BACKGROUND:Nemaline myopathy (NM) is a congenital muscle disease associated with weakness and the presence of nemaline bodies (rods) in muscle fibers. Mutations in seven genes have been associated with NM, but the most commonly mutated gene is nebulin (NEB), which is thought to account for roughly 50% of cases.RESULTS:We describe two siblings with severe NM, arthrogryposis and neonatal death caused by two novel NEB mutations: a point mutation in intron 13 and a frameshift mutation in exon 81. Levels of detectable nebulin protein were significantly lower than those in normal control muscle biopsies or those from patients with less severe NM due to deletion of NEB exon 55. Mechanical studies of skinned myofibers revealed marked impairment of force development, with an increase in tension cost.CONCLUSIONS:Our findings demonstrate that the mechanical phenotype of severe NM is the consequence of mutations that severely reduce nebulin protein levels and suggest that the level of nebulin expression may correlate with the severity of disease.

published_or_final_version / Surgery / Doctoral / Doctor of Philosophy

As part of the effort to increase the reliability of the MMT Observatory (MMTO) 6.5m Primary Mirror Coating System, the specified filament has changed from a configuration in which the aluminum charge is hand wound around a tungsten filament to a configuration in which the aluminum is integrally wound with the tungsten at the time of filament manufacture. In the MMTO configuration, this filament consists of the three strands of tungsten wire and one strand of aluminum wire. In preparation of a full system test utilizing two hundred filaments fired simultaneously, an extensive testing program was undertaken to characterize these filaments using a four filament configuration in the MMTO small coating chamber (0.5m) and then a forty filament configuration in the University of Arizona Steward Observatory coating chamber (2m). The testing using the smaller coating chambers has shown these filaments provide very predicable coatings from test to test, and with the proper heating profile, these filaments greatly reduce the likelihood of aluminum drips. The initial filament design was modified during the course of testing by shortening the unwound filament length to closer match the aluminum load required in the MMTO coating chamber. This change increased the aluminum deposition rates without increasing the power delivered of the filament power supplies (commercial welders). Filament power levels measured at the vacuum chamber feedthroughs, currents, and deposition rates from multiple coating tests, including a full system test, are presented.

Natural fibre composites (NFCs), which are polymers reinforced with cellulosic bast fibres, have the potential to be applied into a range of building products. They are seen as an alternative to glass fibre reinforced plastics (GFRP) in some applications, because of natural fibres (NF) relatively high strength and low density. Moreover, natural fibres have a set of beneficial traits, such as thermal insulation, thermal stability, biodegradability, and are inherently renewable. Those characteristics are of importance when NF are used as reinforcements in polymer composites, but developments in mechanical performance, reliability and economic viability are still required in order to be adopted fully by industry. The goal of this thesis was the development of a processing methodology for NFC laminate and subsequent material characterisation to assess the developed material suitability for building applications. Research objectives included materials selection, processing route development for laminates and tubes, manufacture of NFC laminates and analysis of mechanical properties in order to find an optimal composition. Hemp and flax fibres were selected as the reinforcement, because both have high mechanical properties and are important bast fibre crops in the European region with established cultivation and processing methods. As a matrix, fossil-fuel based and partially bio-derived thermoset resin systems were used. Handling and processing methodologies were developed for laminates and composite tubes based on filament winding and compression moulding techniques. The effects of the selected factors, namely material composition, volume fraction, processing parameters, reinforcement linear density, yarn twist, lamination sequence, yarn waviness and hybrid hemp-wool reinforcement were subsequently described in mechanical properties analysis of laminates. The influence of weathering conditions on the mechanical performance of the NFCs was examined. Furthermore, a study of NFC tubes under compression was performed. Results showed that the developed laminates reinforced with NF yarns have sufficient mechanical properties to be utilised in sandwich panels and/or tubes. However, a low resistance to moisture-related weathering restricts the developed NFCs for indoor applications.

This thesis is an investigation of the TPE filament for Fused Deposition Modelling (FDM) manufacturing method. All the investigations aim to optimize the quality of the filament in order to make Thermoplastic Elastomer (TPE) material possible for FDM manufacturing method. Optimization experiments were made to find out key parameters in the extrusion process that determine the quality of the filament. With the optimal parameters, further investigation of the additive content in the TPE granulate was made to solve the current problem of the filament in practical 3D printing, which the high surface friction massively affects the FDM manufacturing feasibility. The filaments were manufactured by the desktop extruder 3devo filament extruder and the surface friction tests were performed on TribotesterTM. Additionally, discussion was made to summarize the pros and cons of TPE material as well as the significance of 3D printing TPE. Potential application and benefits are mentioned for combining the property of TPE and the advantage of FDM manufacturing. Current state-of-art extrusion equipment and FDM technology are also summarized.

Actin's ability to engage in a wide range of physiological functions requires that it be subject to complex spatial and temporal regulation. This regulation is achieved internally through monomer-monomer contacts and externally through interactions with actin binding proteins. The first part of my thesis focused on better understanding the role of inter-monomeric ionic interactions proposed between subdomains 2 and 3 of opposing monomers in F-actin stabilization. I studied several yeast actin mutants: A167R to disrupt a proposed ionic attraction with R39, A167E to mimic a proposed ionic attraction in muscle actin, and D275R to disrupt a proposed ionic attraction with R39. I investigated the effects of mutations in vivo, effects on filament polymerization characteristics and appearance in vitro, as well as interaction of the mutants with the filament severing protein cofilin. While both in vivo and in vitro data demonstrated the importance of the R39-D275 interaction for yeast actin and the interaction of the filament with cofilin, disruption of this interaction alone did not cause filament fragmentation. Conversely, results with A167 do demonstrate the in vivo and in vitro importance of another potential R39 ionic interaction for filament stabilization. In the second part of my work I used amide proton hydrogen/deuterium (HD) exchange detected by mass spectrometry as a tool to gain structural insight into yeast and muscle actin and profilin isoform differences and the actin-profilin interaction. The yeast and muscle actin HD analysis showed greater exchange for yeast G-actin compared to muscle actin in the barbed end pivot region and areas in subdomains 1 and 2, and for F-actin in monomer-monomer contact areas. These results suggest greater flexibility of the yeast actin monomer and filament compared to muscle actin. For yeast-muscle hybrid G-actins, the muscle-like and yeast-like parts of the molecule generally showed exchange characteristics resembling their parent actins. There were a few exceptions to this rule, however: a peptide on top of subdomain 2 and the pivot region between subdomains 1 and 3. These exhibited muscle actin-like exchange characteristics even though the areas were yeast-like, suggesting that there is crosstalk between subdomains 1 and 2 and the large and small domains. Hybrid F-actin data showing greater exchange compared to both yeast and muscle actins are consistent with mismatched yeast-muscle actin interfaces resulting in decreased stability of the hybrid filament contacts. Actin-profilin HD exchange results demonstrated a possible differential interaction of specific profilin isoforms with specific actin isoforms. While profilin binding mostly caused a decreased exchange for yeast actin peptides, it caused an increase in exchange for muscle actin peptides. Many of the changes observed were in peptides that line or contact the nucleotide cleft, consistent with profilin's ability to alter the kinetics of nucleotide exchange.

Conductive anodic filament (CAF) is a failure mode in printed wiring boards (PWBs) which occurs under high humidity and high voltage gradient conditions. The filament, a copper salt, grows from anode to cathode along the epoxy-glass interface. Ready and Turbini (2000) identified this copper salt as the Cu2(OH)3Cl, atacamite compound. This work has investigated the influence of polyethylene glycol (PEG) and polyethylene propylene glycol (PEPG) fluxing agents on the chemical nature of CAF. For coupons processed with PEPG flux, with and without chloride, a copper-chloride containing compound was formed in the polymer matrix. This compound was characterized using x-ray photoelectron spectroscopy (XPS) as CuCl and an electrochemical mechanism for the formation of the chloride-containing CAF has been proposed. For PEG flux, with and without chloride, it has been shown that CAF only formed, but no copper containing compound formed in the matrix. It appears for PEG fluxed coupons, a PEG-Cu-Cl complex forms, binds the available Cu and acts as a barrier to the formation of CuCl in the polymer matrix. Meeker and Lu Valle (1995) have previously proposed that CAF failure is best represented by two competing reactions – the formation of a copper chloride corrosion compound (now identified as Cu2(OH)3Cl) and the formation of innocuous trapped chlorine compounds. Since no evidence of any trapped chloride compounds has been found, we propose that the formation of CAF is best represented by a single non-reversible reaction. For coupons processed with a high bromide-containing flux, bromide containing CAF was created and characterized using transmission electron microscopy (TEM) to be Cu2(OH)3Br. In addition, a copper-containing compound was formed in the polymer matrix and characterized using XPS as CuBr. An electrochemical mechanism for the formation of bromide-containing CAF has been proposed based on the XPS data. .

Conductive anodic filament (CAF) is a failure mode in printed wiring boards (PWBs) which occurs under high humidity and high voltage gradient conditions. The filament, a copper salt, grows from anode to cathode along the epoxy-glass interface. Ready and Turbini (2000) identified this copper salt as the Cu2(OH)3Cl, atacamite compound. This work has investigated the influence of polyethylene glycol (PEG) and polyethylene propylene glycol (PEPG) fluxing agents on the chemical nature of CAF. For coupons processed with PEPG flux, with and without chloride, a copper-chloride containing compound was formed in the polymer matrix. This compound was characterized using x-ray photoelectron spectroscopy (XPS) as CuCl and an electrochemical mechanism for the formation of the chloride-containing CAF has been proposed. For PEG flux, with and without chloride, it has been shown that CAF only formed, but no copper containing compound formed in the matrix. It appears for PEG fluxed coupons, a PEG-Cu-Cl complex forms, binds the available Cu and acts as a barrier to the formation of CuCl in the polymer matrix. Meeker and Lu Valle (1995) have previously proposed that CAF failure is best represented by two competing reactions – the formation of a copper chloride corrosion compound (now identified as Cu2(OH)3Cl) and the formation of innocuous trapped chlorine compounds. Since no evidence of any trapped chloride compounds has been found, we propose that the formation of CAF is best represented by a single non-reversible reaction. For coupons processed with a high bromide-containing flux, bromide containing CAF was created and characterized using transmission electron microscopy (TEM) to be Cu2(OH)3Br. In addition, a copper-containing compound was formed in the polymer matrix and characterized using XPS as CuBr. An electrochemical mechanism for the formation of bromide-containing CAF has been proposed based on the XPS data. .

Background and aims: It is known that smooth muscle possesses substantial mechanical plasticity in that it is able to adapt to large changes in length without compromising its ability to generate force. It is believed that structural malleability of the contractile apparatus underlies this plasticity. There is strong evidence suggesting that myosin thick filaments of the muscle are relatively labile and their length in vivo is determined by the equilibrium between monomeric and filamentous myosin. The equilibrium in turn is governed by the state of phosphorylation of the 20-kD regulatory myosin light chain (MLC20, or RLC). It is known that phosphorylation of the myosin light chain favors formation of the filaments; it is not known how the light chain phosphorylation affects the lability of the filaments. The major aim of this thesis was to measure the mechanical integrity of the filaments formed from purified myosin molecules from bovine airway smooth muscle, and to determine whether the integrity was influenced by phosphorylation of the myosin light chain. Methods: Myosin was purified from bovine trachealis to form filaments, in ATP containing zero-calcium solution during a slow dialysis that gradually reduced the ionic strength. Sufficient myosin light chain kinase and phosphatase, as well as calmodulin, were retained after the myosin purification and this enabled phosphorylation of RLC within 20-40 s after addition of calcium to the filament suspension. The phosphorylated and non-phosphorylated filaments were then partially disassembled by ultrasonification. The extent of filament disintegration was visualized and quantified by atomic force microscopy. Results: RLC phosphorylation reduced the diameter of the filaments and rendered the filaments more resistant to ultrasonic agitation. Electron microscopy revealed a similar reduction in filament diameter in intact smooth muscle when the cells were activated. Conclusion: Our results suggest that RLC phosphorylation is a key regulatory step in modifying the structural properties of myosin filaments in smooth muscle, where formation and dissolution of the filaments are required in the cells’ adaptation to different cell length.

In recent years, due to the rapid development of electronic products, non-volatile memory has become more and more important. However, flash memory has faced some physical limits bottleneck with size scaling-down. In order to overcome this problem, alternative memory technologies have been extensively investigated, including ferroelectric random access memory (FeRAM), magneto resistive RAM (MRAM), phase-change RAM (PRAM), and resistive RAM (RRAM). All of this potential next generation non-volatile memory, the resistive random access memory has most advantages such as simple structure, lower consumption of energy, lower operating voltage, high operating speed, high storage time and non-destructive access, which make it be the most potential candidate of the next generation non-volatile memory. Many studies have proposed to explain the resistance switching phenomenon, which is due to the metallic filament or the oxygen vacancies. Therefore, in order to investigate the influence of resistance switching characteristic by metal or oxygen, we choose the non-metal contained boron oxy-nitride film as the insulator layer and successfully make the resistance has the switchable characteristic of this device. Furthermore, we improved the iv stability by using the Gadolinium-doped method in the boron oxy-nitride based film. In addition, we observed the negative current differential phenomenon during the set process, which can further controlled by lower operating voltage to achieve the interfacial resistance switching. We think that is due to the formation of nitrogen titanium oxide at the interface between insulator layer and titanium nitride electrode, which caused the Schottky barrier formation and reduced the current flow. In addition, current conduction fitting can also confirm this hypothesis. Besides, titanium nitride easily bond with oxygen ions; moreover, the oxygen ions can be easily disturbed at higher temperature ambient. We believed there may easily form the nitrogen titanium oxide layer in higher temperature environment; which also improve by a series of varied temperature experiments. However, this nitrogen titanium oxide layer formed naturally very easily, resulting in an inevitable problem of data retention time, which wish to be resolved in the future.