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In cancer therapy, tumour hypoxia is an important predictor of disease progression, treatment outcomes, and metastatic potential. To date, methods for reducing tumour hypoxia have delivered limited success owing to either a lack of proven efficacy, unwanted side effects, or logistical limitations. Therefore, there remains an urgent requirement for an effective hypoxia reduction mechanism that is safe, cost effective, biocompatible, and easy to use. To address this, oxygen-filled micro and nanobubbles have been investigated as a method of oxygen delivery and have shown promising results in improving therapeutic response. To develop and optimise oxygen carriers and other hypoxia reduction strategies, however, consistent methods for measuring changes in tissue oxygenation need to be established. The ideal method should be noninvasive and quantitative, allowing tumour oxygen measurements to be obtained before, during, and after treatment. This thesis examines whether clinically available MRI techniques can be used to detect the change in tissue oxygenation caused by the nano-carriers. A combination of phantom and preclinical experiments, mathematical modelling, and the analysis of data from two clinical trials are used. First, original results from phantom and preliminary in vivo MRI experiments are reported, which demonstrate the feasibility of detecting the effects of oxygen carriers. Second, three separate mathematical models are developed and presented in order of increasing complexity 2 first addressing simple bodily fluids, then blood, and finally, tissues 2 each of which are useful for areas of MRI research beyond the focus of this thesis. Third, original research results from recently completed clinical studies of oxygen-enhanced MRI are analysed and used to assess the viability of using these MRI measurements to monitor oxygen delivery from nanobubbles in a clinical MRI setting. Lastly, the broader potential applications of the conclusions and tools presented in this thesis are discussed.

The physiological oxygen environment in vivo (physioxia) typically ranges from 1.0-12.5% O₂. However, in vitro mammalian cell culture largely utilises an O₂ concentration ([O₂]) of 18.6% O₂ (which accounts for the partial pressure of water in the incubator) during the growth stage. As such, most mammalian cells are grown in a non-physiological [O₂] during in vitro cell culture. The effect of growing mammalian cells long term in 18.6% O₂ on the subsequent cellular response to redox-active compounds compared to cells grown in physioxia is not well understood. It was shown that human non-melanoma squamous cell carcinoma cells (A431) cultured long term under 18.6% O₂ were resistant to auranofin (thioredoxin reductase inhibitor), and H₂O₂-induced cell death compared to A431 cells grown in 3.0% O₂ (physioxia in human skin). A431 cells grown in 18.6% O₂ were also resistant to H₂O₂, mercaptosuccinic acid, and cumene hydroperoxide-induced lipid peroxidation compared to A431 cells grown in 3.0% O₂. Auranofin-induced ROS generation and oxidative stress was lower in A431 cells grown in 18.6% O₂ compared to A431 cells grown in 3.0% O₂ for 96 h under the same treatment conditions. Catalase and glutathione reductase enzyme activities were higher in A431 cells grown in 18.6% O₂ compared to A431 cells grown in 3.0% O₂ for 96 h. Additionally, the expression levels of nuclear Nrf-2 protein, and NAD(P)H quinone oxidoreductase (NQO-1) protein, were about two-fold higher in A431 cells grown in 18.6% O₂ compared to the levels in A431 cells grown in 3.0% O₂ for 96 h. Auranofin treatment did not induce NQO-1 protein expression in A431 cells grown in 18.6% O₂ whilst A431 cells grown in 3.0% O₂ for 96 h showed higher auranofin-induced NQO-1 protein expression levels. These data showed that A431 grown in 18.6% O₂ exhibited resistance to auranofin and H₂O₂-induced cell death compared to A431 cells grown in physioxia under the same treatment conditions. This resistance involves, in part, higher levels and activities of certain antioxidant proteins and enzymes as conferred by long term growth in standard cell culture [O₂] compared to physioxia. It is concluded that the in vitro testing of redox-active compounds on squamous cell carcinoma cells grown in 18.6% O₂ may yield artefactual in vitro data compared to such in vitro testing on A431 cells grown in physioxia. It is advised that the in vitro testing of redox-active compounds on human non-melanoma squamous cell carcinoma cells should be performed on such cells grown in physioxia for at least four days prior and not on such cells grown chronically in a standard cell culture [O₂].

Three-dimensional, porous collagen tissue engineering scaffolds can be produced using freeze-drying. However, the stochastic nature of ice nucleation may lead to architectural variability between structures created using nominally similar preparation conditions and this can impact subsequent tissue ingrowth and function. Another common problem in tissue engineering is inadequate nutrient supply to the scaffold centre, which can lead to imbalanced cell distribution and tissue necrosis. The aim of the thesis was to address these issues by investigating three different aspects of ultrasound application on the performance of collagen-based tissue engineering scaffolds. The work has addressed underpinning research questions, covering scaffold fabrication, the stimulation of cell migration within the structure, and the ability to provide targeted oxygen delivery. Collagen scaffolds were fabricated by freeze-drying and crosslinking using N-(3-Dimethyl-aminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) with N-Hydroxysuccinimide (NHS). Scaffolds were freeze-dried at -20 °C and crosslinked with a 15:6:10 molar ratio of EDC, NHS and collagen carboxylic acid groups respectively. Micro-computed tomography (MicroCT) was used to characterise scaffold architecture and to quantify the pore dimensions. Pore size variability was noted through the thickness of the scaffold which, for consistency, required sample trimming before further testing. For this reason, a possible method to reduce the stochastic nature of the ice nucleation was sought. Ultrasound was applied to supercooled collagen suspensions (40 kHz frequency, 0.2 W cm−2 acoustic intensity) to investigate whether ice nucleation could be stimulated. It was found that the technique was an effective trigger, as characterised by an instantaneous increase in temperature to the equilibrium freezing temperature. It was also noted that the nucleation temperature correlated with pore diameter. The addition of 0.25 mg ml−1 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)/polyethylene glycol-40 (PEG-40) stearate phospholipid microbubbles in collagen slurry produced significantly smaller pore sizes (p ≤ 0.05) with mean diameter of 84 ± 4 µm, compared to 102 ± 6 µm without microbubbles. The presence and impact of cavitation were confirmed through imaging of ultrasonic nucleation with microbubbles. The work confirmed that ice nucleation could be controlled, as hypothesised, but it was noted that the concomitant reduction in pore size would not be advantageous for cellular infiltration into the scaffold. However, the cavitation caused by ultrasound offered further possibilities to influence cellular behaviour within the scaffold. The effects of cavitation and local pressures produced by high-intensity focused ultrasound (HIFU) on directional migration in human dermal fibroblast (HDF) culture were investigated by measuring cavitation response and cell distributions. HIFU was applied (0.5 MHz) to HDF cultures with 0.25 mg ml−1 polystyrene cavitation agents in collagen scaffolds for one, three or five days consecutively to investigate the impact of repeated HIFU exposure. HDF culture was seeded at one end of the scaffold, with ultrasound applied parallel to the length at the same end. After 5 days, HDF cells were distributed 15% further along the scaffold length, parallel to the direction of ultrasound, relative to the control samples (p ≤ 0.05). It was also noted that, as HIFU exposure increased, total cell counts decreased (p ≤ 0.01). For this reason, methods were considered to increase cell viability via targeted oxygen delivery using ultrasound. The feasibility was explored of using DSPC/PEG-40 stearate phospholipid microbubbles as oxygen delivery vehicles. Oxygen release was measured during ultrasound exposure (40 kHz, 0.3 W cm−2) and the effects on cell metabolism and cell viability in HDF and HT1080 populations cultured in collagen scaffolds were investigated. Oxygen-loaded microbubbles were delivered to the centre of the cell cultures, using either 10 minutes or 30 seconds of exposure to ultrasound, daily, for 10 days. While the metabolism of both HT1080 cells and HDF cells was reduced, and the viability of HT1080 cells also decreased, the viability of HDF cells was enhanced. Scanning electron microscopy (SEM) revealed that HT1080 cells exhibited less cell binding with ultrasound, and cavities were produced in the collagen structure due to cavitation pressures. This work established the use of an oxygen-releasing biomaterial with targeted characteristics to improve cell viability of 3D HDF culture and inhibit viable HT1080 development. The work carried out in this thesis developed three distinct approaches with ultrasound to improve current techniques used in collagen-based tissue engineering. The results from experiments using different acoustic frequencies and intensities have advanced knowledge across the areas of scaffold production, cell migration and oxygen delivery.

Despite the significant improvements in Assisted Reproductive Technologies (ART) there is an extensive area that remains unknown in regards to the effect of embryo culture on the long-term health of the offspring. The preimplantation embryo has specific requirements depending on the developmental stage; however, it remains unclear how the different compositions of commercial culture media can affect embryo development. The in vitro factors that most influence embryo health include cryopreservation methods, culture media components, and oxygen levels. Several studies have reported alterations in birth weight, and a higher risk of congenital anomalies when comparing between assisted and naturally conceived babies. Others have reported effects on epigenetic regulation, gene expression and imprinting, suggesting that embryo culture medium may have a potential impact on the further development of the newborn. Nevertheless, the most commonly reported alterations in offspring health outcomes include foetal growth, birth weight, childhood growth and long-term diseases such as metabolic, cardiovascular, and neurological. Due to the number of reports indicating the possible effect of the environment on embryo development, it is crucial to be aware of the mechanisms involved in order to prevent embryo alterations and long-term diseases. Therefore, the objective of this thesis was to determine whether specific components of embryo culture such as glucose and oxygen had any effect on the epigenetic patterns, at transcript levels, as well as on mitochondrial function. The analysis of transcripts was assessed to determine differential expression of genes related to pluripotency, mitochondrial function, apoptosis, oxidative stress, DNA methylation, glucose transporters, among others. The epigenome analysis (Bisulfite Sequencing) was carried out to determine differential DNA methylation levels on human blastocyst cultured in high or low glucose (0.9mM or 3.5mM). Finally, mitochondrial function was evaluated through the measurement of mitochondrial membrane potential (MitoProbe™ JC-1 assay) and through the analysis of mitochondrial respiration (Seahorse XF Cell Mito Stress Test) by directly measuring the oxygen consumption rate (OCR). In order to accomplish the objectives of this project and to obtain the most relevant clinical data, this research was conducted using preimplantation human embryos at blastocyst stage (day 6) and a model of human embryonic stem cells (HESCs). HESCs are pluripotent stem cells derived from the inner cell mass (ICM) of human blastocysts. The combination of the analysis on human embryos and HESCs allowed the validation of initial protocols, set up experimental conditions and the evaluation of different culture conditions. Firstly, I evaluated combined oxygen levels (5% and 20%) and glucose concentration (5mM and 17.5mM) on HESCs to determine their effect on transcript levels and mitochondrial function. Findings revealed altered transcript levels and mitochondrial function when cells were exposed to different concentrations of either oxygen or glucose, as well as to the combinations of both. However, the most significant effects were observed with changes in glucose concentration. Secondly, oxygen levels (5% and 20%), glucose concentration (0.9mM and 3.5mM) and the effect of cryoprotectants were evaluated on human blastocysts to determine their effect on transcript levels, mitochondrial function and DNA methylation. Findings revealed altered gene expression due to changes in oxygen but more significantly due to changes in glucose. Also, the analysis of DNA methylation showed significant differences as a result of glucose concentration. Furthermore, for the evaluation of mitochondrial function, the analysis of membrane potential was only carried out on blastocysts, treated/untreated with cryoprotectant. On the other hand, the measurement of OCR for mitochondrial respiration was only possible to determine the baseline on fresh blastocyst but not to make a comparison between culture conditions due to technical limitations. Overall, these findings suggest that altered culture conditions during early embryo development have the potential to affect several pathways that could be linked to the development of diseases. This project provides the first evidence about specific components of the embryo culture that could alter embryo development. Therefore, the conditions evaluated in this project are worth further investigation to elucidate the altered mechanisms and their direct association with the development of diseases.

Hypoxia is an adverse prognostic feature in cancer, conferring resistance to treatments including immunotherapy. In this thesis, image analysis of human tumour histology samples and murine models is utilised to evaluate some aspects of the interactions between tumour oxygenation, cellular oxygen sensing and immunity. In Chapter 1, a novel image analysis pipeline is validated and utilised to analyse the distribution, relative to hypoxia, of tumour infiltrating leukocytes in pimonidazole-treated head and neck squamous cell carcinomas before comparing the observed distributions to those in clear cell renal cell carcinoma, a tumour characterised by perturbed oxygen sensing. Leukocyte-specific distribution patterns were shared by both tumour types, with lymphoid cells largely restricted to better oxygenated perivascular regions and myeloid cells being more diffusely distributed including within profoundly hypoxic necrotic regions. In Chapter 2, this pipeline is used to evaluate immunohistochemically labelled hypoxia, vascular and immunological markers in lung cancer samples from participants in a clinical study evaluating whether atovaquone, an oxidative phosphorylation inhibitor, reduces tumour hypoxia. Although scoring of all markers was equivalent between groups, given that atovaquone-treated tumours were more hypoxic at baseline, this may represent a meaningful finding. Chapter 3 concerns histological phenotyping of transgenic mice in which hypoxia-inducible factor (HIF) was up-regulated through reversible prolyl hydroxylase domain-containing protein 2 (Phd2) gene silencing, leading to regulatory T lymphocyte dysfunction and autoimmunity. Given that regulatory T lymphocytes suppress anti-tumour immunity, in Chapter 4, the effects of host Phd2 silencing are assessed in murine tumour models. Disappointingly, this intervention affected neither tumour growth nor immune contexture.

Cambridge was a well-established town and an important trade centre in the High and Late Medieval period (11th-16th century). Historical accounts and archaeological evidence indicate that the Late Medieval period was a tumultuous chapter in English history, with episodes of war, climate change, famine and plague. It is argued that the social and economic changes, particularly those associated with the Black Death of 1348-50, would have significantly affected food production and procurement in England and that dramatic population reduction may have resulted in changes in population replacement and diversity. However, it is not fully understood to what extent events in the Late Medieval period affected towns like Cambridge, and whether they were experienced universally across the local population. This thesis utilises contextualised isotope analysis of human and faunal skeletal remains from cemetery sites excavated from Cambridge and its hinterlands to investigate what life was like for the population living in the High and Late Medieval period. Isotope sampling focussed on High and Late Medieval skeletal remains, but to evaluate the scale of change across the longue durée, skeletal remains dating from the Neolithic to the Post-Medieval period were also sampled. A multi-tissue (bone, dentine, enamel) multi-isotope (δ13C, δ15N, δ18O, 87Sr/86Sr) approach was used to understand how diet and mobility varied at an intra- and inter-individual and population scale. Collagen from 246 faunal bones, 227 human tooth roots and 314 human ribs was analysed for δ13C and δ15N, carbonate from 295 tooth crowns was analysed for δ18O and δ13C, and 170 tooth enamel samples were analysed for 87Sr/86Sr. Successful results were combined with previously generated data from the area, totalling 252 faunal isotope results and 1174 human isotope results. The results of the isotope analysis have allowed for the characterisation of diet across multiple sites, indicating variation based on social status, religious beliefs and locality. Isotope analysis has identified a small number of individuals who were unlikely to have spent their childhood in Cambridge and may have migrated into the town. Potential localised, short-distance migration in women has also been identified. Most importantly, this thesis concludes that the data does not show conclusive temporal changes in the High and Later Medieval periods which could be linked to events in the 14th century. Some general temporal change in diet may be present, but this is likely to be linked to the increasing popularity of marine fish consumption following the 'fish event horizon'.

Oxygen is required for life and is a major determinant of mammalian cell fate. Vital systems are driven by the activity of oxygen-dependent enzymes within fundamental processes, such as cellular metabolism and gene transcription. The hypoxia-inducible factors (HIFs), which are regulated by oxygen-sensing hydroxylases, have a central role in maintaining oxygen homeostasis in cells throughout the body. This work explores the role of HIF signalling in T cells and how modulation of this signalling may be harnessed to potentiate the immune response against cancer. The thesis is divided in two parts, each considering a key aspect of HIF activity: i) the metabolic consequence of transcriptional activity downstream of HIF and ii) the metabolic control of the regulators lying upstream of HIF. Chapter 2 describes the discovery of HIF1-dependent modulation of vitamin B6 metabolism via pyridoxal phosphate phosphatase (PDXP), and the effect of pharmacological targeting of vitamin B6-dependent enzymes in primary and malignant T cells. Vitamin B6-dependent enzymes are shown to be essential for the proliferation and effector differentiation of T cells in vitro and required to support T cell expansion and effective anti-tumour responses in vivo in mice. These findings highlight HIF1-dependent vitamin B6 metabolism as a key modulator of T cell fate and a promising potential target to improve cancer immunotherapy. In Chapter 3, the role of factor inhibiting HIF (FIH) in directing T cell metabolism and fate is explored. Using a mouse T cell-specific FIH knockout model, FIH is shown to regulate T cell differentiation in an oxygen-dependent manner. Furthermore, by considering a metabolic network of related enzymes that compete for the same cosubstrates, FIH activity is predicted, and demonstrated, to be optimal under conditions where oxygen levels are non-limiting and HIF levels are maximised. The therapeutic benefit of targeting FIH to limit in vivo tumour growth in mice is evaluated by deleting FIH in both T cell and tumour cell compartments. Taken together, these findings describe a dynamic metabolic feedback loop in which HIF activity modulates pathways that are critical to T cell proliferation and differentiation, and in turn is regulated by metabolic competition between HIF hydroxylases and other cell fate-determining hydroxylases. This interdependence allows for amplification of targeted metabolic alterations via downstream transcriptional responses as a strategy to improve anti-tumour T cells function.

Zirconium alloys are widely used in light water reactors as fuel cladding, acting as the primary barrier between the nuclear fuel and the coolant, due to the excellent neutron transparency, acceptable corrosion resistance and good mechanical properties of the zirconium. While in service these alloys degrade due to exposure to high temperature aqueous environments and strong radiation fields. As these Zr alloy components often perform critical safety roles, their degradation in service must be understood. In this thesis I use the capabilities of the NanoSIMS 50L, a magnetic sector mass spectrometer capable of nanoscale lateral resolution, to analyse the distribution of hydrogen and oxygen in corroded Zircaloy-4 and Zircaloy-2 and the impact that irradiation has on oxide growth and distribution of oxygen in the oxide layer. In this study an array of H+ irradiated samples was produced. Two batches of these samples were oxidised in simulated pressurised water reactor coolant for 52 and 131 days before being irradiated at the Dalton Cumbrian facility at 350 °C with 1 MeV H+ ions using the BABY beamline to doses of 0.25 and 0.75 dpa. These samples were then returned to an autoclave isotopically spiked with 5% H2 18O and 50% 2H2O at 320 °C for forty days. This produced an array of samples with oxides that were pre-transition (92 days) and at the point of transition (171 days). Alongside these samples were unirradiated samples, isotopically spiked with 2H, that were made available by Jacobs plc. Samples were analysed with NanoSIMS and correlative techniques and the results of these analyses are presented in three papers. In the first of these papers I show that oxygen transport across oxide layers is dominated by transport along cracks and pores, while hydrogen diffusion appears to be lattice diffusion based and limited in extent. Additionally, I apply a technique to image areas sequentially with the NanoSIMS that allows for analysis of more material by an order of magnitude than typical analysis. In the second paper I show that hydrogen segregates to the interfaces between secondary phase particles, most likely Zr(Fe,Cr)2 intermetallics, in the zirconium base metal in Zircaloy- 2 and Zircaloy-4. Additionally, I show that prolonged analysis with NanoSIMS leads to the accumulation of subsurface irradiation damage in the sample which destroys hydrogen trapping sites. In the final paper, which focusses on the impact of irradiation on oxygen diffusion and zirconium oxide growth, I show that the impact of proton irradiation varies greatly depending on the oxide thickness during irradiation. For pre-transition oxides the oxide growth rate and oxygen diffusivity are increased, but this leads to inhomogeneous oxide growth. When oxides approaching transition are irradiated this leads to increased cracking and spallation of the oxide layer.

The chemical weathering of silicate rocks with carbonic acid is thought to play an important role in the consumption of atmospheric carbon dioxide (CO₂), which regulates global climate over million-year timescales. However, the climatic implications of chemical weathering of carbonate rocks with sulfuric acid, a process that can release geologically stored carbon to the atmosphere, are not thoroughly understood. Depending on the reaction environment the lithologically-sourced carbon released from the sulfuric acid weathering of carbonates can either degas as CO₂ instantaneously to the atmosphere, or can be transferred as bicarbonate (HCO₃⁻) to the oceans to be precipitated as carbonate, releasing CO₂ on million-year timescales. It is important to consider the timescale of CO₂ release to assess whether a river basin is a transient, or long-term source of carbon to the atmosphere. Few studies have highlighted the importance of this weathering reaction and less have quantified the impact of CO₂ release from sulfuric acid weathering of carbonates in large scale catchments. Quantifying carbonate weathering with sulfuric acid requires the source of riverine sulfate (SO₄²⁻) to be determined. This comes predominantly from two sources: sedimentary sulfate and sulfide. Sulfate released from the weathering of gypsum or anhydrite plays no role in the carbon cycle. Oxidative weathering of sedimentary sulfides, predominantly pyrite, produces sulfuric acid which can react with carbonates to release CO₂. Here, new coupled sulfur, δ³⁴S_SO₄, and oxygen, δ¹⁸O_SO₄, isotope data on dissolved riverine sulfate and river water isotopes, δ¹⁸O_H₂O, from one of the world's largest rivers, the Mekong in Southeast Asia, are presented. A new two end member mixing model is used to partition sources of dissolved sulfate. Importantly, sulfate sources cannot be distinguished using δ³⁴S_SO₄ alone, and hence δ¹⁸O_SO₄ must also be determined. The Mekong is the world's 10th largest river in terms of discharge, yet is disproportionately understudied compared to many other large rivers where chemical weathering rates have been investigated. This study sampled 36 tributaries and 10 locations along the main channel to calculate the carbon budget of the Mekong River. Samples were collected over three field seasons, as well as a bi-monthly time-series from 2014-2017. HCO₃⁻ and SO₄²⁻ concentrations in the Mekong are up to 2709μmol/L and 720.3μmol/L respectively, and generally decrease downstream from the headwaters on the Tibetan Plateau. Samples display up to 20.7‰ difference in Δ¹⁸O_SO₄⁻H₂O, and δ³⁴S_SO₄ ranges by ~ 13.5‰ over the basin. The proportion of sulfate derived from the oxidative weathering of pyrite, f_pyr, calculated by the two end member mixing model varies between 0.18 and 0.83 in Mekong tributaries with a mean of 0.60 in the main channel. Sources of dissolved inorganic carbon (DIC) are partitioned using a forward model which incorporates the source of acidity in weathering reactions (carbonic or sulfuric) and the use of individual Ca/Na and Mg/K ratios in silicate end members of each tributary. Charge balance calculations with partitioned cations suggest the most likely reaction pathway of CO₂ release from sulfuric acid weathering of carbonates is instantaneous degassing. The weathering of carbonates by carbonic acid accounts for on average 81% of the total DIC flux for all tributaries, whilst carbonic acid weathering of silicates contributes on average 19% (ranging 7-60%) of the total DIC flux in tributaries. Using a framework to track partitioned Ca²⁺ ions, CO₂ consumption or release in the Mekong basin is shown graphically. On timescales shorter than carbonate precipitation, all Mekong tributaries are a sink of atmospheric CO₂. Annually, the instantaneous release of CO₂ from oxidative pyrite--driven weathering of carbonates is ~ 16 times smaller than the drawdown of atmospheric carbon by carbonic acid weathering of both silicate and carbonate minerals. On million-year timescales, the headwater regions in China and one karst dominated tributary in the Middle Mekong release CO₂. The carbon budget of the Mekong varies throughout the year: during the monsoonal months the Mekong consumes atmospheric CO₂ but CO₂ is released during the dry season. Carbonic acid weathering of carbonates is carbon-neutral on million-year timescales, whereas the release of CO₂ from sulfuric acid weathering of carbonates is large enough to marginally offset the long-term sequestration of atmospheric CO₂ by carbonic acid weathering of silicates. Time-series samples and discharge measurements at Chroy Changvar, close to the mouth, are used to determine that the Mekong River basin is an annual net source of 0.01tC.km⁻².yr⁻¹ to the atmosphere. This differs to previous estimates, which indicate atmospheric CO₂ consumption by the Mekong. This thesis highlights the importance of determining the origin of sulfate in the world's largest rivers for the global carbon cycle, particularly in catchments with a high proportion of carbonate lithology.

The composition of the atmosphere has changed substantially over Earth's history, with important implications for past climate. A number of case studies will be presented which employ coupled climate model simulations to assess the strength of these chemical feedbacks on the climate. The eruption of Mount Samalas in 1257 led to the largest stratospheric volcanic injection of aerosol precursor gases in the Common Era, however climate model simulations of the last millennium typically overestimate the resulting climatic cooling when compared with tree-ring proxy records. A novel configuration of the Met Office UM-UKCA climate model is presented which couples an atmosphere-ocean general circulation model to a rigorous treatment of the relevant atmospheric chemistry and microphysical aerosol processes. This permits the climate response to a particular stratospheric injection of reactive volatile gases to be quantified and for the first time to date applied to a historical volcanic eruption. This model configuration compares favourably to observational data for simulations of the 1991Mount Pinatubo eruption. Results from an ensemble of model simulations are presented, with different assumptions about the sulfur dioxide and halogen loadings based on a recent geochemical reconstruction. These show a muted climate response, in reasonable agreement with tree ring records. Emissions of halogenated compounds lead to an increase in the sulfur dioxide lifetime, widespread ozone depletion and a prolonged climatic cooling. Strong increases in incident ultraviolet radiation at Earth's surface also occur. Oxygen levels may have varied fromas little as 10% to as high as 35% in the Phanerozoic (541Ma - Present). An increase in atmospheric oxygen increases atmospheric mass which leads to a reduction in incident shortwave radiation at Earth's surface due to Rayleigh scattering. However, this is offset by an increase in the pressure broadening of greenhouse gas absorption lines. Dynamical feedbacks also lead to increased meridional heat transport, warming polar regions and cooling tropical regions. An increase in oxygen content using the HadCM3-BL and HadGEM3-AO climate models leads to a global mean surface air temperature increase for a pre-industrial Holocene base case, in agreement with idealised 1D and 2D modeling studies. Case studies from past climates are investigated using HadCM3-BL which show that in the warmest climates, increasing oxygen may lead to a temperature decrease, as the equilibrium climate sensitivity is lower. For the Maastrichtian (72.1 - 66.0Ma), increasing oxygen content leads to a better agreement with proxy reconstructions of surface temperature at that time irrespective of the carbon dioxide content. There is considerable uncertainty in the timing of the rise in atmospheric oxygen content from values around 1% in the Neoproterozoic (1000 Ma - 541 Ma) to the 10- 35% values inferred in the Phanerozoic with respect to two global glaciation episodes (717-635Ma). Results of simulations with HadCM3-BL which investigate the impact of oxygen content on the Neoproterozoic Snowball Earth glaciations are presented. These demonstrate that a smaller reduction in carbon dioxide content is required to initiate a Snowball Earth at low oxygen content. Geological evidence suggests the presence of a basaltic large igneous province before the Sturtian Snowball Earth episode. This could have caused episodes of paced explosive volcanism, injecting sulfate aerosol precursors into the stratosphere. Results of simulations to investigate the impact of different volcanic aerosol emission scenarios are presented. 500 Tg SO2 is investigated with a range of aerosol sizes. For aerosol size distributions consistent with the aerosol evolution in the aftermath of the Mount Pinatubo eruption, the Earth enters a Snowball Earth in between 30 and 80 years. Using a larger size of aerosols, consistent with a larger eruption, does not lead to a Snowball Earth. These simulations show that changes to the chemical composition of the atmosphere, whether reactive gases or bulk chemical composition may have played an important role in the past climate of Earth.

Pre-eclampsia, fetal growth restriction and stillbirth are major pregnancy disorders throughout the world. The underlying pathogenesis of these diseases is defective placentation characterised by inadequate invasion of extravillous trophoblast (EVT) cells into the uterine decidua. This invasion is necessary to transform the uterine arteries, ensuring an adequate maternal blood supply into the intervillous space for normal fetal growth and development. The mechanisms that regulate trophoblast invasion remains poorly understood, but it is known to be influenced by a number of factors in the uterine environment. These include interactions with maternal immune cells as well as cytokines and the products from the uterine glands. In this thesis, physical factors, specifically, tissue stiffness and oxygen are studied as regulators of trophoblast invasion. The mechanical environment is known to regulate cell fate and the migratory behaviour of cells. Despite invasion of EVT cells through decidual tissue rich in extracellular matrix (ECM) proteins, there has been no study investigating how tissue stiffness might regulate EVT invasion. Oxygen has also long been investigated as a regulator for trophoblast invasion, but evidence is conflicting on whether low oxygen promotes or inhibits invasion. This is in part because of the wide variation in methods used and the over-reliance on trophoblast cell lines. To examine the effects of tissue stiffness and oxygen tension, a robust in vitro method to examine the motility and migration of primary EVT cells in three-dimensions (3D) was first established. This system offers significant benefits compared with two-dimensional (2D) systems used previously. Importantly, only cells expressing the HLAG antigen, a marker for the extravillous phenotype are analysed. The stiffness of decidual tissues at the maternal-fetal interface was determined using atomic force microscopy. In patient matched samples, a 3-4 fold increase in stiffness was found where the placenta implants into the decidua, compared to where there is no implantation. Migration of single EVT cells under different matrix stiffness and oxygen concentrations in 3D were investigated. To determine whether EVT migration is directed, a microfluidic system was established, which models the oxygen gradient at the maternal-fetal interface in the first trimester of pregnancy. This system is simple and economical to setup, and permits analysis of the migration dynamics of trophoblast cells in 3D and in real-time under different oxygen concentrations. In conclusion, the change in stiffness at the site of implantation, is further evidence of the dramatic change that occurs in the uterine wall during pregnancy. A microfluidic system to study whether primary EVT cell invasion is directed under oxygen gradients was developed.

Novel techniques, mainly three-dimensional (3D) reconstructions from image stacks and finite element analysis (FEA), were combined to study the oxygen transport mechanism in the human placenta and its relationship to placental structure. Initial research relates to the development of a platform suitable for realistic computational simulations. The work shows how the 3D architecture of a terminal villus can be accurately reconstructed from confocal laser scanning microscopic (CLSM) images. By combining the resultant 3D structures of terminal villi with finite element analysis, the diffusion of oxygen from the maternal bloodstream towards the fetal blood across the villous membrane is assessed. The results correlate with theoretical studies demonstrating that image-based computational modelling is a robust platform to explore the structure-function relationship in the placenta. Following work deals with the study of blood flow through the fetal capillary network, with particular interest in its role on the oxygen transport capacity of the terminal villi. The computational models are corroborated by a particle image velocimetry (PIV) experiment. The study shows that the variation in capillary diameter is key for effective oxygen uptake by the fetus. The fetus invests minimum energy needed for the blood to travel fast enough in order to provide oxygenated blood, but at the same time slow enough to allow for good oxygenation. This is achieved by the combination of narrow and dilated segments. Additionally, the results demonstrate that there is no vortical flow or whirling. In the subsequent work, the effect of blood properties is investigated. The calculated oxygen flux is 75 times higher than in the previous study (blood flow models), highlighting the importance of haemoglobin molecules in transporting oxygen. Fetal blood affinity is shown to improve fetal oxygen uptake by 11.5%. However, when accounting for haemoglobin concentration the data suggest that the different villous structures have a constant oxygen transport capacity. The methodology developed herein helps to elucidate the structure-function relationship in the human placenta. Additionally, 3D image-based multi-physics computational modelling is demonstrated to be a powerful tool to investigate in detail the mechanics of transport in the human placenta. This technique has the potential to enlighten on the development of pregnancy complications and serve as an in vivo diagnostic tool.

Pulse oximeters are low-cost, easy to use, and effective at detecting hypoxemia (low blood oxygen levels), a common complication of bronchiolitis, asthma, and pneumonia, the leading infectious cause of death in children worldwide. However, pulse oximeters are often unavailable in lowincome settings, and if available, often underused, yet little research investigates why. In this thesis, I examine pulse oximeter implementation in low-income settings, focusing on Kenyan hospitals as a case study, and using a mixed-methods approach. I conducted a systematic literature review, examining how pulse oximeter use with children at admission to hospital impacts health outcomes; I then conducted quantitative analyses of 28,000 children admitted to seven Kenyan hospitals to determine with which children pulse oximeters are used, and pulse oximetry's impact on treatment provision; these analyses informed the qualitative research component, for which I conducted interviews with 30 healthcare workers (HCWs) and staff in 14 Kenyan hospitals and employed theoretical frameworks to determine how HCWs decide whether to use pulse oximeters, and the barriers to pulse oximetry. I found that pulse oximeter use varies substantially between and within Kenyan hospitals over time. After adjusting for case-mix and signs of illness severity, HCWs were most likely to use pulse oximeters with children with a very high respiratory rate, indrawing and/or who were not alert; children who obtained a pulse oximeter reading were more likely to be prescribed oxygen than if a pulse oximeter was not used; and children with a reading below 90% were more likely to be prescribed oxygen than those with higher readings, suggesting that HCW decision-making is influenced by international and national guidelines. However, HCWs sometimes cannot use pulse oximeters when they intend to, because of insufficient pulse oximeter availability, largely due to inefficient and confusing procurement processes and repair delays. Furthermore, HCWs sometimes use pulse oximeters incorrectly or misinterpret their results, because of insufficient training. Pulse oximeter promotion programme planners can use the recommendations I provide to effectively target barriers to pulse oximeter uptake in low-income settings. Increased pulse oximetry implementation could enable early detection of hypoxemia, improving accurate diagnosis, and supporting prompt, effective treatment, which could help reduce mortality in children needing oxygen, in line with Sustainable Development Goal 3.

Oxygen Transport Membranes (OTMs) can drastically reduce the energy and cost demands of processes that require pure oxygen, as they offer the possibility to combine a separation unit with a chemical reactor. One of the most commercially viable applications of OTMs is the partial oxidation of hydrocarbons for syngas production. A typical OTM configuration is a sequential arrangement of layers, i.e. an inactive support, a fuel oxidation layer, a dense layer and an oxygen reduction layer. However, one of the limitations of the OTM system is the low catalytic activity and stability of the materials currently used for the fuel oxidation layer. Moreover, the traditional deposition techniques that are used for the catalysts preparation are difficult to perform, as the fuel oxidation layer is buried deeply in the structure of the OTM. To simplify the OTM fabrication and improve the catalysts activity and stability, this thesis explores the exsolution of Ni nanoparticles from two different host lattice compositions, as potential materials for the fuel oxidation layer of OTMs. The (La₀.₇₅Sr₀.₂₅)(Cr₀.₅Mn₀.₄₅Ni₀.₅)O₃ (LSCMNi5) perovskite was selected, as the first candidate material for the OTMs. During reduction, the exsolution of Ni nanoparticles from the perovskite lattice took place and enhanced significantly the catalytic activity of the material regarding methane conversion. However, these nanoparticles were oxidised during the first hours of the testing and slowly reincorporated into the perovskite structure, leading to drop in the performance. Thereafter, the (La₀.₇₅Sr₀.₂₅)(Cr₀.₅Mn₀.₄₅Ni₀.₅)O₃ (LSCMNi5) perovskite was selected as an alternative composition. When the oxide lattice was sufficiently reduced, the exsolution of Fe-Ni alloy nanoparticles occurred. The catalytic testing suggested that the Fe-Ni alloy nanoparticles on LSCFNi5 presented lower activity for methane conversion comparing to the Ni nanoparticles on LSCMNi5, but higher stability in oxidising conditions. By increasing the Ni doping on the B-site of LSCF to 15 mol%, the catalytic activity of the material regarding methane conversion was increased and exceeded that of LSCMNi5. A CH₄ conversion of 70% was achieved, which was 20 times higher than that of the initial LSCF perovskite. Therefore, by tailoring the perovskite composition and the exsolution of the Fe-Ni alloy nanoparticles, it was possible to synthesize a material for the fuel oxidation layer of OTMs, which combined the high catalytic activity of Ni and the good redox stability of Fe.

Anthropogenic climate change is one of the main challenges affecting the globe, with particular implications for the oceans. Marine climate change research has moved forward rapidly in recent years, and a range of physical model outputs are available that can be used by ecologists to help predict how species might be affected into the future. Policy makers require a level of understanding of how certain species and their ranges might change so that they can respond with sustainable management actions. This thesis aims to make use of a number of modelling techniques to explore implications of past and future conditions for marine species, and to appraise those tools that can be used under differing circumstances. Policy questions are answered relating to changes in the abundance and distribution of marine species. The links between historical climatic conditions and Barents Sea cod abundance are explored using Generalised Additive Models using data collected in the middle of the 20th century. This valuable historical data indicated that cod have temperature preferences and expand and shift their distributions based on environmental conditions. A simpler modelling technique is used to examine how oxygen conditions have changed in recent decades in the North Sea, how they might change in the future, and what implications this has for commercial fish species. The models show that oxygen conditions have improved recently and that they will not decrease to levels that result in large negative effects in the coming century. Species distribution modelling using a combination of global and downscaled model outputs shows that the UK will become more suitable for some non-native and harmful algal species in the 21st century, and less suitable for others. The model outputs contribute to the understanding of climate change effects and development of management tools to ensure the resilience of marine ecosystems into the future.

Fetal growth restriction (FGR) is a common pregnancy complication resulting in increased neonatal mortality and morbidity. The aetiology of fetal growth restriction is not fully understood, but abnormalities in placental development are, leading to abnormalities in placental structure which are thought to affect supply of oxygen to the fetus. The source of fetal hypoxia is unknown due to the difficulty in obtaining oxygenation data in the context of pregnancy using existing techniques. There is also an absence of data relating to oxygenation in FGR pregnancies. Oxygen-Enhanced MRI (OE-MRI) and Blood Oxygen-Level Dependent (BOLD) MRI permit noninvasive acquisition of data related to changes in the concentration of dissolved oxygen (pO2) and changes in hemoglobin saturation (sO2) under air- and oxygen- breathing (hyperoxic challenge).The aim of this project was to determine whether MRI methods can provide information relating to placental oxygenation in normal and FGR-compromised pregnancy, to investigate fetal brain oxygenation and to assess the potential confound of placental perfusion changes under hyperoxic challenge. After optimization of sequences in non-pregnant volunteers, similar pO2 and sO2 increases under hyperoxic challenge were seen in normal and FGR pregnancy. This suggested placental oxygenation was similar and that fetal extraction of oxygen may be a likelier cause of fetal hypoxia. Normal fetal brain oxygenation was found not to increase under hyperoxic challenge, which may be due to hemodynamic adaptation to limit cerebral hyperoxygenation. Finally, the robustness of these oxygenation results was supported by the lack of placental perfusion changes observed under hyperoxia using Arterial Spin Labeling (ASL).In conclusion, MRI methods successfully provided information on placental and fetal oxygenation in normal and abnormal pregnancy, obtaining novel data informing the aetiology of FGR and the physiology of the fetal brain.

The main aim of the work reported is the design of proof-of-concept of at point-of-use hydrogen peroxide electrogeneration from air. The experimental work discussed within this thesis explores five major areas: the kinetics of electrocatalysis, ion-pairing, change of solvent media, the electrode surface modication by a redox mediator, and the electrochemical reduction of oxygen within enhanced mass transport systems. The electrocatalytic rates and mass transport of two oxygen reduction redox meditors, viz. anthraquinone and methyl viologen, are studied in aqueous solutions. The investigation is facilitated through the use of a boron-doped diamond electrode, allowing the catalytic response to be clearly delineated from that of the direct oxygen reduction process. The use of simulation software is highlighted in combination with experimental voltammograms to extract kinetic data. Specifically, the voltammetric features, such as the `reverse' peak and the `split waves', are given particular attention. Consequently, it is possible to deconvolute the electrocatalytic reaction mechanisms. The reactivity of the viologen radical cation is comparable to the semiquinone radical anion in aqueous solution ((4.8~6)x10^9 M^-1 s^-1), but over a far wider pH range (pH 2.5 - pH 8.5). The change of local proton concentration, and sequential electron transfers play key roles here. Moreover, the reduced reactivity of semiquinone is observed upon formation of ion-pairs with tetrabutylammonium cations in alkaline solutions. The electro-reduction of oxygen and its mediated pathways are also investigated in non-aqueous media; in particular the thermodynamics, the kinetics, and mass transport involved in these processes. Through a variable temperature study in electrolytic acetonitrile solution, the oxygen dissolution is quantitatively shown to be an endothermic process. Moreover, the diffusion coeficients and concentration of oxygen upon change of acetonitrile mole fraction is also explored in water-acetonitrile mixtures. The rates of bimolecular reactions are extracted from simulation programs, involving semiquinone in anhydrous acetonitrile and viologen radical cation in ethanol, and show a 3 - 4 orders of magnitude reduction compared to that in aqueous solution. Although the solubility of oxygen is ca. 6 - 8 times larger in non-aqueous solvents, the much reduced homogeneous rates limit the electrogeneration of hydrogen peroxide in pure organic media. Novel surface modification methodologies for graphitic surfaces with covalently attached anthraquinonyl groups are studied and characterised. The anthraquinonyl-modified carbon surfaces show much reduced overpotentials required for oxygen reduction. In the final chapter, utilising the new surface modification methodology and novel designs, two gravity-feed flow cells for electrochemical reduction of oxygen in aqueous solutions are proposed and characterised, one based upon the tubular electrode geometry. The other exhibits much enhanced current conversion by using a porous reticulated vitreous carbon electrode. The latter may provide a prototype hydrodynamic system to produce dilute hydrogen peroxide solution at point-of-use.

Amperometric gas sensors are widely used for environmental and industrial monitoring. They are sensitive and cheap but suffer from some significant limitations. The aim of the work undertaken in this thesis is the development of ‘intelligent’ gas sensors to overcome some of these limitations. Overall the thesis shows the value of ionic liquids as potential solvents for gas sensors, overcoming issues of solvent volatility and providing a wide potential range for electrochemical measurements. Methods have been developed for sensitive amperometry, the tuning of potentials and especially proof-of-concept (patents Publication numbers: WO2013140140 A3 and WO2014020347 A1) in respect of the intelligent self-monitoring of temperature and humidity by RTIL based sensors. Designs for practical electrodes are also proposed. The specific content is as follows. Chapter 1 outlines the fundamental principles of electrochemistry which are of importance for the reading of this thesis. Chapter 2 reviews the history and modern amperometric gas sensors. Limitations of present electrochemical approaches are critically established. Micro-electrodes and Room Temperature Ionic Liquids (RTILs) are also introduced in this chapter. Chapter 4 is focused on the study of analysing chronoamperometry using the Shoup and Szabo equation to simultaneously determine the values of concentration and diffusion coefficient of dissolved analytes in both non-aqueous and RTIL media. A method to optimise the chronoamperometric conditions is demonstrated. This provides an essential experimental basis for IL based gas sensor. Chapter 5 demonstrates how the oxidation potential of ferrocene can be tuned by changing the anionic component of room temperature ionic liquids. This ability to tune redox potentials has genetic value in gas sensing. Chapters 6 and 7 describe two novel patented approaches to monitor the local environment for amperometric gas detection. In Chapter 6, an in-situ voltammetric ‘thermometer’ is incorporated into an amperometric oxygen sensing system. The local temperature is measured by the formal potential difference of two redox couples. A simultaneous temperature and humidity sensor is reported in Chapter 7. This sensor shows advantageous features where the temperature sensor is humidity independent and vice versa. The Shoup and Szabo analysis (Chapter 4) requires ‘simple’ electron transfer and as such the reduction of oxygen in wet RTILs can be complicated by dissolved water. Chapter 8 proposes a method to stop oxygen reduction at the one electron transfer stage under humid conditions by using phosphonium based RTILs to ‘trap’ the intermediate superoxide ions. Chapters 9 and 10 report the fabrication of low cost disposable electrodes of various geometries and of different materials. The suitability of these electrode for use as working electrodes for electrochemical experiments in aqueous, non-aqueous and RTIL media is demonstrated. Their capability to be used as working probes for amperometric gas sensing systems is discussed.

Holographic sensors are photonic layered structures contained in analyte sensitive lms that upon illumination produce monochromatic reflections (λ). The present work reports the fabrication of oxygen and ammonia sensors in Nafi on membranes and hydrocarbon and volatile organic compound sensors in poly(dimethylsiloxane) (PDMS) films. A holographic recording technique was developed to suit these materials consisting of the in situ formation of nanoparticles of 18nm average diameter and their subsequent ordered ablation with a 300mJ laser. The wavelength of the monochromatic reflections depends principally on the refractive index of the resulting layers (n) and the separation between them (Λ). Changes in these parameters are generated by the analyte-sensor interactions and their magnitude can be correlated to the analyte concentration. The strength of these interactions is determined by the thermodynamic properties of the analytes, such as the cohesive energy density (δ^2), and this, was coupled with a photonic model for the prediction of the holographic response. After exposure to different concentrations of the analytes, the kinetics of the responses were determined and the lowest detection limits (LDL) established as follows: Hydrocarbons in PDMS holograms 1% (v/v) in 3s for a range of concentrations from 0-100%; ammonia in Nafi on holograms 0.16% in 100s in the 0-12.5% range; the LDL for oxygen sensing could not be determined although the response was recorded down to 12.5% and up to 100% in 100s. Holographic sensors show competitive responses comparable to commercially available gas sensors for biomedical diagnostics and industrial process monitoring because of their facile fabrication and their shared sensing platform allowing multiplexing.

The functions of the human placenta depend on the syncytiotrophoblast (STB), a highly specialised multinucleated epithelium. STB transfers nutrients to the fetus and secretes hormones that are required to support fetal development. Throughout normal pregnancy STB is renewed by cell turnover which involves proliferation, migration and fusion/differentiation of cytotrophoblasts, balanced by apoptosis/autophagy. Dysregulation of STB renewal underlies pre-eclampsia (PE), a serious pregnancy disease, but the mechanisms remain to be elucidated. Potassium (K+) channels facilitate fusion and differentiation, cellular events critical for tissue renewal. K+ channels are regulated by partial pressure of oxygen (pO2), reactive oxygen (ROS) and nitrogen (RNS) species. K+ channel function could be modulated in PE which is characterised by altered placental pO2 and increased oxidative and nitrative stress. This thesis tested the hypotheses that K+ channels participate in STB renewal and human chorionic gonadotropin (hCG) secretion and that dysregulation of K+ channel function, by pO2 and/or elevated ROS/RNS, could contribute to the abnormal STB turnover and endocrine secretion that is a feature of PE. Three aims were pursued:1) To determine whether pO2 and ROS modulate STB hCG secretion through an effect on K+ channels. Placental villous tissue was cultured in 1%, 6% and 21% pO2 and K+ channel activity (86-rubidium (86Rb) efflux) and hCG secretion were determined. STB voltage-gated K+ channel (KV) activity and hCG secretion was greater in 21% than 6% pO2 and KV blockers reduced hCG secretion at 21% but not 6% pO2. ROS had pO2-dependent effects on both hCG secretion and STB K+ channel activity but causal links between the two remain to be established. Altered pO2 and elevated ROS in PE could modulate KV channel activity leading to altered STB hCG secretion and compromised STB renewal;2) To determine whether intermediate conductance calcium-activated K+ channels (IKCa) participate in cytotrophoblast morphological and/or biochemical differentiation. Cytotrophoblasts were isolated from normal term placentas and studied at 15/42h (mononucleate) and 66h (multinucleate) of culture. Cytotrophoblasts expressed IKCa protein, and IKCa was activated by the opener DCEBIO, or hyposmotic cell swelling, but was normally quiescent. Chronic activation of IKCa by DCEBIO inhibited cytotrophoblast multinucleation and hCG secretion, an effect which may be mediated by altered cytotrophoblast volume homeostasis. Inappropriate activation of IKCa could compromise STB turnover and volume homeostasis in PE;3) Having demonstrated that pharmacological activation of IKCa inhibits cytotrophoblast differentiation in vitro, the aim was to determine whether IKCa is activated by nitrative stress. Acute exposure to nitrative stress reversibly activated TRAM-34 (IKCa blocker)-sensitive 86Rb efflux from cytotrophoblasts. Chronic nitrative stress inhibited cytotrophoblast hCG secretion but whether this reduction is through an effect on IKCa remains to be confirmed. This suggests that under conditions of increased nitrative stress in PE, IKCa could be activated contributing to dysregulated STB renewal. Overall the results confirm the hypotheses that K+ channels participate in processes of STB renewal and that their dysregulation by altered pO2 and/or ROS/RNS could contribute to abnormal STB renewal in PE. K+ channels play a fundamental role in regulating a wide range of cellular functions and knowledge of K+ channels and their regulation in STB is fundamental to understand placental physiology and pathophysiology.