Wing trailing vortex paths in formation flight

Includes bibliographical references.Formation flight has been shown to reduce the induced drag for a formation of aircraft. The mechanism by which this is achieved is caused by the wake velocity field of the aircraft. This field is dominated by wing-tip trailing vortices. The paths of these vortices become too complex for rigid wake models downstream of the second aircraft in the formation. To tackle this problem, a combined vortex lattice and vortex filament numerical model was developed. For each simulation the vortex lattice model determined the lift distribution which was applied to the vortex filament model. The vortex filament model used the Burnaham-Hallock vortex profile with a core size of 5% of the wing span to eliminate numerical instabilities. Individual components of the model were verified successfully against literature and the overall approach was validated against wind tunnel data. The wind tunnel data was extracted from apparatus designed and build as part of this study. The apparatus consisted of two NACA 0012 rectangular planform wings mounted in various formation positions and a tuft grid placed downstream of the wings to visualise the vortex paths. Test were performed with both wings at 8◦ angle of attack. Span-wise wing-tip overlap distances were set at 38%, 10%, 0% and -10% of the span, where 0% implies wing-tip alignment and a positive value indicates a wing-tip overlap. Vertical separations were set at -3%, 0% and 3% of the span for each span-wise wing-tip overlap condition apart from 38% which was only tested at 0 vertical separation. The formation outboard vortex paths were predicted well within the 3% span accuracy of the tuft grid. The predictions of the paths of the formation inboard vortices, however were less accurate. The errors were attributed to a combination of bias errors in the experimental apparatus as well as the pseudo-viscous effects of the Burnham-Hallock vortex profile

Mechanisms of amyloid fibril-mediated toxicity

Amyloid diseases are a group of protein misfolding disorders characterised by the formation of highly ordered filamentous assemblies known as amyloid fibrils. Soluble aggregation intermediates whose formation precedes that of mature fibrils are commonly considered the major source of toxicity in amyloid diseases. Oligomer toxicity has often led to mature amyloid fibrils being referred to as inert end products of aggregation. Recent evidence, however, has shown that fibrils themselves are capable of facilitating toxicity by a variety of mechanisms. One such protein, β2-microglobulin (β2m), has been shown to form amyloid fibrils that bind to liposome membranes causing deformations and disruption in a pH dependent manner. In this thesis, the in vitro mechanisms of β2m fibril-induced membrane disruption are explored to elucidate why reducing the pH from 7.4 to 6.4 leads to an enhancement in fibril membrane disruption. A combination of chemical kinetics, NMR and liposome dye-release assays show that at both pH values, membrane disruption is mediated through the shedding of soluble species induced upon diluting fibrils formed at low pH into either buffer. Fibril depolymerisation at pH 6.4 leads to the persistence of membrane-active non-native species, whereas depolymerisation at pH 7.4 is driven rapidly to membrane-inactive native monomer. Further analysis reveals that these non-native species are structurally disordered, spherical particles that display significant surface-exposed hydrophobicity. The observed pH-dependent formation of oligomers shed during depolymerisation is likely to play an important role in mediating cellular effects upon incubation with fibrils. Consistent with this, chemical cross-linking of fibrils and co-incubating fibrils with Hsp70-1A prevents the depolymerisation of β2m fibrils at both pH 7.4 and 6.4 in vitro and reduces metabolic defects associated with β2m fibril depolymerisation. The results suggest that kinetically stabilising fibrils to prevent molecular shedding could be a means of helping to remedy amyloid-associated toxicity

Human immunodeficiency virus infection and older adults: A retrospective single-site cohort study from Johannesburg, South Africa

Introduction: HIV-infected adults aged over 50 years in South Africa are increasing. This study explored differences between baseline characteristics and 12-month outcomes of younger and older HIV-infected adults initiated on antiretroviral therapy (ART). Additionally, associations with outcomes within the older group were sought.Methods: We retrospectively reviewed treatment-naive HIV-infected adult patients at ART initiation. Patients aged 18.0–39.9 years were compared to patients aged over 50 years using log-binomial regression for baseline characteristics and 12-month outcomes. Within the older group, outcome associations were found using multivariate regression.Results: The older cohort (n = 1635) compared to the younger cohort (n = 10726) comprised more males (47.2% vs. 35.4%, PR 1.52, p < 0.05), smokers (12.9% vs. 9.7%, PR 1.32, p < 0.05) and overweight patients (26.0% vs. 20.0%, PR 1.32, p < 0.05). Fewer older patients had tuberculosis (10.2% vs. 15.3%, PR 0.67, p < 0.05), other opportunistic infections (16.9% vs. 23.3%, PR 0.70, p < 0.05), World Health Organization stage 3/4 disease (39.9% vs. 43.2%, PR 0.89, p < 0.05), anaemia (22.8% vs. 28.4%, PR 0.77, p < 0.05), liver dysfunction (17.1% vs. 21.3%, PR 0.83, p < 0.05) or low CD4+ count < 100 cells/mm3 (56.3% vs. 59.9%, PR 0.71, p < 0.05).Mortality was higher in the older cohort (11.3% vs. 7.5%, PR 1.48, p < 0.05). Virological suppression was greater in the older cohort (89.5% vs. 86.5%, PR 1.28, p < 0.05) but CD4+ restitution was lower (62.8% vs. 75.0%, PR 0.61, p < 0.05). There was no difference in treatment complications between the groups.Within the older cohort, associations with death were as follows: age > 55 years (PR 1.47, p < 0.05), an AIDS-defining condition (PR 2.28, p < 0.05), raised ALT (PR 1.53, p < 0.05) and CD4+ < 100 cells/mm3 (PR 2.15, p < 0.05). Associations with favourable treatment response at 12 months were unemployment (PR 1.18, p < 0.05) and raised ALT (PR 1.19, p < 0.05). Associations with a treatment complication at 12 months were unemployment (PR 1.12, p < 0.05), smoking (PR 1.20, p < 0.05) and nevirapine use (PR 1.36, p < 0.05) but secondary education was protective (PR 0.87, p < 0.05).Conclusion: HIV-infected South African adults aged over 50 years differ in characteristics and outcomes compared to their younger counterparts and justify specialised management within HIV treatment facilities

Imaging Drug Uptake by Bioorthogonal Stimulated Raman Scattering Microscopy

Stimulated Raman scattering (SRS) microscopy in tandem with bioorthogonal Raman labelling strategies is set to revolutionise the direct visualisation of intracellular drug uptake. Rational evaluation of a series of Raman-active labels has allowed the identification of highly active labels which have minimal perturbation on the biological efficacy of the parent drug. Drug uptake has been correlated with markers of cellular composition and cell cycle status, and mapped across intracellular structures using dual-colour and multi-modal imaging. The minimal phototoxicity and low photobleaching associated with SRS microscopy has enabled real-time imaging in live cells. These studies demonstrate the potential for SRS microscopy in the drug development process

Kinetic analysis of bioorthogonal reaction mechanisms using Raman microscopy

Raman spectroscopy is well-suited to the study of bioorthogonal reaction processes because it is a non-destructive technique, which employs relatively low energy laser irradiation, and water is only very weakly scattered in the Raman spectrum enabling live cell imaging. In addition, Raman spectroscopy allows species-specific label-free visualisation; chemical contrast may be achieved when imaging a cell in its native environment without fixatives or stains. Combined with the rapid advances in the field of Raman imaging over the last decade, particularly in stimulated Raman spectroscopy (SRS), this technique has the potential to revolutionise our mechanistic understanding of the biochemical and medicinal chemistry applications of bioorthogonal reactions. Current approaches to the kinetic analysis of bioorthogonal reactions (including heat flow calorimetry, UV-vis spectroscopy, fluorescence, IR, NMR and MS) have a number of practical shortcomings for intracellular applications. We highlight the advantages offered by Raman microscopy for reaction analysis in the context of both established and emerging bioorthogonal reactions, including the copper(i) catalysed azide-alkyne cycloaddition (CuAAC) click reaction and Glaser-Hay coupling

Ratiometric sensing of fluoride ions using Raman spectroscopy

Ratiometric Raman spectroscopy represents a novel sensing approach for the detection of fluoride anions based on alkyne desilylation chemistry. This method enables rapid, anion selective and highly sensitive detection of fluoride in a simple paper-based assay format using a portable Raman spectrometer

Label-free imaging of lipid droplets in prostate cells using stimulated Raman scattering microscopy and multivariate analysis

Hyperspectral stimulated Raman scattering (SRS) microscopy is a powerful imaging modality for the analysis of biological systems. Here, we report the application of k-means cluster analysis (KMCA) of multi-wavelength SRS images in the high wavenumber region of the Raman spectrum, as a robust and reliable method for the segmentation of cellular organelles based on the intrinsic SRS spectrum. KMCA has been applied to the study of the endogenous lipid biochemistry of prostate cancer and prostate healthy cell models, whilst the corresponding SRS spectrum of the lipid droplet cluster enabled direct comparison of their composition. The application of KMCA in visualising the lipid droplet (LD) content of prostate cell models following inhibition of de novo lipid synthesis (DNL) using the acetyl-coA carboxylase inhibitor, 5-(tetradecyloxy)-2-furoic acid (TOFA) is demonstrated. This method identified a reliance of prostate cancer cell models upon DNL for metabolic requirements, with a significant reduction in the cellular lipid droplet content after treatment with TOFA, which was not observed in normal prostate cell models. SRS imaging combined with KMCA is a robust method for investigating drug-cell interactions in a label-free manner

Label-free cytometric evaluation of mitosis via stimulated Raman scattering microscopy and spectral phasor analysis

Hyperspectral stimulated Raman scattering (SRS) microscopy is a robust imaging tool for the analysis of biological systems. Here we present a unique perspective, a label-free spatiotemporal map of mitosis by integrating hyperspectral SRS microscopy with advanced chemometrics to assess the intrinsic biomolecular properties of an essential process of mammalian life. The application of spectral phasor analysis to multi-wavelength SRS images in the high-wavenumber (HWN) region of the Raman spectrum enabled the segmentation of subcellular organelles based on innate SRS spectra. Traditional imaging of DNA is primarily reliant on using fluorescent probes or stains which can affect the biophysical properties of the cell, here we demonstrate the label-free visualization of nuclear dynamics during mitosis coupled with an evaluation of its spectral profile in a rapid and reproducible manner. These results provide a snapshot of the cell division cycle and chemical variability between intracellular compartments in single cell models, which is central to understanding the molecular foundations of these fundamental biological processes. The evaluation of HWN images by phasor analysis also facilitated the differentiation between cells in separate phases of the cell cycle based solely on their nuclear SRS spectral signal, which offers an interesting label-free approach in combination with flow cytometry. Therefore this study demonstrates that SRS microscopy combined with spectral phasor analysis is a valuable method for detailed optical fingerprinting at the subcellular level

A Palette of Minimally Tagged Sucrose Analogues for Real‐Time Raman Imaging of Intracellular Plant Metabolism

Sucrose is the main saccharide used for long-distance transport in plants and plays an essential role in energy metabolism; however, there are no analogues for real-time imaging in live cells. We have optimised a synthetic approach to prepare sucrose analogues including very small (≈50 Da or less) Raman tags in the fructose moiety. Spectroscopic analysis identified the alkyne-tagged compound 6 as a sucrose analogue recognised by endogenous transporters in live cells and with higher Raman intensity than other sucrose derivatives. Herein, we demonstrate the application of compound 6 as the first optical probe to visualise real-time uptake and intracellular localisation of sucrose in live plant cells using Raman microscopy

A new class of ratiometric small molecule intracellular pH sensors for Raman microscopy

Intracellular pH (pHi) homeostasis is intertwined with a myriad of normal cellular behaviors as well as pathological processes. As such, small molecule probes for the measurement of pHi are invaluable tools for chemical biology, facilitating the study of the role of pH in cellular function and disease. The field of small molecule pHi sensors has traditionally been dominated with probes based on fluorescent scaffolds. In this study, a series of low molecular weight (<260) oligoyne compounds have been developed which exhibit pH sensitive alkyne stretching frequencies (νalkyne) in Raman spectroscopy. The modular design of the compounds enabled tuneability of their pKa(H) through simple structural modification, such that continuous pH sensitivity is achieved over the range 2-10. Alkyne stretching bands reside in the 'cell-silent' region of the Raman spectrum (1800-2600 cm-1) and are readily detectable in a cellular environment with subcellular spatial resolution. This enabled the application of a pH sensitive oligoyne compound to the ratiometric sensing of pHi in prostate cancer (PC3) cells in response to drug treatment. We propose that probes based on Alkyne Tag Raman Imaging offer an entirely new platform for the sensing of pHi, complementary to fluorescence microscopy