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    Lipid-based nanosystems, including solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs), cationic lipid nanoparticles, nanoemulsions, and liposomes, have been extensively studied to improve drug delivery through different administration routes. The main advantages of these systems are their ability to protect, transport, and control the release of lipophilic and hydrophilic molecules (either small-molecular-weight molecules or macromolecules); the use of generally recognized as safe (GRAS) excipients that minimize the toxicity of the formulations; and the possibility to modulate pharmacokinetics and enable the site-specific delivery of encapsulated payloads. In addition, the versatility of lipid-based nanosystems has further been demonstrated for the delivery of vaccines, the protection of active cosmetic ingredients, and the improvement of moisturizing properties of cosmetic formulations.Lipid-based nanosystems are well established and there are already different commercially approved formulations for various human disorders. This success has paved the way for the diversification of the pipeline of development, to address unmet medical needs for several indications, such as cancer, neurological disorders, and autoimmune, genetic, and infectious diseases.This Special Issue aims to update readers on the latest research on lipid-based nanosystems, both at the preclinical and clinical levels. A series of 15 articles (six reviews and nine studies) is presented, with authors from 12 different countries, showing the globality of the investigations that are being carried out in this area

    Proceedings of 14th international symposium Modern trends in livestock production

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