VictorJuiz/thesis huggingface dataset:
Characterising the Notch-ligand binding interaction, and its modulation by glycosylation

The Notch signalling pathway is universally conserved in all metazoan species, and is involved in many aspects of cell fate determination and tissue homeostasis, during development and in adult organisms. Several developmental diseases are associated with defective Notch signalling, and the Notch pathway has been implicated in a growing number of cancers.
The Notch signalling pathway requires direct cell-cell contact for ligand binding and receptor activation to occur. Specific domains within the Notch receptors and ligands have been identified as necessary for the interaction to take place, and a series of enzymes are known to regulate Notch signalling via glycosylation. Other domains beyond the minimal ligand binding region of the Notch receptor are also known to influence binding.
The aim of this study was to characterise the molecular basis for ligand binding by the Notch receptor, and how this is regulated by glycosylation. The effects on ligand binding of specific amino acid substitutions and sugar modifications were tested using prokaryotically -expressed proteins, and a series of constructs containing additional domains N-terminal of the ligand binding region was produced prokaryotically and eukaryotically to test how additional domains might affect ligand binding.
Binding was assessed by a flow cytometry-based binding assay and by SPR in order to investigate how particular modifications affected ligand binding. These assays indicated that an evolutionarily-conserved hydrophobic site exists within the central β-sheet of EGF12 in the Notch receptor that is directly adjacent to the O-fucosylation site within this domain. The GlcNAc-fucose disaccharide modification at this position was found to increase binding of hNotch1 to both Jagged1 and DLL4. Additional EGF domains N-terminal to the ligand binding region showed opposite effects on binding to these two ligand classes, suggesting that the precise mode of binding may vary slightly between different Notch ligands.

Keywords: glycosylation
Notch
Jagged
ligand-binding
Delta
Advisor: Handford, P
Optogenetic analysis of inhibitory circuits in the neocortex

Information processing in the brain involves circuits composed of different classes of neurons. Inhibitory interneurons are by far the most diverse subgroup in terms of physiology and morphology, and the principles that govern their connectivity are far from understood. Their excitatory counterparts, pyramidal and spiny stellate cells, have been found to be governed by a compartmentalized horizontal and vertical structure – forming layers and columns – as well as by a stereotypic or “canonical” pattern of connections between them.
However, it has remained unclear whether similar, general organizing principles exist for inhibitory circuits. To map the sources of inhibitory inputs to neocortical pyramidal cells, a Cre-lox-based mouse knock-in line, that conditionally expresses the light-sensitive ion channel channelrhodopsin-2 in GABAergic neurons, was generated. Expression levels were sufficient to drive interneuron spiking at up to 40 Hz, but too low to activate synaptic terminals allowing perisomatic activation and thus mapping of local connections. I found, that inhibitory inputs to excitatory cells in all layers in primary motor (M1), somatosensory (S1), and visual cortex (V1) derive largely from the same layer and putative column. However, four translaminar inhibitory connections were found, which differ significantly both, between as well as within a cortical area. Most notably, (some) excitatory cells in layers 2/3, 4 and 5B of V1 receive prominent, putative feedback inhibition from their direct or indirect output layers. Furthermore, while a columnar structure is visible also in inhibitory circuits, their laminar organization is degenerate in an area-specific manner. Neocortical inhibitory microcircuits thus display significant variations, which potentially reflect the localization of function.

Keywords: optogenetics
neocortex
neuroscience
physiology
inhibition
Advisor: Miesenböck, G
Sharing findings on sickle cell disorder in international collaborative biomedical research: an empirical ethics study in coastal Kenya

Against the background of a dilemma experienced by researchers during a genomics study at an established biomedical research centre in Kenya, the broad aims of this thesis are to develop appropriate responses to important ethical questions on sharing information on a common and serious genetic condition, sickle cell disorder, and assess the responsibilities of researchers in this regard. Using an empirical approach to normative reflection across two phases of qualitative research, I explore the nature of important moral concerns related to sharing sickle cell disease information from researchers’ and community members’ points of view; and develop a bottom-up normative analysis around the questions generated. This analysis interweaves community experiences, processes of community reasoning and ex situ normative reflection; placing community views and values centrally while referencing these to wider ethical debates, commentaries and guidelines in the literature. Two main outputs of this thesis are to provide recommendations for information sharing on SCD findings in the genomics study in Kilifi; and to propose a set of key issues to consider for this type of information in other studies and geographic settings. I conclude that researchers have a strong responsibility to share SCD information on affected children with families as a form of ancillary service (validating tests, counselling and care); but less responsibility to actively share carrier information. Concurrent responsibilities are working collaboratively with the Ministry of Health/District General Hospital to plan and implement services for SCD; ensuring counselling services support family stability as far as reasonably possible; and to build forms of community engagement and informed consent that counter risks of diagnostic interpretations of research.

Keywords: family counselling
international research ethics
disclosure genetic findings
empirical ethics
Africa
Kenya
sickle cell disease
Advisor: Parker, M
Molyneux, C
Fitzpatrick, R
Foam as a soil conditioner in tunnelling: physical and mechanical properties of conditioned sands

Earth pressure balance (EPB) tunnelling machines are commonly used for the construction of tunnels in soft soils. These machines use the excavated soil in a pressurised head chamber to apply a support pressure to the tunnel face during excavation. How well an unstable face is supported in an EPB machine depends on effectively transferring a constant pressure from the support medium to the surface of the face. If the support pressure is not constant, but instead changes, the varying pressure inevitably leads to collapse of the face or heave on the surface ground. A machine may be designed to work in “Ideal ground” conditions. However, natural soils rarely have these properties, and conditioning of the soil is usually necessary to change its properties to suit the machine. Effective soil conditioning significantly improves the machine performance and control of the soil flow through the screw conveyor. However, for soil conditioning as commonly used in practice, the effects of different conditioning treatments on soil properties and the machine performance are not clearly understood, and problems with EPB machine operations related to the soil properties are often encountered. This thesis presents experimental investigations of direct shear box tests on conditioned sands, compressibility tests on conditioned sands and a model EPB screw conveyor operating with sandy soils. Index tests were performed to investigate effects of foam and polymer conditioning treatments on the plastic fluidity of different Leighton Buzzard (L.B.) sands and Thanet sand. The index tests allowed assessment of conditioning treatments for sandy soils, and optimum ranges of treatments for Leighton Buzzard sand and Thanet sand are suggested. In the series of shear box tests, performed on conditioned Thanet sand and conditioned L.B. sands, one of the most important findings was that the pore water pressure controls the strength of the sand foam mixtures. An increase in foam injection ratio (FIR) might produce an increase of pore water pressure and a decrease of shear stress. In the series of compressibility tests, performed on conditioned Thanet sand and conditioned L.B. fine sand, one of the most important findings was that increasing FIR does not increase maximum gas expelled. The FIR and the initial relative density of the specimen have to be related to the capability of the specimen to retain the gas and establish a coexistence between grains of soil, water and gas bubbles. Finally, from the series of model (1:10 scale), screw conveyor tests, performed on conditioned Thanet sand, conditioned L.B. fine sand and conditioned Garside sand, it can be concluded that the Oxford screw conveyor model can be used as a trial machine to study the effects of different operating conditions on conditioned sand specimens similar to those used on site. This can allow choice of conditioning methods to improve the performance of an EPB machine at a given site.

Keywords: sand
tunnelling
Advisor: Houlsby, G
A critical study of the Liber historiae Francorum

Although the Liber Historiae Francorum is the only surviving contemporary chronicle which treats the fifty years spanning the turn of the seventh to the eighth century in Frankish Gaul, it has been generally mistrusted by students of the period. This thesis is an attempt to discover whether the reputation is just or whether the chronicle might yield new information about the later Merovingian age. In so doing it asks three questions: 1) Is the work's latest edition, which is now nearly 100 years old, still the best version of the text we can achieve? 2) How accurate is the LHF's description of events? 3) Does a study of the author and his attitudes yield any insight into the nature of late Merovingian politics and society? The study concludes that the edition is an accurate rendering of the surviving manuscripts and an adequate basis for study of the work. This is true despite the fact that the contents of one early manuscript, which was unknown to the work's editor, calls into question the currently held assumptions concerning order, date, and composition of the LHF's recensions. Although the chronicle does indeed contain many errors in its earlier parts, for the period from the 650's (LHF-43) to the end of the work, there are a surprising number of instances where its version of events is either as believable as or more probable than the currently accepted view. An analysis of the author and his attitudes outlines a picture of late Merovingian politics as conducted by factions of leading Neustrians, whom the chronicler calls the Franci, and who could find unity in the loyalty due the legitimate Merovingian king.
The operation of sexual selection in the red junglefowl

Sexual selection acts on traits that increase the reproductive success of an individual in competition with other individuals of the same sex over reproductive opportunities, through intra-sexual competition and inter-sexual mate choice. Because males benefit more from remating than females, they are subject to more intense sexual selection. Modern genetic tools have shown that females often mate promiscuously, thus prolonging sexual selection after insemination through intra-sexual (sperm competition) and inter-sexual (cryptic female choice) episodes. Despite increasing interest in post-copulatory sexual selection, the implications of sperm competition, cryptic female choice and underpinning mechanisms remain little understood. This thesis adopts an integrated approach to quantify the relative importance of post-copulatory episodes in the operation of sexual selection, elucidate their proximate mechanisms in the red junglefowl Gallus gallus. By combining behavioural observations of replicate groups with paternity data, I show that female promiscuity decreased the total opportunity for sexual selection in a group, but accounts for an unexpectedly large proportion of the variance in male reproductive success. By comparing the operation of sexual selection on multiple male traits, I show that post-copulatory sexual selection reinforced pre-copulatory sexual selection for male social dominance and that female preferred to mate with compatible males.
I used experiments to study the mechanisms of post-copulatory sexual selection by studying the effect of seminal fluid in sperm competition and cryptic female choice in relation to male status and relatedness. Following previous work indicating that seminal fluid products influence sperm quality in this species, I tested in vivo whether the seminal fluid of an ejaculate acts differentially towards sperm from the same ejaculate and rival sperm, and found no evidence for this idea. Finally, I show that cryptic female choice can drastically bias the outcome of sperm competition, and that female fowl might bias paternity toward unrelated males.

Keywords: sexual selection
red jungle fowl
MHC
post-insemination sexual selection
intra- and inter-sexual selection
gallus gallus
cryptic female choice
Advisor: Pizzari, T
Pedunculopontine nucleus stimulation for gait and postural disorders in Parkinson's disease

The pedunculopontine nucleus (PPN) is a reticular collection of neurons at the junction of midbrain and pons. The PPN in animal models appears topographically organised and functionally related to locomotion and arousal. In Parkinson’s disease, the PPN degenerates and is susceptible to abnormal basal ganglia output. In patients with Parkinson’s disease, low frequency PPN stimulation is proposed to improve gait freezing and postural instability. However, the therapeutic mechanisms, optimal clinical application and precise effects on gait and posture of PPN stimulation are unclear.
Here, a topographic arrangement of the PPN was supported by local field potential recordings in parkinsonian patients. In the PPN region, beta oscillations were recorded rostrally and alpha oscillations caudally. Alpha oscillations, consistent with their putative role in allocating attention, correlated with gait performance and attenuated with gait freezing. Thus the caudal PPN subregion may be the most relevant target for gait disorders.
Accordingly, an unblinded clinical study suggested that stimulation of the caudal PPN subregion was beneficial for gait freezing, postural instability and falls. In a double-blinded study using spatiotemporal gait analysis, caudal PPN stimulation reduced triggered gait freezing, with bilateral stimulation more effective than unilateral. However, akinesia including akinetic gait did not improve with PPN stimulation. Accordingly, dopaminergic medication requirements did not change.
Mechanisms underlying gait freezing and PPN stimulation were explored with reaction time experiments. Parkinsonian patients with severe gait freezing and postural instability demonstrated a ‘block’ to pre-programmed movement. This was evidenced by prolonged simple reaction times and the absence of ‘StartReact’, whereby pre-prepared responses are normally accelerated by loud acoustic stimuli. PPN stimulation improved simple reaction time and restored Startreact. The relief of this ‘motor block’ with PPN stimulation may therefore explain the associated improvement in gait freezing and postural instability, as these tend to occur in circumstances requiring triggered, pre-prepared adjustments.

Keywords: Parkinson's disease
deep brain stimulation
gait freezing
pedunculopontine nucleus
postural instability
Advisor: Aziz, T
Arthur Johnston and the fostering of Scottish letters

Traditionally, Arthur Johnston has been judged proxime accessit to George Buchanan in the world of Scottish neo-Latin poetry, and particularly in the versification of the Book of Psalms. The thesis offers a counterpoint to that theme. More of his poetry came under scrutiny at the close of the nineteenth century, when an edition of his Parerga and Epigrammata of 1632, turned scholarly attention to his secular poems. This study examines the poems written between 1599 and 1622 during Johnston’s peregrenatio academica in Europe – poems which depict him at the moment of his emergence onto the public stage, and which offer insights into his life, and the worlds he occupied, during those years. Part one of the thesis will examine his early years and his move into the academic world in Aberdeen and at Heidelberg University. Part two will consider the years he passed as a teacher of philosophy at the Huguenot Academy in Sedan, the independent principality on the northern border of France. It will look, too, at the evidence of his year spent in Padua, where he studied to become a physician. Part three will focus on the years 1619-22 when his longest secular poems were composed. He wrote and published with an eye to achieving a post in the medical circle around James VI and I. The thesis concludes by considering the retreat he made from Europe and London to his home in Aberdeen, and looks briefly at one of the small poems he wrote in 1623-24. Throughout, themes emerge of Johnston’s irenic preferences, and his response to the disturbance to intellectual life brought about by Calvinist division, and by the crisis heralded by the Bohemian Revolt.

Keywords: theology
anatomy
George Buchanan
Sedan
Scotland
Daniel Tilenus
Respublica litterarum
Bohemian Crisis
Padua
philosophy
James VI/I
neo-Latin poetry
Heidelberg
irenicism
Advisor: Brigden, S
Norbrook, D
A commentary on Catullus 64, lines 1-201

The thesis consists of detailed commentary on the first 201 lines of Catullus 64, together with an edited text and apparatus criticus. It represents about half a planned commentary on the whole poem, which will also include an introduction. The commentary begins by discussing the poem’s Argonautic opening, its use of allusion to negotiate generic relationships with epic and tragedy, and its exploration of narrative, pictorial and first-person ‘lyric’ modes. As the narrative jumps to Peleus’ wedding, the commentary examines the complicated moral signals about Roman luxury and the golden age sent by the description of the gleaming palace surrounded by abandoned fields. The transition to the description of Ariadne prompts an examination of how this ‘disobedient’ ecphrasis emphasises details that a picture could not literally convey, together with an analysis of the male narrator’s objectifying presentation of a woman in distress. The ecphrasis proper is then disrupted by a ‘flashback’ covering Ariadne’s first encounter with Theseus and his fight with the Minotaur: the commentary explores the ways in which shifting focalisation complicates the reader’s judgement of Theseus’ heroism. Finally, the thesis looks at Ariadne’s speech as an intertextual node, investigating the meanings generated by its relationships with other speeches from both earlier and later in Greek and Roman poetic traditions, and examining how each theme or topos is used in this particular situation both by the alluding poet and by Ariadne herself. More discursive notes introducing the various sections are interspersed with shorter lemmata considering textual, metrical, linguistic and cultural-historical issues as well as literary interpretation. The commentary aims both to open up the possibilities of meaning offered by individual words and phrases, and to advance critical understanding of key aspects of the whole poem, such as its narratorial voice, engagement with visuality and place in literary history.

Keywords: commentary
ecphrasis
Ariadne
Catullus
epyllion
Advisor: Hardie, P
Heyworth, S
The role of ASPP2 in intestinal cell polarity and homeostasis

The intestinal epithelium is an important model that integrates various biological processes, including cell proliferation, cell differentiation and inflammation. Recent evidence has linked intestinal homeostasis with cell polarity. ASPP2, a known tumor suppressor, has been implicated in the maintenance of epithelial integrity. In this thesis, how ASPP2 is involved in intestinal epithelial cell polarity, and whether its function modulates the epithelial homeostasis in the intestine is examined.
For this purpose, the intestinal epithelia of transgenic Balb/c ASPP2 ∆exon3 and ASPP2 conditional knockout mice were studied. The expression of ASPP2 under energetic stress was investigated, and the consequences of the loss of ASPP2 in a DSS-challenged mouse model were evaluated. ASPP2 was found to be localized at the tight junctions in the intestinal epithelium. The loss of ASPP2 in the intestine resulted in epithelial disorganization, and compromised epithelial cell turnover by promoting cell proliferation and influencing Notch-regulated differentiation. Interaction between ASPP2 and the LKB1 complex was also observed, and the expression and localization of ASPP2 was proven to be influenced by glucose levels in an AMPK-dependent manner. Various signs of intestinal inflammation were detected in ASPP2-null mice, and the increased susceptibility of these mice to develop DSS-induced colitis further supported the observations.
Overall, ASPP2 was identified as a gatekeeper of the intestinal tight junctions, and new insights into the complex involvement of ASPP2 in cell metabolism and intestinal inflammation were uncovered.

Keywords: ASPP2
tight junctions
inflammation
intestine
Advisor: Lu, X
Development of tissue and imaging biomarkers of rotator cuff tendinopathy

Rotator cuff tendinopathy accounts for the majority of cases of shoulder pain, however, the aetiology and pathophysiology of the condition is poorly understood and treatment interventions for the condition have been introduced on an empirical basis, guided only by unproven theories of biological and structural changes in and around the affected tissue.
This thesis explores changes in the provision of surgical interventions for rotator cuff tendinopathy, showing a rapid, sustained increase over the last decade. Investigation into the use of ultrasound as a clinical and research tool led to the development of an independent learning method for surgeons using the modality for shoulder imaging and highlighted the technological limitations of contrast enhanced ultrasound in assessing the microvascularity of the supraspinatus tendon.
Development of a novel biopsy method for sampling the supraspinatus tendon permitted collection of tissue samples from several cohorts of individuals with rotator cuff tendinopathy. These studies, for the first time, described tissue changes across the whole spectrum of pathology suggesting the possibility of an early inflammatory phase of the condition followed by tissue senescence and reduced viability as pathology progressed. Paired samples taken before and after treatment identified reduced tissue activity in response to glucocorticoid therapy and a potential healing response from the supraspinatus tendon following surgical repair. Significant deterioration in tissue activity and viability was shown as age increased in a large cohort of individuals, highlighting the major impact of ageing as a factor in the onset and progression of rotator cuff tendinopathy.
The techniques introduced provide standardised, reproducible methods for further evaluation of rotator cuff tendinopathy and the development of novel therapeutic interventions.

Keywords: rotator cuff
ultrasound
supraspinatus
orthopaedics
tendinopathy
shoulder
Advisor: Carr, A
Beard, D