Cardiac Hypertrophy, Heart Failure, and Regulation of Microtubule Stability

This work focused on the regulation of the different isotypes of beta-tubulin during cardiac hypertrophy, with the specific hypothesis that increased levels of the beta I isotype leads to the increase in microtubule density seen during hypertrophy. Partial cDNAs for the three beta-tubulin isotypes expressed in the heart (I, II, and IV) have been cloned and used to show that there is a differential upregulation of the beta I and beta II isotypes during cardiac hypertrophy. Messenger RNA half-life studies of cells isolated from hypertrophied myocardium have shown that this upregulation is due to increased transcription of the genes encoding those isotypes. This work has also shown the presence in terminally differentiated cells of the co-translational autoregulatory mechanism by which increased free tubulin protein levels leads to decreased mRNA levels. The beta-tubulin isotypes that are expressed in the heart have been cloned into tetracycline-regulated expression vectors and adenoviral vectors. Overexpression in CHO cells has shown that, contrary to some previous reports, overexpression of beta I, beta II, and beta IV does not affect microtubule stability as measured by taxol resistance. In addition, overexpression of beta I in adult cardiocytes does not affect microtubule stability. However, overexpression of the predominant heart microtubule associated protein, MAP 4, is alone enough to cause increased microtubule stability in vitro. Because there is an increase in MAP 4 protein levels during cardiac hypertrophy in vivo, we believe that this is the primary mechanism of increased microtubule stability in the hypertrophied myocardium. This work also focused on development of a mouse model in which to study the effects of microtubule stability on the development of heart failure in vivo. Mutation of the beta-tubulin protein in a string of leucine residues destabilized the microtubules in CHO cells. Constructs encoding these mutant tubulin proteins have been inserted into a heart-specific vector and cloned into transgenic mice. Several lines have been initially characterized for levels of expression, and one line has shown heart-specific expression of a mutated beta-tubulin protein. These lines will be used for future studies of the effect of destabilized microtubules on the development of heart failure

Effects of Irradiation and Thermal Annealing on the Mechanical and Microstructural Properties of Bulk Metallic Glasses

A series of ion irradiation and annealing experiments have been performed on Zr52.5Cu17.9Ni14.6Al10Ti5 “BAM-11” and Cu60Zr20Hf10Ti10 bulk metallic glass (BMG) specimens to evaluate their irradiation- and temperature-induced microstructural and mechanical property evolution. These experiments covered four main themes, namely, ion irradiation, neutron irradiation, thermal annealing, and helium implantation. For the ion irradiations, samples were exposed to 9 MeV Ni and 5.5 MeV C ions at temperatures ranging from room temperature to 360 oC. For the Ni ion irradiations the samples were exposed to midrange (~1.5 m depth) doses of 0.5 and 10 displacements per atom (dpa), while the C ion irradiations samples were irradiated to a midrange dose of 0.5 dpa. For the neutron irradiations, samples were irradiated by neutrons (E \u3e 0.1 MeV) at ~70 oC to fluences of 1.4 × 1020 n/cm2 and 1.4 × 1021 n/cm2 (doses of 0.1 and 1 dpa). Thermal annealing experiments involved heating the samples to various temperatures ranging from 25 - 770 oC. For the helium implantation experiments, amorphous and partially crystallized BMGs were exposed to helium fluences of 2 × 1015 and 5 × 1015 cm-2 . The mechanical property and microstructural characterization included nanoindentation, compression testing, bend testing, Xray diffraction (XRD), neutron diffraction, thermal desorption analysis (TDS), and nuclear reaction analysis. From the experiments, several important conclusions were obtained. The results of the XRD and nanoindentation characterizations of the ion irradiated and thermal annealed specimens indicate good stability during irradiation at 25 to 290 oC up to at least 10 dpa but suggest that the BAM-11 BMG is not suitable for irradiation environments where temperatures exceed 300 oC for prolonged periods of time. As for the neutron irradiation and thermal annealing experiments, significant softening was observed in the sample irradiated by neutrons, while postirradiation annealing led to a marked increase in hardening. Neutron diffraction results indicated vi that primary knock-on events caused rejuvenation (disordering) while annealing resulted in structural relaxation. The results of the TDS experiments found that for the lower He implantation fluence, He desorbed more quickly in the partially crystallized alloy, indicating a structural effect on the mobility of He

Bridging Control-Centric and Data-Centric Optimization

With the rise of specialized hardware and new programming languages, code optimization has shifted its focus towards promoting data locality. Most production-grade compilers adopt a control-centric mindset - instruction-driven optimization augmented with scalar-based dataflow - whereas other approaches provide domain-specific and general purpose data movement minimization, which can miss important control-flow optimizations. As the two representations are not commutable, users must choose one over the other. In this paper, we explore how both control- and data-centric approaches can work in tandem via the Multi-Level Intermediate Representation (MLIR) framework. Through a combination of an MLIR dialect and specialized passes, we recover parametric, symbolic dataflow that can be optimized within the DaCe framework. We combine the two views into a single pipeline, called DCIR, showing that it is strictly more powerful than either view. On several benchmarks and a real-world application in C, we show that our proposed pipeline consistently outperforms MLIR and automatically uncovers new optimization opportunities with no additional effort.Comment: CGO'2

Analysis, preliminary design and simulation systems for control-structure interaction problems

Software aspects of control-structure interaction (CSI) analysis are discussed. The following subject areas are covered: (1) implementation of a partitioned algorithm for simulation of large CSI problems; (2) second-order discrete Kalman filtering equations for CSI simulations; and (3) parallel computations and control of adaptive structures

Characterisation of self-assembled engineered proteins on gold nanoparticles and their application to biosensing

PhD ThesisThe use of gold nanoparticles (AuNP) has a long and varied history, thought to cover several thousand years. More recently the unique properties of nanoscale materials have stimulated extensive work on nanoparticles and other nanomaterials leading to their use in novel technologies. AuNPs have been of particular interest for bioscience applications due to their biocompatibility and the ease with which biological molecules can be conjugated to their surface. In this study the assembly of engineered proteins, specifically the transmembrane domain of Escherichia coli outer membrane protein A (OmpATM), onto the surface of AuNPs was investigated both in solution and with the particles attached to a SiO2 substrate. AuNPs were adhered to SiO2 surfaces using a novel silane treatment developed by the industrial sponsor and were characterised using spectroscopy, electron and atomic force microscopy. The addition of a single cysteine residue to the OmpATM structure was shown, by UV-Vis and fluorescence spectroscopy, to increase protein binding at equilibrium and form higher stability protein-AuNP complexes in solution. Following this, engineered OmpATM proteins containing tandem antibody-binding domains from Streptococcal protein G were assembled on the AuNP surface and their structure interrogated using neutron and light scattering. This revealed an oriented protein layer where the functional domains extend away from the AuNP surface and are available to bind antibodies. OmpATM-AuNP conjugates were used to develop biosensing assays using both well-established methods, such as lateral flow assays, and novel spectroscopic methods, which use the unique optical properties of AuNPs. Detection of influenza A nucleoprotein, an antigen used to clinically diagnose influenza, was achieved using a bespoke anti-nucleoprotein single-chain antibody domain fused to OmpATM and assembled on 20 nm diameter AuNPs. The results demonstrate that engineered OmpATM proteins conjugated to AuNPs can be used to develop novel diagnostics using a range of read out technologies

Membrane functionalisation using polyrotaxanes with amphiphilic cyclodextrins

This work is aimed at the design and characterisation of a new family of tethered ligands, called sliding tethered ligands (STLs). They are based on topological complexes between polymers and amphiphilic cyclodextrins (CDs), which can be inserted into phospholipid membranes. At first we investigate the membrane insertion properties of amphiphilic cholesteryl CD derivatives, which are suitable membrane anchors for the STLs. With the help of neutron reflectivity it can be demonstrated that the CD residues show a remarkable conformational adaptability and that the CD cavities remain accessible upon insertion into lipid model membranes. We have developed a synthetic pathway to assemble the STLs from polyrotaxanes with a controlled low number of mono-modified azido-α-CDs, threaded on a polyethylene glycol (PEG) chain. Using newly developed in-situ capping methods the polyrotaxanes are endcapped with adamantane ligands, which can be recognized by a β-CD receptor. Furthermore a cholesteryl anchor is attached to the threaded CD in order to enable the STL to insert into membranes. We demonstrate that STLs readily insert into phospholipid (DPPC) model membranes using IR Absorption Reflection Spectroscopy and investigating the film morphology by Brewster Angle Microscopy and Atomic Force Microscopy. Applying neutron reflectivity it is shown, that for sufficiently high polymer densities the STLs form polymer brushes, which follow the scaling laws predicted by the mean field theory. Using the surface force apparatus it is evidenced that model membranes modified with STLs and cholesteryl β-CD receptors give rise to typical tethered ligand - receptor interaction profiles

52nd Rocky Mountain Conference on Analytical Chemistry

Final program, abstracts, and information about the 52nd annual meeting of the Rocky Mountain Conference on Analytical Chemistry, co-endorsed by the Colorado Section of the American Chemical Society and the Society for Applied Spectroscopy. Held in Snowmass, Colorado, August 1-5, 2010

Co-crystallisation of energetic materials – a step-change in the control of properties and performance of munitions

The research described in this thesis seeks to explore a concept that has the potential to make a step-change for the control of the properties of energetic materials (sensitivity, long-term storage, processability, performance, etc.), resulting in safer munitions with enhanced performance. This concept is co-crystallisation and involves crystallisation of the energetic material with one or more molecular components in order to modify the properties of the composition. The concept has been demonstrated in the pharmaceutical sector as a successful means of altering the physical properties of active pharmaceutical ingredients, e.g. solubility, bioavailability, stability to humidity. This project therefore aims to exploit the concepts of crystal engineering and co-crystallisation as applied to selected energetic materials in order to achieve the following objectives: (i) develop an enhanced understanding of how structure influences key properties such as sensitivity, (ii) control the sensitivity of existing, approved energetic materials, and (iii) identify new energetic materials with enhanced properties, e.g. reduced sensitivity, higher performance, and increased thermal stability. The compound 3,5-nitrotriazolone (NTO) was crystallised with a selection of co-formers to produce salts and co-crystals. The structure properties of these materials were explored using single-crystal and powder X-ray diffraction, and structural features were correlated with properties such as crystal density, difference in pKa of co-formers, thermal properties, and sensitivity to impact. Detonation velocities of the co-crystals were calculated based on densities, chemical composition, and heats of formation. Co-former molecules included a series of substituted anilines, substituted pyridines (including 4,4’-bipyridine, 2-pyridone), and substituted triazoles. A co-crystal was formed between NTO and 4,4’-bipyridine on crystallisation from ethanol, whilst a salt was formed when crystallised from water. Upon heating the salt to 50ºC, the co-crystal was formed. Structural differences between the salts formed by NTO with 3,5-DAT and 3,4- DAT were correlated with structural features. 3,5-DAT.NTO is substantially less impact sensitive than 3,4-DAT.NTO, and this is attributed to the layered structure of 3,5-DAT.NTO. An investigation into triazole-based NTO salts under high pressure was conducted. A new polymorph of 3,5-DAT.NTO was discovered upon increasing the pressure to 2.89 GPa. The high-pressure phase appears to retain the layered structure and remains in this phase up to 5.33 GPa, although it was not recoverable upon decompression to atmospheric pressure. The compression behaviour of the unit cell volume for phase I of 3,5-DAT.NTO has been fitted to a 3rd-order Birch- Murnaghan equation of state (EoS) with V0 = 957.7 Å3, B0 = 8.2 GPa and B’0 = 14.7. The unit cell was found to be most compressible in the a and c directions. Under high pressure 3,4-DAT.NTO does not give any indication of a phase change occurring up to 6.08 GPa. The coefficients of the 3rd-order Birch-Murnaghan EoS have been determined to be V0 = 915.9 Å3, B0 = 12.6 GPa and B’0 = 6.5