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The human skin surface is covered with a lipid film that forms the interface between the viable epidermal layers and outer environment, and influences the colonisation of bacteria. Hydrolysis of the epidermal lipid ceramide via ceramidase yields sphingosine, an antimicrobial lipid that is able to kill staphylococci rapidly. Studies of patients with atopic dermatitis have indicated a correlation between levels of sphingosine and S. aureus colonisation of human skin, yet the contribution of sphingosine to the composition of the human skin microbiome and microbiota colonisation is unclear. This study investigates the effect of the host epidermal antimicrobial lipid D-sphingosine on S. aureus and S. epidermidis, with a focus on discovering components that contribute to survival of the former. The methods RNA-Seq and qPCR were used to dissect the transcriptional responses of S. aureus Newman and S. epidermidis Tü3298 after challenge with D-sphingosine. S. aureus displayed a large set of differentially expressed genes (1331), which contrasted with a smaller set for S. epidermidis (340). There was a clearly defined stimulon that was common between the species that included pathways for energy maintenance, amino acid and ion transport and metabolism, and protein repair. Previously reported transcriptomic responses to multiple cell wall and membrane-targeting antimicrobials shared commonalities with the response to sphingosine that supports its reported mode of action at the cell surface with membrane damage. The VraSR cell wall stress stimulon was upregulated in both species. Each species displayed pronounced upregulation of a putative phosphate-specific ABC transporter system encoded by the pstS-pstCAB operons. Experiments identified a contribution of this transport system to sphingosine survival and phosphate supplementation was shown to markedly increase sphingosine survival. Previous studies by the Horsburgh group identified experimentally evolved S. aureus strains with high levels of D-sphingosine resistance after serial passage with increasing concentrations of the lipid. SNPs associated with increased resistance were determined by genome resequencing and some of these were localised to the farER genes encoding a FarR-regulated FarE (mmpL) lipid efflux transporter. An isogenic farR SNP mutant of S. aureus was generated using molecular genetics approaches and showed a 10-fold higher MIC for D-sphingosine compared with the wild type Newman strain, indicating that the purported FarER fatty acid efflux system may have the capacity to mediate transport of sphingosine. The study findings of a link between sphingosine challenge and phosphate metabolism indicates future research potential to study host spingosine-1-phosphate in staphylococcal lifecycles, during health and disease.

BACKGROUND AND AIMS: Pancreatic cancer (PC) has a poor prognosis and effective diagnostic tools and therapies are currently unavailable. This project has explored the role of T cell responses in PC patients as a potential therapy. METHODS: Serum mesothelin (MSLN) levels were tested in patients with pancreatic diseases. CD4+ and CD8+ T cell responses against full-length overlapping MSLN peptide pools were identified. CEA691, together with pre-chosen MSLN547 and WT1-126 were used to stimulate T cell lines from PC patients. Antigen-specific function and phenotype were characterised in the ex-vivo expanded T cell lines after repeated Ag exposure. Further experiments used inhibitory receptor blockade to augment the function of T cells isolated from cancer patients. RESULTS: Soluble MSLN was elevated in PC patients compared to normal controls, but could not distinguish the malignant from benign pancreatic disease. MSLN-specific CD4+ T cell responses were significantly increased in PC patients compared to controls, indicating an expansion of MSLN-specific CD4+ T cell in PC patients, and new epitopes were identified. It was possible to generate CEA691, WT126 and MSLN547-specific HLA-A*02 restricted T cell lines from PC patients. The ex vivo expanded CEA691-specific T cells demonstrated Ag-specific cytotoxicity and were able to recognize and kill HLA-A2+ CEA+ pancreatic cell lines in vitro. Blockade of PD-1 and TIM-3 further enhanced T cell function in vitro. CONCLUSION: We found that tumour associated antigen (TAA)-specific T cells were identifiable in the peripheral repertoire of patients with pancreatic cancer, and the function of these cells were preserved in some cases. Such antigen-specific function and cytotoxicity was more common in relatively early stages of the disease than the terminal stage, and PD-L1 blockade further improved the responses, suggesting that TAA-specific T cells together with checkpoint inhibition could be potential strategies for the treatment of pancreatic cancer.

This thesis addresses the mathematical modelling of end-to-end packet delay for Quality of Service (QoS) provisioning in multi-hop wireless networks. The multi-hop wireless technology increases capacity and coverage in a cost-effective way and it has been standardised in the Fourth-Generation (4G) standards. The effective capacity model approximates end-to-end delay performances, including Complementary Cumulative Density Function (CCDF) of delay, average delay and jitter. This model is first tested using Internet traffic trace from a real gigabit Ethernet gateway. The effective capacity model is developed based on single-hop and continuous-time communication systems but a multi-hop wireless system is better described to be multi-hop and time-slotted. The thesis extends the effective capacity model by taking multi-hop and time-slotted concepts into account, resulting in two new mathematical models: the multi-hop effective capacity model for multi-hop networks and the mixed continuous/discrete-time effective capacity model for time-slotted networks. Two scenarios are considered to validate these two effective capacity-based models based on ideal wireless communications (the physical-layer instantaneous transmission rate is the Shannon channel capacity): 1) packets traverse multiple wireless network devices and 2) packets are transmitted to or received from a wireless network device every Transmission Time Interval (TTI). The results from these two scenarios consistently show that the new mathematical models developed in the thesis characterise end-to-end delay performances accurately. Accurate and efficient estimators for end-to-end packet delay play a key role in QoS provisioning in modern communication systems. The estimators from the new effective capacity-based models are directly tested in two systems, faithfully created using realistic simulation techniques: 1) the IEEE 802.16-2004 networks and 2) wireless tele-ultrasonography medical systems. The results show that the estimation and simulation results are in good agreement in terms of end-to-end delay performances.

The thesis describes the development and use of ultra-fast MRI techniques to quantitatively characterise the dissolution process of controlled drug release dosage forms. Implementations and validations of two quantitative single shot RARE based magnetic resonance imaging (MRI) protocols are described. Quantitative T2 (spin-spin relaxation time constant) and diffusion weighted single shot RARE images, both with acquisition time of less than 3 minutes, were achieved by preconditioning the standard RARE sequence with a hard pulse CPMG echo train or an alternating phase bipolar pulsed field gradient spin echo (APGSTE) diffusion sequence respectively. T2-preconditoined RARE and diffusion-preconditioned RARE experiments were carried out on 5 phantoms with T2 values between 29 – 2200 ms. The optimisation of the phase encoding start value (PESV) parameter in the MRI pulse sequence was carried out, and four centric phase encoding schemes were also investigated. Phantoms imaged using the conventional RARE phase encoding scheme under the optimal PESV value of -0.2, showed a percentage error difference of 11 % for absolute water concentration (Cwat) maps, and an error of 2 % in T2. Similarly an error of less than 2 % in the molecular self diffusion coefficient (D) was found compared to the corresponding reference value. The two quantitative preconditioned RARE pulse sequences were then used to follow the dissolution of hydroxypropylmethyl cellulose (HPMC) in both static and flowing environments. Finally, a case study was carried out on a coated tablet for the purpose of examining tablet coating efficiency upon scale up, where a pilot scale coated tablet was compared to a lab scale coated tablet. Two quite different dissolution behaviours of the two types of tablets were observed.

The PhD program developed a novel tracking model to assess the stability of diet over time. The ‘Two-stage Estimation Model for Tracking Analysis’ (TEMTA) overcomes difficulties of previous tracking methods and can be widely applied to future epidemiological studies. The model was applied to the MRC National Survey of Health and Development (NSHD) to track fruit, vegetable and fat intake from ages 4-36 years and 36-43 years. Physical activity and under-reporting were also tracked for NSHD participants from ages 36-43 years old. Diet, especially vegetable and fat intakes, were shown not to track from childhood to adulthood. Diet and physical activity were tracking in adulthood but under-reporting was not. In addition, an accumulative life course model was applied to investigate the longitudinal patterns of fruit consumption and obesity. Results suggest the importance of keeping a healthy diet from an early age in life. It is indicated from this study and recent healthy diet can have immediate benefit to health. Discussions and recommendations are made in terms of the advantages and limitations of the TEMTA model, research and public health implications suggested by the tracking analysis in NSHD, and further research opportunities presented by the work.

This thesis describes the high-throughput screening of chrome/PHD domain constructs from a number of chromatin remodelers and subsequent biochemical and biophysical characterization of several constructs of human CHD3, CHD4 and CHD5. Results from NMR spectroscopy, size exclusion chromatography and small angle neutron scattering al indicate that these proteins are either misfolded, or that they form higher-order structures, and that this is likely caused by the praline rich/highly charged sequences conserved amongst the CHD3/4/5 double chromodomains. Pull-down experiments of three double PHD domain-containing constructs suggest that unmodified and methylated histone H3/H4 peptides bind to these proteins. A Drosophila homologue of human CHD3, dCHD3 which lacks the problematic sequences was then investigated. This expressed well but had low solubility. A refolding protocol was, however, successfully developed to recover dCHD3 from inclusion bodies. This protein was the only double chromodomain throughout the entire project that produced a high-quality NMR spectrum – in addition, it eluted from a gel filtration column as expected. Extensive crystallization trials were performed for all the constructs that could be produced in appreciable amounts and good purity. However, crystals formed only for the dCHD3 construct. High resolution reflections were observed, but the crystals proved not to be of the intact protein.  NMR structural studies of the human PHF8 PHD domain constructs are described – signals of 82% of the backbone residues were assigned with the missing residues most likely being unstructured.

In this thesis I use the most widely prescribed anticoagulant drug, warfarin, as a model to investigate the effect of genetic determinants on drug efficacy and safety. Following a literature review of all the genes involved in warfarin pharmacokinetics and pharmacodynamics, 35 candidate genes were selected for investigation. Two independent Swedish cohorts of warfarin-treated patients were analysed. First linkage disequilibrium maps were constructed for each gene. Selected SNPs integrated with putative functional variants were genotyped in 201 patients recruited at the Uppsala University. A panel of 216 haplotype tag SNPs were then derived to analyse an independent cohort of 1496 patients from the prospective Warfarin Genetic Study in Sweden. The two studies were analysed separately for genetic association to warfarin dose requirement (single marker and haplotypic tests).  Common SNPs in the vitamin K epoxide reductase gene (VKORC1) are significantly associated with dose in the Uppsala and WARG studies (p=1.9 x 10-15 and 6.5 x 10-100, respectively). Cytochrome P450 2C9 (CYP2C9) has been known to affect dose requirement and was confirmed in both Swedish cohorts (p= 2.3 x 10-5 and 4.9 x 10-32). The two genes together explain ~40% of warfarin dose variation. SNPs in microsomal epoxides hydrolase (EPHX1) and orosomucoid 1 (ORM1) genes do not show a broad effect but are associated with dose in both studies. Genes encoding PROC, APOE, CALU, PDIA2 and GGCX showed nominal association with dose in the Uppsala study. Likewise, PROS1, CYP1A1, CYP3A4, PDIA5, PDIA3 and F10 showed nominal association to dose in the WARG study. Most of these minor effects if real are most likely to be population/treatment specific. A model taking in to account genetic factors (VKORC1 and CYP2C9*2/*3) and non genetic factors (age, gender, and drug interaction) together explained more than 50% inter-individual dose variance. We analysed 64 patients from the Uppsala and WARG studies with recorded severe bleeding episodes using the same 216 common SNPs. Case-control analysis found SNPs in PDIA4, P4HB, and NR1I3 to be associated (p≤0.01) with bleeding. Using a recessive model, patients with a gastrointestinal bleeding sub-phenotype in the WARG cohort showed association with common variants in PD1A6 (P=0.0014, odds ratio = 6.98). We sequenced the exons of 11 of the candidate genes in 36 bleeders and 12 non-bleeders (Uppsala study). However, no high prevalence mutation was discovered.

The increasing use of granular materials in the chemical, petrochemical, food and pharmaceutical industries, both as intermediate and as end products, gives rise to the need for a better understanding of granule dissolution and disintegration (GDD). However, little is understood about the physical principles controlling the release of active ingredients from granules, a crucial knowledge for the design of product structure in order to optimise product performance. The aim of this work is to bring a scientific approach to the study of GDD in a systematic way. which has not been addressed hitherto. The work here is to highlight the testing of these hypotheses and provide insight into GDD mechanisms. The scientific concern in this thesis is to attempt to reveal the unique behaviour of GDD that is different from homogeneous material dissolution. Unlike homogeneous material dissolution, which depends mainly on the chemical properties of the solute and solvent, GDD is more complicated because the heterogeneous system involves mixed insoluble particles and soluble binder. Further complexity lies in the fact that various physical properties such as granule size, primary particle size, binder-solid ratio and internal pore structure play in influencing GDD behaviour. This thesis reports experiments and theoretical analysis of such GDD behaviour. Four hypotheses - reaction-limited consumption of binder, transport limit for binder removal, transport limit for particle removal and additional Hmit for particle and binder removal - were proposed to cover all of the possible GDD mechanisms. Theories were developed and experiments were set up to test the theory and give explanation of various limiting cases of GDD. A bimodal population balance model was developed. Furthermore, a dynamic shrinking-shell and core model was proposed to clarify GDD mechanisms. The analysis, which has its origin in comparing binder and particle transport coefficients, enables prediction of whether particle or binder transport will hmit GDD process. This evolved into a Sherwood number analysis, which provides a direct physical account of the controlling step. Based on a non-synchronous binder and particle transport analysis, a shrinking shell and core model was proposed to highlight that binder transport may be limited by internal particle shell build-up in the granule. A dynamic model was then developed to provide quantitative analysis of the binder Sherwood number, which was tested by experiment. A rigorous deduction of the model provided a clear physical meaning of the parameters in the equations. To demonstrate the theory and test hypotheses, an ionic binder, zeolite-NaLAS was used in high shear granulation, low shear granulation, fluidised bed granulation and spray drying granulation to provide a range of granule structures and to allow investigation of GDD by both PSD and electrical conductivity. Granules were characterised using mercury porosimetry, sieving, bulk density, particle size analyser and SEM to provide information concerning their various properties, such as binder content, internal pore structure and volume fraction, granule size and primary particle size distribution, as well as surface morphology and granule internal structure. Two main methodologies were adopted to investigate GDD behaviour including a): monitoring electrical conductivity and granule diameter simultaneously for a single granule; b) monitoring the evolution of the PSD by laser light scattering. Those results were analysed to support the theory. In general, the phenomena in GDD are quite different from the general cases of dissolution.

Three examples of application of the anti-integrability concept in Lagrangian systems are proved, concerning the continuation of a class of trajectories from the anti-integrable limit. All three examples were proposed by Robert S. MacKay. The first example arises in adiabatically perturbed systems. With an assumption that the adiabatic Poincaré-Melnikov function has simple zeros, we constructed a variational functional whose critical points give rise to a sequence of homoclinic trajectories for the unperturbed Lagrangian in the adiabatic limit but a sequence of multi-bump trajectories under perturbations. We found there is a compact set, which is a Cantor set, such that the Poincaré map induced by the phase flow restricting to it is conjugate to the Bernoulli shift, in our case, with three symbols. Hence the approach of the anti-integrability to the adiabatically perturbed problems is equivalent to the one which combines the Poincaré-Melnikov method and the Birkhoff-Smale theory. The second example occurs in the Sinai billiard system. The anti-integrable limit is the limit when the radius of the scatterer-disc goes down to zero, and the system becomes "δ-billiards". The orbits of the δ-billiards are the anti-integrable orbits which are piecewise straight lines joining zero-radius discs to discs, and are easily obtained. Under some non-degeneracy conditions, we proved all anti-integrable orbits can be continued to the small radius case, and found that any periodic orbit has infinitely many homoclinic orbits as well as heteroclinic orbits to any others. These exists a compact set, which is also a Cantor set, such that the billiard map restricted to it is conjugate to a subshift of finite type with an arbitrarily given number of symbols. We studied in the third example when the scatterers are approximated by repulsive potentials such as the Coulomb potential ε/r, where ε and r are non-negative numbers and r is the distance from the potential centre. In the Coulomb potential case, the anti-integrable limit is the ε → 0, and the system becomes the δ-billiard system. Then we found that the results in the Sinai billiards also hold here when ε > 0 but small. More general type of repulsive potentials were also investigated and a sufficient condition under which anti-integrable trajectories persist was given.

This thesis is concerned with the colour change and microstructural evolution of wet paint when subject of shear. When different processing shear histories are applied to certain paints, the resulting colour of the paint can vary. Paint consists of pigment particles suspended in a mixture of resin and solvent, and this multi-phase structure results in a complex microstructure. For this reason there can be many possible causes of this colour change, one of which was thought to relate to the organisation of the pigment dispersion. Rheological measurements of paint were carried out to investigate whether there was a link between rheology and colour change. The measurements were performed using a Rheometrics Dynamic Spectrometer (RDS II) at low shear rates and a Multi-Pass Rheometer (MPR) at high shear rates. Although the shear ranges of the RDS II and MPR did not overlap, extrapolation of data obtained at low shear rates matched those from the MPR at high shear rates. The rheological response of the material was described using a viscoelastic K-BKZ integral constitutive equation in combination with a Wagner damping factor. Systematic tests under controlled conditions were successfully performed, and the results of three paint samples were compared. The experimental findings indicated that the observed colour change behaviour of paint was related to the modification of size and size distribution of pigment agglomerates, both during and after shear. In order to correlate pigment size with the resulting paint colour, an attempt was made to model these experimental findings. A mathematical model was developed on the basis of the Mie scattering theory. This model was verified by a case study, and its prediction gave semi-quantitative agreement with the experimental findings.

Research in High Voltage Thin Film Transistors (HVTFTs) has been driven by the need for devices with reduced on-state resistance and high blocking capability to improve the performance of large area electronic applications. Conventional HVTFTs give unsatisfactory performance because of the high on-state resistance, low breakdown voltage (Offset Drain HVTFT), high possibility of oxide failure and requirement for extra external bias line (Metal Field Plate HVTFT) etc. This thesis presents a novel high voltage thin film transistor structure - Semi-Insulating Field Plate HVTFT (SIFP HVTFT) which utilises a semi-insulating layer as the field plate to modify the conductivity in the offset region. The new structure has demonstrated and enhanced on-state performance relative to conventional offset drain device and a much improved blocking capability compared with all existing high voltage thin film transistor structures. Unlike conventional offset drain TFTs, during the "on-state", the channel formed under the gate can be extended into the offset region by the potential on the semi-insulating field plate which is controlled by the bias on the gate and the drain. Equivalent circuit models for the SIFP HVTFT have been developed for the device analysis. New concepts such as "extended channel" have been proposed for the first time to illustrate the device physics underlying the improvement in performance of the new device. The superior blocking capability is attributed to a very small leakage current flowing through the semi-insulating field plate which increases the radius of potential curvature near the drain. This leakage current, however, does not significantly contribute to the main current flowing through the device. It is concluded that the SIFP HVTFT presents a new concept to the high voltage thin film transistor family and has clearly shown many advantages over conventional HVTFTs.

Carpal tunnel syndrome (CTS) is a compression neuropathy of the median nerve. Patients may experience discomfort in the hand and wrist, altered sensation and functional deficit. The management of mild to moderate CTS involves local corticosteroid injection (CSI) and night splinting (NS). The aims of the studies presented in this thesis were to describe the epidemiology of CTS and to develop prognostic models to predict future likely outcomes in patients presenting with CTS in a primary care setting. Evidence for predictors of treatment effect (CSI or NS) were also explored. The estimated crude prevalence of patients presenting with CTS in UK primary care (using data from the Clinical Practice Research Datalink - CPRD) increased from 26.03 per 10,000 person years in 1993 (95% CI 25.10 to 27.00) to 36.08 per 10,000 person years (95% CI 35.45 to 36.72) in 2013. The proportion of patients having carpal tunnel release surgery (CTR) changed over the study period increasing from 19.35% in 1993 to a peak in 2009 at 29.32% and then decreasing to 27.3% in 2013. A systematic review of 16 cohort studies reporting on the course and prognosis of CTS showed that prolonged symptom duration, a positive Phalen's test, and thenar wasting were associated with poor outcome over a follow-up period of between six months and 12 years. However, not all associations were statistically significant and many studies were deemed to be at increased risk of bias (particularly relating to study attrition, confounding, and/or statistical analysis or reporting), hence the overall judgement regarding their predictive value remained inconclusive. A CPRD cohort of 91,412 patients with CTS was developed. 20.24% of the cohort had a recorded episode of CTR within 3 years of diagnosis. A prognostic model predicting time to surgical intervention was developed using Cox regression and included age, obesity, alcohol use, smoking and having multi-site pain, an inflammatory condition or a neck symptoms (adjusting for region and deprivation). The predictive capability of the model was limited, C statistic 0.59 (95% CI 0.58 to 0.59). Multivariable linear regression modelling using data from a randomised controlled trial (INSTINCTS) was used to develop a prognostic model predicting patient reported outcome at six months. The final model included the baseline Boston Carpal Tunnel Questionnaire score, the symptom severity score, and the absence of any other neck or upper limb symptom. The optimism-adjusted model calibrated poorly, overestimating the severity of outcome in patients with less severe observed CTS, and underestimating the severity of outcome in patients with more severe observed CTS. A second systematic review including four RCTs summarised evidence for predictors of treatment effect. The results from one trial indicated that the effect of CSI was larger in patients with more severe nerve conduction and baseline symptom scores. Exploratory analysis of a small set of a priori defined candidate predictors of treatment effect using INSTINCTS data suggested that at six months, CSI was less effective than NS in patients with unilateral symptoms compared to those with bilateral CTS. However, results need to be carefully interpreted given the lack of significant interaction, the possibility of a unit of analysis error and a very small sample size. In summary, patients with CTS presented in primary care with increasing frequency between 1993 and 2013. However, since around 2008, the proportion of patients receiving surgical treatment was observed to decrease, despite being considered clinically effective for most patients. Lower rates of surgery may be associated with changes in access to the procedure. This highlights the need for optimal management to be provided in primary care. Assuming the CPRD population to be representative, at least 20% of patients presenting with CTS did not respond well to their initial management in primary care. There is likely to be clinical benefit in identifying this group early in the course of their symptoms, and explore any differential treatment response, in order to better target treatment to the individual and identify those who are likely to require surgery. However, the prognostic models developed in this thesis performed poorly. It seems that the prediction of CTS is complex and potentially includes prognostic factors not measurable in CPRD or trial data. Similarly, no confirmatory evidence was found that could be used to match treatment options to individuals. Therefore, patients with CTS can be initially managed in primary care using current guidance and should be routinely followed up and referred for surgery if they fail to experience initial satisfactory improvement.

This thesis considers, from a theoretical perspective, the design of mechanisms to accomplish the objective described in the title. Two cases of this problem are considered: information as represented by data points in a machine-learning context, and information as represented by beliefs in a general prediction context. While there is significant literature on the acquisition or aggregation problems as considered separately, past approaches are often inapplicable or inefficient when considering both together. For the case of data, the thesis proposes an active procurement approach whereby data points are selectively purchased depending on their utility to the learning algorithm. A model is proposed and a purchasing scheme designed that interacts in black-box fashion with the user’s choice of learning algorithm. For a large class of problems, via a specific choice of learning algorithm, risk and regret bounds are proven as a function of budget and the “monetary difficulty” of the problem. For the case of beliefs, the thesis proposes a theory of substitutes and complements of pieces of information. In particular, this theory is used to analyze prediction markets, which are natural and popular mechanisms for simultaneously acquiring and aggregating beliefs. In addition, the thesis examines several additional problems involving information acquisition and aggregation in the fields of crowdsourcing and mechanism design with and without money., Engineering and Applied Sciences - Computer Science