VictorJuiz/thesis huggingface dataset:
Mechanisms of cone dysfunction in inherited retinal degenerations
Keywords: Ophthalmology

This thesis examines the mechanisms of cone dysfunction in two different inherited retinal diseases: Achromatopsia and Retinitis Pigmentosa (RP). The first, minor, component of the thesis explored and characterised the loss of cone function in a novel mouse model of achromatopsia (cpfl10). The second, major, component of the thesis explored the loss of cone function in a mouse model of RP. I hypothesised that the cone function lost in RP could be restored using AAV vectors to over-express down regulated cone genes. This thesis subsequently developed and tested two AAV vectors delivering CRX and OPN1LW genes to photoreceptors in RP mice.
Achromatopsia is a congenital condition characterised by absent or diminished cone function due to mutations of a single gene, typically those involved in the cone phototransduction cascade. This thesis provides a detailed phenotype of a novel cpfl10 mouse model of Achromatopsia that arose spontaneously in a local mouse colony. The causative (NM_013927)c.692G>A; p.(R231H) missense mutation in Cngb3 was also identified and explored.
Retinitis pigmentosa (RP) is an inherited retinal degeneration in which secondary loss of cones follows the initial death of genetically abnormal rod photoreceptors. Strategies to prolong cone photosensitivity or survival would offer an important treatment. Such universal interventions could be effective regardless of which genetic mutation is underlying primary rod death.
The molecular mechanisms by which cone photoreceptors lose photosensitivity in RP are poorly understood. I examined the gene expression profile of the cone phototransduction cascade in the Rho-/- mouse model of RP and correlated these changes to declining retinal function. The cone opsins were substantially down-regulated, as was the photoreceptor transcription factor Crx. I hypothesised that the cone function lost in RP could be restored using AAV vectors to over-express these down regulated cone genes. Using an AAV gene therapy vectors, I delivered the human homologs CRX and OPN1LW to the retinas of the same Rho-/- mouse model. Both vectors successfully transduced photoreceptors and RPE cells across three different doses. Transduced retinas did not show any improvement in cone function or survival.
Whilst the observed down-regulation of cone genes correlates with cone function loss, it appears that both down-regulation and dysfunction arise from an unknown common cause. Ongoing research into mechanisms of metabolic starvation, oxidative stress and trophic factors in cone cell death offer promise for a common intervention to rescue cone photoreceptors.
Photofragmentation studies of metal ion-molecule complexes and metal oxides
Keywords: Infrared spectroscopy
Metal clusters
Photodissociation
Velocity map imaging

Gas phase metal-containing complexes provide suitable systems in which to study fundamental binding motifs between a metal ion and molecules in the absence of any solvent, support or competing charge effects. In this thesis, metal-containing species are explored experimentally using infrared resonance enhanced photodissociation (IR-REPD) spectroscopy and velocity map imaging (VMI). The experimental results are further interpreted with the aid of spectral simulations based on density functional theory (DFT). These are the first studies reported using a newly built IR-REPD spectrometer equipped with a purpose-built laser ablation source to allow for the study of single metal ion-molecule complexes. The laser ablation source is shown to efficiently produce various complexes including Rh+(CO2)n, VO2+(N2O)n and Au+(CH4)n and the IR-REPD spectrometer has been characterised against a well-studied system of V+(CO2)n complexes. In order to record the IR-REPD spectra for small metal ion-molecule complexes, an argon atom is employed as the inert messenger.
A combined IR-REPD spectroscopy and DFT investigation of M+(CO2)n complexes (where M = Co+, Rh+ and Ir+) reveals a common [M+(CO2)2] core structure for all three considered metal ions. Additional ligands, which are not directly bound to the central metal ion, experience lower perturbation as evident in the reduced blue-shift for the ligand in the outer coordination shells. A further IR-REPD/DFT study involving CO2 complexation around NbO2+ and TaO2+ ions reveals a strongly-bound core of four CO2 ligands around the MO2+ ion (M = Nb, Ta). A significant increase in the intermolecular bond distances for the second coordination sphere ligands coincides with a decrease in the calculated binding energies.
Velocity map imaging is employed to explore the rich photodissociation dynamics of VO in the vicinity of C4Σ- - X4Σ-(v',0) vibronic transitions in VO. The final quantum state distribution was observed to be strongly dependent on the intermediate vibronic state of VO via which the dissociation threshold is reached. This work provides a refined value for the VO dissociation energy of D0(VO) = 53190 ± 261 cm-1 in excellent agreement with available literature.
Streamlined synthesis of taxol analogues
Keywords: Organic chemistry
enzymatic catalysis
biocatalyisis
Late stage C-H functionalisation
oxidation with P450 enzymes
synthesis

This thesis centres on the synthesis of taxol analogues via late-stage hydroxylation with P450 enzymes. To accomplish this, the taxane core, specifically taxa-4(5),11(12)-dien-2-one, was synthesised by classical synthetic methods, and subsequently oxidised using P450BM3 mutants.
Chapter 1 introduces enzymatic catalysis, and the advantages and disadvantages of its application to organic synthesis. Additionally, an overview of taxol, including its discovery, mode of action, biosynthesis and large-scale production, and a summary of the previously reported approaches to the taxane core are described.
Chapter 2 details the problems encountered and solutions implemented when reproducing Baran’s route to taxa-4(5),11(12)-dien-2-one. Furthermore, approaches to some of its intermediates and an alternative route to taxa-4(5),11(12)-dien-2-one, which is based on Baran’s, are discussed.
Chapter 3 describes the development of a new, practical and short synthetic route to taxa-4(5),11(12)-dien-2-one which, ultimately, led to 1,3-di-epi-taxa-4(5),11(12)-dien-2-one. Additionally, the application of this route to the synthesis of a model compound and attempts to convert this racemic synthesis into an enantioselective route are reported.
Finally, the enzymatic oxidation of taxa-4(5),11(12)-dien-2-one and related molecules using P450BM3 mutants is explored in Chapter 4. A preliminary study to determine the substrate enantioselectivity of the mutants is also described, along with the biological assays of the oxidised compounds produced during the study.
Ethical business: an ethnography of ethics and multiplicity in commercial settings
Keywords: Management Studies
Ethnography
Ethical Trading
Multiple Realities
Science and Technology Studies
Ethical Business
Chimera Realities
Multiplicity
Anthropology of Business
Chimera Object
Business Studies
Modern Commercial Setting

This thesis is a study of ethics and multiplicity as found within contemporary commercial settings. Drawing on Science and Technology Studies (STS) sensibilities and ethnographic-style research, the thesis proposes that current ethical phenomena should be understood as a user-enacted chimerical object: an object that is multiple in its ontology and as much enacted by what it is, as what it is not.
This research is particularly pertinent now because the term 'ethical' has become commonplace in modern Western life, including crucially within commercial activities. In certain uses, doing ethics becomes synonymous with doing business. Despite the increasing prevalence of what is considered 'ethical business', the exploration of how the term is appropriated and enacted remains largely under-examined.
Through examination of research material gathered during extensive ethnographic studies in three self-avowedly 'ethical organisations' - an ethical start-up, an ethical confectionery company, and an ethical consultancy - the thesis addresses this research gap. By focusing on the users of ethical business, the investigation questions traditional market assumptions of homogeneity within producing organisations, the supposed linear transfer of ethical knowledge, what we can know about 'users', and the genesis of novel ethical realities. Through this questioning the thesis provides new insights on the ethical object. The thesis additionally builds upon questions of how far we can push the boundaries of what we can know about knowledge, and whether it is possible to bring the mess of investigation back into the reporting. Developing previous applications of constitutive reflexivity, the research symmetrically investigates the appropriateness of my application of STS sensibilities to ethical business as a new research area, and interrogates my thesis as an ethical object in order to address the underlying question(s) of whether 'STS means ethical business?'
Analysis of two pore channel proteins in Dictyostelium development
Keywords: Calcium channels
Dictyostelium
Cell signaling

Calcium is a ubiquitous intracellular signal responsible for controlling numerous cellular responses including development and proliferation. Calcium (Ca2+) is stored in both neutral and acidic stores and its release through gated channels has been implicated in regulating development in Dictyostelium discoideum. This thesis aims to understand the roles of the calcium channel proteins, in particular the two-pore channel proteins (TPCs), found on acidic stores in Ca2+ signalling during Dictyostelium development.
Bioinformatic analysis indicates conservation of a gene encoding an orthologue of TPC2 in Dictyostelia and, similar to plant TPCs, a Ca2+ sensing domain is predicted along with a novel potential calmodulin binding site. To investigate the role of intracellular Ca2+ channels, a series of strains was generated, disrupted in one or more of genes encoding the channels TPC2 and mucolipin (TRP-ML), predicted to be located on acidic stores, and IplA, located on neutral stores. All disrupted strains, including one lacking all three channels, are able to complete development. However, strains lacking TPC2 show a pronounced delay in early development, correlating with reduced expression of some early developmental genes. Vesicles derived from tpc2-null cells show normal Ca2+ release compared to parental cells but an increased rate of Ca2+ uptake. During early development, the pH of acidic vesicles is increased in the absence of TPC2. However development of tpc2-null cells showed increased sensitivity to weak bases in producing fewer aggregates but resistance to sodium chloride and weak bases in later development suggesting a complex role for TPC during development.
In vivo cytosolic Ca2+ responses were analysed in strains expressing an ultra-sensitive Ca2+ indicator YC-Nano 15. Growing tpc2- and iplA- cells have lower basal cytosolic Ca2+ than parental Ax2 cells. Intercellular Ca2+ waves were observed in aggregates from Ax2, mcln- and tpc2- cells but were greatly reduced in iplA- aggregates, as was the increase in cytosolic Ca2+ in response to extracellular cAMP. In tpc2- aggregates, wave frequencies were reduced and the response to cAMP addition abolished after treatment with caffeine, a known adenylyl cyclase inhibitor in Dictyostelium.
This work demonstrates that TPC2 plays a role in the early stages of Dictyostelium development. TPC2 is important for pH regulation in acidic vesicles and cytosolic Ca2+ levels, either or both of which could influence development either directly or via changes in early developmental gene expression.
The in-vivo study of pain in neuromyelitis optica
Keywords: neuroinflammation
MRI
pain
Devic's
neuromyelitis optica

A thesis exploring the aetiology of pain in neuromyelitis optica (NMO).

Neuromyelitis optica is a severe autoimmune neuroinflammatory disorder characterised by longitudinally extensive myelitis and severe optic neuritis. An oft-neglected symptom of the condition is severe, intractable chronic pain that can be detrimental to quality of life and is marked out by its severity and prevalence in comparison to the related disorder, multiple sclerosis.
The experiments within this thesis make use of both conventional and advanced MRI techniques applied to the spinal cord and brain, and 1H NMR spectroscopy of blood plasma to explore the mechanisms driving chronic pain in NMO. The principle findings are:

i. Thoracic lesions are associated with greater pain, irrespective of lesion length or cervical lesion volume. They are associated with spinothalamic tract damage in the cervical cord that correlates with the severity of pain. A possible autonomic aetiology is proposed.
ii. Periaqueductal grey (PAG) to dorsolateral prefrontal cortex and to pregenual anterior cingulate cortex functional connectivity is correlated with pain severity, conversely PAG to rostroventromedial medulla connectivity is negatively associated with pain severity. Disruption of descending pain modulatory circuits is considered.
iii. PAG glutamate concentration is negatively correlated with pain scores and is higher in low pain patients compared to controls and high pain patients. The discussion includes consideration of pain vulnerability.

Chronic pain in NMO can devastate lives. The studies undertaken in this thesis are some of the earliest MRI imaging studies directed at understanding pain in NMO and the association of pain with thoracic lesions and the aberrations in the descending pain modulatory network are novel findings. There is more to be done and my hope is that this body of work will serve as a stepping-stone to a better understanding of chronic pain in NMO and the future development of effective treatments.
On privacy
Keywords: privacy

This thesis concerns the ethics and political philosophy surrounding privacy. It investigates what privacy is, what is at stake in its loss, and how it relates to other rights and values.
The first part sets the groundwork for the rest of the thesis. Chapter One delves into the origins of privacy. I argue that privacy is not a recent cultural product, but rather a need buried deep in our evolutionary and human history.
The second part of the thesis is dedicated to conceptual issues. Chapter Two clarifies the relation between privacy and the public and private divide. I argue against the popular belief that privacy is an issue that belongs solely to the private sphere. Chapter Three reviews the most influential definitions of privacy that have been offered in the legal and philosophical literature, and points out some of their shortcomings and strengths. In Chapter Four, I develop my own definition of privacy as remaining personally unaccessed, as well as an account of the right to privacy as a right to a robustly demanding good. I also map out the moral significance of privacy perceptions, and privacy-related obligations.
The third part of the dissertation concerns practical issues. Chapter Five inquires into the relationship between security and privacy. I argue that mass surveillance is a disproportionate, unnecessary, and ineffective response to the threat of terrorism. I also argue that encryption should be widely used, as it can curtail the mass surveillance of content and protect people without seriously obstructing criminal investigations. Chapter Six explores the relationship between privacy and transparency. I argue that transparency should sometimes be limited in the interest of privacy. Chapter Seven deals with the questions of whether we can lose privacy to computer algorithms, and whether decision-making algorithms can violate our right to privacy. I answer both questions in the negative, as algorithms are currently neither our peers nor moral agents responsible for their actions.
The conclusion sketches some of the lessons learnt over the course of this investigation.
Investigating resistance mechanisms to ATR inhibitors
Keywords: DDR (DNA damage response)
resistance
ATR
AZD6738

The kinase activity of Ataxia-Telangiectasia and Rad3 related (ATR) protein plays a key role in controlling the DNA damage response (DDR) by acting through downstream targets to stabilise DNA replication forks, induce a transient cell cycle arrest to facilitate the repair of DNA, and thereby prevent premature mitosis in cells with unrepaired or unreplicated DNA. ATR is activated when single stranded DNA arises, for example, from stalled DNA replication forks or during the intermediate steps of double strand DNA break repair, and it is thought to mediate most of its responses through the downstream checkpoint kinase 1, CHK1. As ATR plays an important role in intra-S phase and G2/M cell cycle checkpoints, it has been proposed that inhibition of ATR activity selectively targets tumour cells that have defects in DNA damage-induced G1 cell cycle checkpoints by causing the accumulation of DNA damage and cell death.
AZD6738 is a selective ATR kinase inhibitor that is currently in early clinical development (phase 1). Since development of resistance has been universally observed with long-term use of kinase inhibitors in the clinic, we aimed to identify potential mechanisms of acquired resistance to AZD6738. In this study, we used an in vitro assay system using PC-9 human non-small cell lung cancer cells to generate independent clonal cell lines (SO25, SO45, and SO46) that were resistant to AZD6738. The resistant cells have increased relative cell viability in the presence of AZD6738 compared to the parental cells and, surprisingly, grow better in the presence of the inhibitor than in its absence. AZD6738 resistant cells are also cross-resistant to other chemically unrelated ATR inhibitors (VE-821 and VE-822) as well as CHK1/2 and WEE1 inhibitors, but not ataxia telangiectasia mutated (ATM) inhibitors. Together, cell cycle, DNA fibre, and RNA-sequencing analyses all point towards differential cell cycle regulation in the resistant clones while western blot analysis revealed activation of ATM signalling in the presence of AZD6738 in parental PC-9, but not in the resistant cells.
Western blot and RNA-sequencing analyses demonstrated that cyclins A2 and E1 are downregulated in the resistant cells. Knockdown of cyclin A2 and/or cyclin E1 using siRNA in parental PC-9 cells increased cell viability in the presence of AZD6738. We conclude that downregulation of cyclins A2 and E1 can confer resistance to ATR kinase inhibitors, and may also confer cross-resistance to other inhibitors targeting certain pathways downstream of ATR signalling (CHK1, WEE1).
Technology and political speech: commercialisation, authoritarianism and the supposed death of the Internet's democratic potential
Keywords: survey
social media
discourse analysis
Internet
public sphere
commercialisation
random sample
China
USA

The Internet was initially seen as a metaphor for democracy itself. However, commercialisation, incorporation into existing hierarchies and patterns of daily life and state control and surveillance appear to have undermined these utopian dreams. The vast majority of online activity now takes place in a handful of commercially owned spaces, whose business model rests on the collection and monetisation of user data. However, the upsurge of political action in the Middle East and North Africa in 2010 and 2011, which many argued was facilitated by social media, raised the question of whether these commercial platforms that characterise the contemporary Internet might provide better venues for political speech than previous types of online spaces, particularly in authoritarian states.
This thesis addresses the question of how the commercialisation of online spaces affects their ability to provide a venue for political speech in different political systems through a mixed-methods comparison of the U.S. and China. The findings of this thesis support the hypotheses drawn from existing literature: commercialisation is negative for political speech but it is less negative, even potentially positive, in authoritarian systems. However, this research uncovers a surprising explanation for this finding. The greater positivity of commercialisation for political speech in authoritarian systems seems to occur not despite the government but because of it. The Chinese state’s active stance in monitoring, encouraging and crafting ideas about political speech has resisted its negative repositioning as a commercial product. In contrast, in the U.S., online political speech has been left to the market that sells back the dream of an online public sphere to users as part of its commercial model. There is still hope that the Internet can provide a venue for political speech but power, particularly over the construction of what it means to be a political speaker in modern society, needs to be taken back from the market.
Dissecting heterogeneity in GWAS meta-analysis
Keywords: meta-analysis
heterogeneity
prior distributions
coronary artery disease
genetic association meta-analyses
Bayesian meta-analysis
LPA
GWAS

Statistical heterogeneity refers to differences among results of studies combined in a meta-analysis beyond that expected by chance. On the one hand, excessive heterogeneity can diminish power to discover genetic signals; on the other, moderate heterogeneity can reveal important biological differences among studies. Given its double-edged nature, this thesis dissects heterogeneity in genetic association meta-analyses from three vantage points. First, a novel multi-variant statistic, M is proposed to detect genome-wide (systematic) heterogeneity patterns in genetic association meta-analyses. This was motivated by the limited availability of appropriate methodology to measure the impact of heterogeneity across genetic signals, since traditional metrics (Q, I2 and τ2) measure heterogeneity at individual variants. Second, given that meta-analyses comprising small numbers of studies typically report imprecise summary effect estimates; GWAS-derived empirical heterogeneity priors are used to improve precision in estimation of average genetic effects and heterogeneity in smaller meta-analyses (e.g. ≤ 10 studies). Third, a critical evaluation of the Han-Eskin random-effects model shows how it can identify small effect heterogeneous loci overlooked by traditional fixed and random-effects methods.
This work draws attention to the existence of genome-wide heterogeneity patterns, to reveal systematic differences among the ascertainment criteria of participating studies in a meta-analysis of coronary disease (CAD) risk. Furthermore, simulation studies with the Han-Eskin random-effects model revealed inflated genetic signals at small effect loci when heterogeneity levels were high. However, it did reveal an additional CAD risk variant overlooked by traditional meta-analysis methods.
We therefore recommend a holistic approach to exploring heterogeneity in meta-analyses which assesses heterogeneity of genetic effects both at individual variants with traditional statistics and across multiple genetic signals with the M statistic. Furthermore, it is critically important to review forest plots for small effect loci identified using the Han-Eskin random-effects model amidst moderate-to-high heterogeneity (I2 ≥ 40%).