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‘The Poor Man’s Disease’: Glasgow’s sanitary battle against typhus in the nineteenth century
Get PDFIntroduction: pages 5-7
Dissecting genetic variation and the application of infrared spectroscopy for predicting health outcomes in Drosophila
No full textAbstract not currently available
Investigating the role of intestinal IFN-γ expression on barrier immunity during enteric helminth infection
Get PDFEnteric helminth infections are a significant global health concern, with over a quarter of the world’s population infected. The infecting parasites often migrate through host tissues and, as large multicellular organisms, they can cause substantial damage. The nematode Heligmosomoides polygyrus bakeri (H. polygyrus) causes a chronic roundworm infection in mice and during early stages of its lifecycle, it enters the intestinal tissue. The main aim of this thesis is to understand the factors that influence the immune responses at this early breach timepoint of infection, with particular interest in the role of IFN-γ production local to the helminth.
Our data show that the early phase of infection is accompanied by barrier disruption and an acute IFN-γ response which precedes the classical Th2 response to helminth infections. Through in vivo neutralisation of IFN-γ, we were able to show that the early production of IFN-γ contributes to increased expression of cell adhesion molecules found in H. polygyrus infection, suggesting a reparative role; and also coordinates and enhances local antimicrobial immunity characterised by increased antimicrobial protein expression and influx of innate myeloid cells to the site of infection.
Enteric helminth infections occur in the context of intestinal microbiota, and in environments where co-infection is frequent. We hypothesised that the antimicrobial responses and epithelial repair responses orchestrated by local IFN-γ could be in response to bacteria potentially invading the ‘breach’ points created by H. polygyrus. Using broad-spectrum antibiotics to eliminate microbiota during H. polygyrus infection, we found that the early IFN γ response is dependent on bacterial presence. Our data suggest that bacterial translocation may not occur, perhaps reflecting the effective antimicrobial and tissue repair responses during the helminth infection. Together our data contribute to our overall understanding of the intestinal environment and immune responses in early H. polygyrus infection. By illustrating a microbiota-driven production of IFN-γ and characterising a role for this cytokine in coordinating local responses, we reveal a potential role for IFN-γ in protecting the host from secondary infection
Micro-optics: From wormholes to medical applications
Get PDFLying within the overarching boundaries of micro-optics, this thesis begins by exploring two different “pixelated” approaches to medical spectacles. The first was aimed as a treatment alternative to surgical corrections for torsional diplopia (double vision due to a relative eyeball rotation around the back-to-front axis). To do this, the spectacles were designed to approximately rotate the view seen through them. In the very basic design consisting of simple wedges (each of which we call a pixel), the component does not actually perform imaging leading to a reduced visual acuity of about 6/420. This was worse than desired, which is why the latter half of the chapter was designated to implementing methods that may improve the visual acuity, consisting of a pupil restriction and a new approach we call derivative control. Using this, the visual acuity was improved to a value of about 6/35, but at the cost of added complexity and bulkiness. The second spectacle pair was designed to help with uncorrected refractive errors by permitting patients to adjust the focusing power of the spectacles through simply rotating two cylindrical lens spirals relative to one another. The combination yields an equivalent Fresnel lens, which is the reason we call these adaptive Fresnel lenses. These were simulated in the ray optical limit, yielding an expected feasible variable range of ±2 diopters. Several configurations were explored where the distances and spiral types were varied, with ray trace simulations confirming the expected view through the adaptive lens.
The other half of the text is dedicated to an extension of the previously published ideal lens cloak. The cloaking principles were explored, yielding conditions for which a light ray within the cloak must remain within the cloak and hence travel in a closed ray trajectory. These were used in the theoretical creation of a novel “extreme omnidirectional ideal lens cloak” which hides an object within from all viewing directions. Furthermore, attractor-like properties were found within the cloak, trapping some light rays entering from the outside. These properties were also found in a more realizable cloak we call the “shifty cloak”, which suggests significant similarities between the two cloaks. The shifty cloak was used to construct a Janus device and an optical wormhole, the latter of which was adversely affected by the attractor properties
Experimental and theoretical studies of Surface Acoustic Wave enhanced Localised Surface Plasmon Resonance sensors
No full textIntegrating different sensing platforms offers several advantages, such as enhanced sensitivity and advanced sensing mechanisms. Surface acoustic wave (SAW) devices have been used for microparticle manipulation for a long time, offering precise and controlled manipulation in a contactless manner. Similarly, localised surface plasmon resonance (LSPR) sensors also offer a label-free sensing platform without engagement of any human contact. LSPR is an extremely sensitive sensing mechanism which can sense a small change in the NPs' dimensions and the surrounding medium. On the other hand, SAW devices offer several approaches for particle manipulation such as mixing, separation and patterning, so SAW can be used for sample preparation, transport and deliver it to the desired location. Integration of the SAW with LSPR offers to development of a complete sensing setup where SAW can be used to manipulate plasmonic particles and LSPR can be utilised for their ultra-sensing capabilities.
This thesis presents a novel approach for integrating SAW devices with LSPR sensing, based on the manipulation of a plasmonic monolayer. For the integration of the SAW with LSPR sensing, a stable and detectable plasmonic particle structure is necessary. A two-dimensional layer of plasmonic nanoparticles offers a key benefit in such configurations where the dimensions of self-assembled particles are comparable to the SAW wavelength, and yet the plasmonic properties remain preserved, essential for an LSPR sensor. Before the integration, we confirmed that the effects of SAW propagation, which generate spontaneous surface deformation, do not alter the plasmonic properties that can affect the resonance spectrum, implying that the LSPR properties remain unchanged.
Theoretically and experimentally, we confirmed that the shear vertical-SAW (SV-SAW) cannot affect the LSPR spectrum of gold nanoparticles. Eventually, we prepared the gold nanoparticle monolayer and used it with the SAW device for the alignment and patterning. The most significant advantage of our novel work is that the pattern remains intact when SAW is off, and even when the microchannels are removed. The experimental SAW setup allowed refractive index (RI) sensing by integrating it with the LSPR setup. To the best of our knowledge, this was the first demonstration that SV-SAW propagation cannot affect the plasmonic structure over the surface of the SAW device and is able to manipulate micro structured metallic monolayers in a contactless manner
Rational verification for Stackelberg Security Games
No full textStackelberg Security Games (SSGs) are a game theory model used to help defenders analyse threats, plan strategies, and estimate resource allocation in cybersecurity. In an SSG, a defender (the leader) commits to a defensive strategy, which an attacker (the follower) observes before choosing their optimal response. However, a key limitation of this model is the absence of a formal approach to verify that the security properties of a defensive strategy hold under Stackelberg equilibrium conditions, particularly against adaptive adversaries. Rational verification addresses this by checking if a temporal logic formula is satisfied in some or all computed equilibria. This thesis introduces a framework that integrates rational verification into SSGs, and defenders can design rational defensive strategies.
We propose a methodology for analysing adaptive adversaries by formalising SSGs, a model that uses rational verification to incorporate equilibria and payoffs to synthesise dynamic strategies. We implement StEVe (Stackelberg Security Games and Equilibrium Verification), a tool that integrates game theory and formal verification for rationality in cybersecurity. The key features of StEVe include Attack Defence Trees (ADTs) to model threats, formalising security properties using temporal logic rPATL (Probabilistic Alternating-time Temporal Logic with Rewards), and an extension of PRISM-games to compute Stackelberg equilibria. StEVe gathers data from public vulnerability databases, such as Common Vulnerabilities and Exposures (CVEs), allowing defenders to tailor strategies to specific characteristics of cyber threat.
Our results show that the adaptive strategies produced by our framework yield a reduction in defender costs and system compromise compared to static approaches. The primary contribution of this work is a methodology and supporting tool that aim to improve the robustness of defensive strategies against adaptive threats by applying rational verification
Elucidating metabolic alterations in myeloproliferative neoplasms: implications for treatment
Get PDFMyeloproliferative neoplasms (MPN) are haematopoietic disorders leading to aberrant expansion of myeloid, erythroid and megakaryocytic lineages, which are classified into three phenotypes namely Polycythaemia Vera (PV), Essential Thrombocythemia (ET) and Primary Myelofibrosis (PMF). The three main somatic mutations leading to MPN are Janus Kinase 2 V617F (JAK2V617F), Janus Kinase 2 exon 12 and Calreticulin (CALR). JAK2V617F was the first driver mutation found in MPNs, with a mutation rate of 95%/60%/55% in PV, ET and PMF respectively. Janus Kinase 2 is one of the four Janus proteins that are constructed to transmit signals from cytokines to downstream pathways. Inside normal cells, Janus Kinase 2 has a dual kinase structure with two domains: an active tyrosine kinase domain JH1 and a catalytically inactive pseudokinase domain JH2. Both domains interact with each other, depending on cellular context with JH2 inhibiting JH1 in the absence of cytokines. In MPN, these mutations lead to the inhibition of JH1 and constitutive activation of downstream signalling pathways including STAT5, RAS/MAPK and PI3K/Akt/mTOR pathways in the absence of growth factors. The discovery of JAK2 inhibitors is beneficial for improving the disease phenotype, but do not eliminate the persistent blood cancer stem cells, which are the cause of the disease. It is therefore important, to find novel therapeutic ways to target blood cancer stem cells. To sustain accelerated growth rate, cancer cells need abnormally high amounts of energy, taken up as nutrients (e.g. glucose), and to increase the breakdown of such nutrients through metabolic pathways. In addition, cancer cells often alter the amount of oxygen they require for the purpose of growing rapidly. Cancer cells therefore alter normal cellular metabolic pathways. Importantly, relying on these alterations makes them vulnerable and targeting altered metabolism and oxygen consumption is an attractive approach to eliminate cancer therapeutically.
My data indicate that metabolic processes are deregulated in MPN. In this thesis, experiments have been performed using advanced assays including Seahorse Metabolic Flux Assay, Liquid Chromatography-Mass Spectrometry, and Flow-cytometry. C57BI/6 background JAK2V617F mice bone marrow cells were used. We have found the increased oxidative phosphorylation and glycolysis rates in MPN Homozygous and heterozygous mice c-kit+ cells. Furthermore, three solute carriers—SLC25A37, SLC2A4, and SLC43A1—were highly expressed in HSCs from homozygous mice, suggesting that targeting glutamine as an anaplerotic pathway could be a potential therapeutic approach
Can serious adverse event rates be used as a metric of trial representativeness for pharmacological interventions? Glucagon-like peptide-1 receptor agonist trials as an exemplar
Get PDFBackground: Randomised controlled trials (RCTs) are considered the gold standard for assessing the efficacy and safety of interventions because they introduce the concept of randomisation, which effectively deals with unmeasured confounders. Trial representativeness is crucial for determining the generalisability of trial findings to the target population. However, various groups, such as older people and those with multiple long-term conditions, are often under-represented in trials. Moreover, there is no gold standard measure of trial representativeness. Current measures are complex, subjective and time-consuming, suggesting the need for new and better measures of trial representativeness. Serious adverse events (SAEs) are likely to be reported in trials and routine care. Moreover, they are reasonably objective, tangible and predictable trial outcomes. Consequently, the SAE rate has been proposed as a potential metric of trial representativeness. However, little is known about its feasibility and validity in measuring trial representativeness. This thesis investigates whether the SAE rate can be used to measure trial representativeness, using glucagon-like peptide-1 receptor agonists (GLP-1 RA) trials as an exemplar.
Methodology: This thesis builds on an ongoing systematic review of novel antidiabetics. GLP-1 RA trials were used as an exemplar for the analysis. Data sources used in this thesis were ClinicalTrials.gov, clinical study reports (CSRs), study protocols, and journal publications. I conducted several approaches to examine whether the SAE rate can be used as a proxy for trial representativeness. First, I examined SAE reporting in RCTs in the published literature to explore the feasibility of using the SAE rate as a metric of trial representativeness. Second, I explored SAE capturing in GLP-1 RA trials to enable the calculation of SAE rates. Third, I compared SAE rates between intervention and control arms to determine if combining SAE rates of trial arms is feasible to increase statistical precision. Fourth, I assessed GLP-1 RA trials using the PRECIS-2 (PRagmatic Explanatory Continuum Indicator Summary) tool to enable fair comparison with the SAE rate and explore the challenges of using this tool. I protocolised and operationalised the PRECIS-2 tool to score GLP-1 RA trials as objectively as possible. Fifth, I compared the SAE rate with the PRECIS-2 tool based on the differences in their associations with several markers of trial representativeness that serve as fair umpires to examine the validity of the SAE rate as a metric of trial representativeness. Finally, I examined the association between eligibility criteria and the SAE rate to examine whether eligibility criteria, a possible driver of trial representativeness, is associated with the SAE rate.
Results: Firstly, I found that SAEs were reported for most GLP-1 RA trials. However, SAE timeframes were not explicitly reported for nearly half of the trials. Major adverse cardiovascular event (MACE) trials had inconsistent reporting of MACE counts in SAE total counts. Secondly, I found no difference in SAE rates between intervention and control arms. Furthermore, the retrospective assessment of GLP-1 RA trials using the PRECIS-2 tool was challenging and time-consuming. The missingness of information required to score recruitment and organisation domains was high. I found no correlations between the domains of the PRECIS-2 score, except for modest correlations between eligibility criteria and recruitment domains and between setting and primary outcome domains. Moreover, I found no association between the SAE rate and the PRECIS-2 score. Additionally, I found that all fair umpires did not strongly favour the SAE rate over the PRECIS-2 tool. However, the direction of the difference in associations for half of the umpires favoured the SAE rate. Finally, I found that trials with permissive eligibility criteria were positively associated with higher SAE rates. Trials with increased continuous eligibility criteria scores were also positively associated with increased SAE rates.
Conclusion: Most GLP-1 RA trials reported sufficient SAE data indicating the ability to calculate SAE rates. SAE rates were similar across trial arms, suggesting the feasibility of combining SAE rates from the intervention and control arms. The SAE rate was not associated with the PRECIS-2 score, and none of the fair umpires strongly favoured the SAE rate. However, the directions of half of the fair umpires favoured the SAE rate and trials with permissive eligibility criteria were associated with increased SAE rates. Therefore, the SAE rate may be useful as a quick-to-measure proxy of the restrictiveness of eligibility criteria. However, given that the triangulation of evidence is inconsistent in supporting the use of the SAE rate as a standalone metric of trial representativeness, examination of the SAE rate would need to be considered carefully in combination with other metrics before making overall judgements about trial representativeness
Synthesis of unnatural fluorescent α-amino acids
Get PDFUnnatural α-amino acids are important probes that can be incorporated into proteins and peptides for biological study using fluorescence spectroscopy. The objective of this MSc research programme was to investigate the synthesis of novel fluorescent unnatural amino acid using transition metal catalysis to implement the key steps.
The aim of the first project was to construct a phenoxathiin α-amino acid and to extend its conjugation by halogenation and subsequent Suzuki-Miyaura reactions. Initially, the phenoxathiin α-amino acid core was prepared using a dual catalytic thioarylation reaction using iron triflimide as a Lewis acid and diphenyl selenide as a Lewis base, followed by a copper mediated cyclisation. Various approaches using succinimide and saccharin reagents were investigated for halogenation but due to either poor conversions or difficult separations, an effective halogenation was not possible. However, the reaction with N-iodosaccharin did give access to a novel phenoxathiinsulfoxide α-amino acid which displayed fluorescent properties with an emission maximum at 372 nm.
The aim of the second project was to develop a synthesis of a nitro-substituted dibenzofuran α-amino acid. The strategy involved the synthesis of a biaryl analogue of tyrosine, followed by a metal-catalysed C–H activation and cyclisation reaction. Initially, the cyclisation precursor was prepared in three steps. This involved aminogroup protection, ortho-bromination and then Suzuki-Miyaura cross-coupling reaction with 4-nitrophenyl boronic acid. The first attempted cyclisation used a palladium catalyst and a per-ester as the oxidant was successful but gave the product in only 6% yield. Other methods were investigated in an effort to improve the efficiency of this transformation. Using a simpler method involving a copper(II)-mediated single electron transfer cyclisation was more efficient and gave the desired nitro-substituted dibenzofuran α-amino acid in 16% yield
Investigating the role of BCL-2 family members in chemoresistance in adult and paediatric core-binding factor acute myeloid leukaemia
Get PDFPaediatric and adult acute myeloid leukaemia (AML) are distinct diseases with different molecular and genetic signatures. Core-binding factor (CBF) AML is a subgroup of AML with chromosomal translocations [t(8;21)] or inversions [inv(16)]. Patients with CBF AML are considered to have favourable prognosis, despite this, relapse and chemoresistance is common in this subgroup.
Changes in apoptotic pathways have been demonstrated to contribute to resistance to established chemotherapies, such as cytarabine (Ara-C), in AML. BH3 mimetics have shown promise for the treatment of AML, however the use of BH3 mimetics in patients with CBF AML that are resistant to Ara-C remains unclear. Therefore, the broad aim of this project was to determine the role of apoptotic proteins in resistance to Ara-C using t(8;21) and inv(16) CBF AML cell lines. This project also sought to assess the use of BH3 profiling as a biomarker of response to BH3 mimetics in CBF AML.
To achieve this, Ara-C resistant models were generated using escalating doses of Ara-C. Baseline BH3 profiling was used to determine how acquisition of Ara-C resistance affected apoptotic priming of CBF AML cell lines. The dependency of these cell lines on anti-apoptotic proteins was assessed using intracellular staining and measurement of response to BH3 mimetics. Dynamic BH3 profiling was performed and compared to response to BH3 mimetics by apoptosis assay.
This project demonstrated that Ara-C resistance has different impacts on the apoptotic priming of paediatric t(8;21) Kasumi-1 and adult inv(16) ME-1 AML cell lines. In both Ara-C resistant cell lines, dynamic BH3 profiling using cytochrome c release recapitulated the apoptotic response typically seen with BCL-2 inhibition. Particularly noteworthy was the up regulation of MCL-1 expression in the resistant lines, suggesting increased reliance on this anti-apoptotic protein for survival. However, treatment with the selective MCL-1 inhibitor, S63845,failed to enhance cytochrome c release or induce apoptosis. In contrast, venetoclax, a BCL-2 inhibitor, induced greater cytotoxicity in Ara-C–resistant ME-1, but not in Kasumi-1.
Results of this project highlight the complexity of BCL-2 family member interactions and highlight that it may not be possible to infer functional apoptotic dependencies from the expression levels of anti-apoptotic proteins. These results suggest that there is a potential for pre-treatment with Ara-C to increase response to venetoclax. However, further work is needed to determine the most effective sequence of treatment with Ara-C and BH3 mimetics. This project adds to previous evidence which suggests that the use of dynamic BH3 profiling for prediction of response to BH3 mimetics may be helpful
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