A key function of the gastrointestinal tract is to break down ingested food and absorb the contained nutrients and water. Within the intestinal tract are a number of specialised epithelial cells that aid in this process as well as maintaining immune homeostasis through their functioning as a physical barrier, and secretion of antimicrobial peptides. The intestinal tract has one of the highest turnover rates within organisms, so because of this, and the variety of functions these cells can carry out, it is essential that the balance of proliferation and cell death is regulated to maintain homeostasis. There are several conserved signalling pathways that are responsible for the proliferation of intestinal cells. Suppressor of cytokine signalling 3 (SOCS3) is produced upon activation of the Janus kinase/Signal transducer and activator of transcription (Jak/Stat) pathway and this is an inducer of negative feedback inhibition and is implicated in regulation of intestinal homeostasis, with SOCS3 dysregulation reported in intestinal pathologies, such as cancer and inflammatory bowel disease. Intestinal epithelial cells are also in close proximity to the commensal gut microbiota and they too are known to regulate intestinal turnover. In the work presented here, we assessed the role of SOCS3 in normal intestinal homeostasis and how microbe-mediated proliferation impacts upon this. These experiments were performed in three different biological models, allowing us to assess the impact of SOCS3-regulated homeostasis at the molecular level, the tissue level, and at an organismal level, and also determine whether the function of SOCS proteins is conserved across different biological systems. Using in vitro human intestinal epithelial cells, mice and Drosophila melanogaster, we observed consistent negative effects upon reduction of SOCS3, which affected proliferation and cytokine profiles, tumour tolerance and, survival and the gut-brain axis, in our respective models. In both in vivo models, we were able to discover functional outcomes due to reduced SOCS3 expression, in the form of facilitation of helminth expulsion and increased stress resistance in mice and Drosophila, respectively, thus suggesting potential benefits of reduced SOCS3 in young animals whose ability to adapt to homeostatic changes is higher. Overall, we were able to deduce that SOCS3 is responsible for maintaining normal intestinal homeostasis, and ultimately host health, at a number of levels within multiple biological systems. However, our results also indicated that SOCS3 is a complex, multi-functional protein, with much investigation still needed to determine its complete role.

Plant extracts are known to have beneficial effects in chronic inflammatory conditions such as diabetes. This study investigated the effects of Trigonella foenum-graecum L. (Fenugreek) seed extracts on inflammatory responses of macrophages. A cell-bioassay guided extraction identified a methanolic extract of fenugreek seeds (FME) with potent anti-inflammatory effects. This extract significantly reduced secretion of pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) in THP-1 macrophages (dTHP-1) stimulated with glycated-BSA. The effect of FME on TNF-α was also reproduced in peripheral blood mononuclear cells (PBMCs) and when dTHP-1 cells were stimulated with LPS. The effect of FME on the polarization of dTHP-1 towards the M2 phenotype at the expense of the M1 phenotype was also confirmed through increased expression of IL-10 and Dectin-1. Although FME did not significantly enhance the gene expression of the M2 marker, IL-1Ra alone, FME acted synergistically in the presence of IL-4, increasing the expression of IL-10, Dectin-1 and IL-1Ra. A C-glycosidic flavonoid, Vicenin-2 (V-2), which was found to be present in FME, demonstrated similar actions to FME, by significantly suppressing pro-inflammatory cytokine release and upregulating M2 marker (IL-10). V-2 also increased Dectin-1 gene expression, synergistically, in the presence of IL-4. The regulatory mechanisms involved in the actions of FME and V-2 were explored using a gene reporter assay. Both FME and V-2 significantly (p < 0.05) inhibited the activity of nuclear transcription factor, NF-κB. Also, FME significantly (p < 0.05) increased the activity of PPAR-γ. V-2 effects appeared to be mediated through homodimerization of the p50 subunit of NF-κB and suppression of IκB-α phosphorylation. V-2 exerted anti-inflammatory effects in endothelial cells (HUVEC) by reducing TNF-α induced MCP-1 expression. V-2 also enhanced release of extracellular vesicles (EVs) from gBSA treated dTHP-1 cells. Additionally, the EVs derived from dTHP-1 cells down-regulated expression of inflammatory markers in HUVECs; ICAM-1 and VCAM-1 significantly, and MCP-1, non-significantly. In conclusion this study suggests a beneficial effect of FME and V-2 in regulating macrophage inflammatory responses. Further studies are required to clarify their role on EV expression, characterization and endothelial cell function.

No description available.

The simultaneous integration of positron emission tomography (PET) and magnetic resonance imaging (MRI) promises to combine superior functional and anatomical information into a single examination, but poses a number of engineering challenges. In particular, the development of MRI-compatible photodetectors is a key issue to realize PET/MRI at its best. In 2009, a novel photodetector, the so-called digital SiPM, was presented. The digital SiPM combines an array of Geiger-mode photodiodes with a direct digital readout and timestamper on a single silicon die. Its high integration and expected immunity to magnetic fields makes the digital SiPM very promising for simultaneous PET/MRI. Related to the development of the world’s first digital SiPM based simultaneous PET/MRI system, this thesis focuses on three specific engineering aspects of simultaneous PET/MRI. First, an MRI-compatible photodetector stack based on digital SiPMs has been developed, investigated and optimized. The final design is virtually immune to the MRI environment and enables the full potential of the digital SiPM to be used for simultaneous PET/MRI, including its time-of-flight capabilities. No evidence was found that the digital SiPM itself is affected by the MRI environment. Second, based on carbon fibre composites, a novel modular radio frequency (RF) shielding concept for MRI-compatible PET detectors has been developed. In contrast to common copper based shields, carbon fibre composites are less susceptible to eddy currents because of their lower sheet conductance. It is shown, experimentally and analytically, that carbon fibre composites can provide higher RF shielding efficiency to sheet conductance ratios than conventional copper shields can do, which makes them beneficial for simultaneous PET/MRI. Third, experimental and simulation approaches for the optical design of pixelated scintillation detectors have been assessed, in particular to improve the depth-of-interaction encoding of pixelated scintillation detectors. A novel pixelated scintillator configuration is proposed, which avoids the use of inter-crystal reflectors.

In this thesis project, we model a sub-part of the adaptive immune system, composed of B and T lymphocytes, which interact to produce a suitable immune response against antigens. From a statistical mechanics perspective, this system can be modelled as a bipartite network with sparse links where the nodes represent B and T cells respectively, signalling via particular proteins called cytokines. Assuming that B lymphocytes evolve on a faster timescale than T cells, we study the dynamics of an effective mono-partite graph of T cells only where the B cells have been integrated out. Interestingly, this system can be mapped into a Hopfield-like associative network, which is able to retrieve and perform multiple immune strategies simultaneously. Using techniques such as Kramers-Moyal expansions for master equations, we carry out a dynamical analysis of the network evolving via Glauber sequential update. We derive equations quantifying the evolution in time of the immune response strength, analysing the nature and the stability of the stationary solutions in different regimes of dilution and network connectivity via linear stability analysis and Monte Carlo simulations. The model has also been extended to include the effect of receptors promiscuity, sampling B-T interactions locally from heterogeneous degree distributions and the effect of the antigens. Finally, we introduce interactions between B lymphocytes, called idiotypic interactions, studying their effect on the system's dynamics. We also analyse the effect of idiotypic interactions in the high storage and finite connectivity regime at equilibrium, using the cavity method to derive equations for the distributions of observables of the system. In particular, we obtain the B clone size distribution, studying its behaviour in different regions of the phase space.

This thesis looks at clinical and biological aspects of vein graft stenoses in order to improve on existing management strategies and to explore the possibility for a new pharmacological therapy using antagonists of the endothelin system. After an overview of peripheral vascular disease, the introductory chapters discuss vein graft surveillance, intimal hyperplasia and properties of the vasoactive peptide, endothelin. The work described consists of clinical and laboratory based research. In the clinical chapters a retrospective study analysed the influence of patient factors on the outcome of lower limb vein grafting in the current era of postoperative vein graft surveillance. Following this, two prospective studies examined specific aspects of graft surveillance. Firstly, the predictive value of pre-discharge duplex vein graft scans was determined. The second study validated the criteria for intervention in duplex detected vein graft flow abnormalities. The first laboratory experiments set out to determine the effect of endothelin and endothelin receptor antagonists on proliferation in isolated venous smooth muscle cells. Following this, an organ culture system, a more representative model of intimal hyperplasia, was used to demonstrate the association between endothelin production and development of intimal hyperplasia. Using the same model, a series of experiments were then performed to determine the effect of endothelin inhibition. Endothelin was inhibited at the level of its synthesis, and by none selective and then selective receptor blockade. The final chapter summarises and concludes the main findings and discusses areas of future work that could arise from this thesis.

The aims of these studies were to characterise the MC IgA receptor, compare it to other established classes of IgA receptor and examine its modulation by cytokines and growth factors. For these studies primary MC cultures were established from the normal poles of kidney's removed from carcinoma. IgA binding was analysed by flow cytometry, and binding specificity determined by competitive inhibition. The receptor identified was a Fc alpha receptor (FcR) with medium affinity (K 107 M-1) for IgA and was immunogenically distinct from the myeloid Fcr, FcRI. However, the two receptors may share a degree of molecular homology as three mRNA transcripts with partial identity to FcRI were detected in all MC cultures. FcR expression was up-regulated in a time and concentration-dependent manner following exposure to IL-1, IL-6, TNF- and IFN-. Co-exposure with the Th2 cytokine IL-4 antagonised the effects of both IL-1 and TNF-. In all cases a single population of receptors was up-regulated and receptor affinity remained unaltered. Immunoblotting of MC lysates revealed five separate IgA-binding proteins, three of which were also isolated by affinity chromatography from surface radioiodinated MC. The specificity of these proteins for IgA was confirmed in competitive re-binding experiments. Whether these proteins represent subunits or multiple forms of a single receptor, or separate IgA receptors is not yet clear. Studies are now in progress to sequence the purified receptor proteins. This will ultimately allow full molecular and biochemical characterisation of the MC FcR.

Doppler ultrasound is used clinically to detect stenosis in the carotid artery. The presence of stenosis may be identified by disturbed flow patterns distal to the stenosis which cause spectral broadening in the spectrum of the Doppler signal around peak systole. This thesis investigates the ability of the short-time Fourier transform (STFT) and the autoregressive (AR) spectral estimators to perform time-frequency analysis of the non-stationary Doppler signal. Quantitative analysis of the degree of spectral broadening was measured using the spectral broadening index (SBI). A real-time system was developed using a modern DSP board combined with an IBM PC-compatible computer to analyse the Doppler signal in real-time using the STFT and AR algorithms. The spectral estimators were compared using simulated Doppler spectra contaminated with noise over a range of signal-to-noise ratios (SNRs) and also real clinical Doppler signals recorded from both healthy subjects and patients with varying degrees of stenosis. The SBI was calculated using the mean and maximum frequency envelopes which were extracted from the STFT and AR sonograms using a threshold at -6 dB of the maximum component of each individual spectrum. The results of the analysis shows a strong correlation between the indices calculated using the FFT and AR algorithms. A qualitative improvement in both the appearance of the AR sonograms and the shape of the individual AR spectra was noticeable, however, the estimation of SBI for short data frames is not significantly improved using AR. The final section of this thesis describes the wavelet transform (WT) and illustrates its application to Doppler ultrasound with two examples. Firstly, it is shown how wavelets can be used as an alternative to the STFT for the extraction of the time-frequency distribution of Doppler ultrasound signals. Secondly, wavelet-based adaptive filtering is implemented for the extraction of maximum blood velocity envelopes in the post processing of Doppler signals.

CD23 is the low affinity IgE receptor. It is a type II integral transmembrane glycoprotein that can be shed from the cell surface forming soluble products of approximately 37, 33, 29, 25 and 16kDa. It appears that membrane and soluble CD23 have opposing regulatory functions and that inhibition of CD23 shedding may have potential to alleviate both allergic and inflammatory diseases. This has focused attention on the endogenous protease(s) responsible for CD23 shedding, leading to the demonstration that both the 37 and 33kDA sCD23 fragments are cleaved from the cell surface by a metalloprotease. In order to characterise the cleavage events within CD23, anti-neoepitope antibodies specific for the newly created amino and carboxy termini of the two predominant cleavage sties within CD23 (producing the 37 and 25kDa soluble CD23 products) were raised. Characterisation of these antibodies demonstrated that the proteolytic cleavage events responsible for creating the 37 and 25kDa sCD23 fragments are independent of each other. Furthermore, two different proteases were shown to be responsible for cleaving these two fragments. The work described in this thesis confirms previous reports that 37kDA sCD23 is cleaved by a metalloprotease, however cleavage of the 25kDa fragment was not inhibited by metalloprotease inhibitors. The production of the two different sized sCD23 molecules by different proteases has important implications for targeting the proteolytic cleavage events to alleviate symptoms of allergic and inflammatory diseases. This emphasises the importance of defining the biological functions of mCD23 and each of the sCD23 molecules.

Simultaneous PET-MR imaging has recently emerged into routine clinical use, with significant role in diagnosis and treatment. The high sensitivity of a PET scanner, combined with the anatomical and functional information given by MR can focus to a molecular level and provide simultaneous information about individual but correlated parameters. The combination of these modalities has also brought new opportunities. One of these is the potential for the MRI to capture and measure patient motion during PET scanning. This motion information, in the form of motion fields can be applied back to the PET and correct any motion artefacts created. For the success of this study many methodological problems have to be resolved. This thesis investigates the methodology for motion correction of PET data by appropriate use of MR motion information, which could in due course become a standard motion correction method for both pre-clinical and human studies. In particular, this study utilized prototype pre-clinical MR-compatible PET inserts. Following the phantom development and during the experimental procedures, various prototype simultaneous MR-compatible PET small-diameter scanners, in conjunction with a 3T clinical MR scanner, were used to validate the phantom and evaluate several different approaches to motion correction. All scanner combinations used were a combination of a removable PET insert operating within a clinical MR scanner so it was necessary to develop robust methods to ensure spatial alignment and temporal synchronization. The data obtained from simultaneous PET and MR acquisition was arranged into gates and reconstructed by deriving motion transformations from the MR data and using this to correct the PET data for the effects of motion using several PET reconstruction approaches. The experimental results obtained with various PET-MR systems demonstrate the feasibility of this approach and the degree of motion correction that can realistically be expected in pre-clinical PETMR investigations.