<p>S-nitrosylation, the prototypic mechanism of redox-based signal transduction, involves the covalent attachment of a nitrogen monoxide group to a Cys-thiol side chain. S-nitrosylation of proteins has been demonstrated to affect a broad range of functional parameters including enzymatic activity, subcellular localization, protein-protein interactions and protein stability. The primary focus of my dissertation was to solve a problem of great importance in the field of S-nitrosylation, which is, to identify denitrosylase(s) i.e., enzymes that remove NO groups from S-nitrosothiols. Recent progress in elucidating the cellular regulation of S-nitrosylation has led to the identification of two physiologically relevant denitrosylating activities that remove the NO group from S-nitrosylated substrates. Thioredoxin/thioredoxin reductase (Trx system) functions as an NADPH-dependent denitrosylase across a broad range of S-nitrosylated proteins (SNO-proteins). S-nitroso-glutathione reductase (GSNOR), which is highly conserved across phylogeny, metabolizes GSNO utilizing NADH as a reducing coenzyme, thereby shifting equilibria between GSNO and SNO-proteins. This dissertation describes the discovery of two novel denitrosylases: one from yeast and the other from mammals. Using technique of column chromatography we have purified these novel denitrosylases to homogeneity and have demonstrated a principal contribution of these enzymes towards S-nitrosothiol metabolism.</p> / Dissertation

Made available in DSpace on 2014-06-11T19:27:12Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-12-21Bitstream added on 2014-06-13T20:08:18Z : No. of bitstreams: 1 gasparotto_g_me_araiq.pdf: 3559380 bytes, checksum: fb3a7ae9b231ef8abae43cc0439f427f (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O presente trabalho consiste no estudo da viabilidade para a obtenção de cerâmicas densas de SnO2 com baixa resistividades, dopados com Zn2+, Nb5+ e Al3+. Os pós cerâmicos foram preparados pelo método químico (método Pechini) e por de mistura de óxidos, usando a sinterização em forno microondas. Fez-se a caracterização do pó por difratometria de raios X (DRX), medidas de área superficial (BET) e termogravimetria e termogravimetria diferencial (TG/DTA). Depois de conformar os pós, fez-se a caracterização microestrutral e elétrica dos compactos. A baixa densificação do material é justificada pela presença de microestruturas em formas de agulhas, constituída pelo agente densificante. Os menores resultados de resistividade à temperatura ambiente foram obtidos para as composições SZ100N e S200ZNA, obtidos pelo método Pechini, cujos valores são, respectivamente, 12,4 e 11,3 Ω.cm. Quando aumentou a temperatura de 50º a 400ºC, os menores resultados de resistividade foram obtidos pelo material preparado por mistura de óxidos com valor próximo a 5 Ω.cm / The present work is to study the viability to obtain high densification of SnO2 with low resistivity doped Zn2+, Nb5+ and Al3+. The ceramic powders were prepared by chemical method (Pechini method) and by solid state reaction, using the microwave sintering. There was the characterization by powder X-ray diffraction (XRD), surface area measurements (BET) and thermogravimetry and differential thermogravimetry (TG/DTA). After conforming the powder, it was the characterization of electrical and microestrutral compact. The low densification of the material is justified by the presence of microstructures in forms of needles constituted of the densifying agent. The lowest specific resistivity at room temperature were obtained for the compositions and S200ZNA SZ100N, obtained by Pechini method, whose values are respectively 12.4 and 11.3 Ω.cm. When the temperature increased from 50 º to 400 º C, the lowest resistivity results were obtained for material prepared by mixing oxides with a value next 5 Ω.cm

The aim of the research presented in this thesis is to investigate and develop novel nitric oxide (NO) delivery systems, specifically designed for application in medical areas. The initial work has focused on utilising metal organic frameworks (MOFs) as a delivery system for this radical gas, NO. Due to their high porosity, high thermal stability and the presence of coordinated unsaturated metal sites (CUSs) when fully activated, the CPO-27 (Coordination Polymer of Oslo) family of MOFs has been selected as a suitable host framework. CPO-27 (Ni), CPO-27 (Mg) and CPO-27 (Zn) have been prepared using reflux and room temperature processes without recourse to the use of any toxic or harmful solvents. The resulting products are characterised by powder XRD (X-ray diffraction) and SEM (Scanning electron microscopy), and their NO adsorption, storage and release properties are reported. The results indicate that the crystallinity, particle size and NO adsorption, storage and release performance are comparable to those of equivalent samples synthesised via traditional solvothermal methods, paving the way for a more easily scalable and environmentally friendly synthetic procedure for these types of MOF. Depending on which metal is employed; the NO uptake, storage and release varies, the more toxic nickel based framework shows enhanced performance in terms of concentration and duration of NO released against either the magnesium or zinc counterparts. In order therefore to reduce the risk of toxicity whilst retaining good performance, Ni (II) ions were doped into the 3D framework of CPO-27 (Mg) and CPO-27 (Zn) using novel water-based reflux and room temperature crystallization methods. Several characterization techniques strongly support the effective incorporation of Ni (II) ions into the 3D framework. Nitric oxide (NO) adsorption/release data, as well as in vitro tests demonstrate that NO dosage and biological response can be tuned via the Ni doping process allowing enhanced performance without the high toxicity of pure Ni MOFs. Such materials would be extremely advantageous and more applicable for use in medical fields. NONOates and other NO-complexes have also been investigated as alternative NO delivery systems. This study has focused on developing NO-drug complexes using a variety of different compounds commonly used by clinicians, namely the antiseptic (chlorhexidine, CHx), the antibiotic (ciprofloxacin) and diuretic (furosemide). A unique high pressure NO loading methodology has been developed to coordinate nitric oxide to these drug molecules and their NO release performance has been evaluated. The resulting NO-drug complexes are characterised using a series of spectroscopic techniques and the collected data highlights that the radical gas coordinates with the secondary amine groups present in the drug molecules. The interaction between the amine group and the gas is reversible; in fact the release of NO from these complexes can be triggered using water (11% RH) and/or UV-light. In addition, chlorhexidine has been incorporated into the pores of the CPO-27 framework. The amount of antiseptic incorporated was determined using a variety of characterisation techniques. The controlled release of significant concentrations of CHx from the CPO-27 materials are achieved by exposing each CHx loaded sample to a water solution, in doing so topical conditions are simulated. The CHx loaded samples have also been activated and NO loaded following the novel high pressure procedure specifically developed during this research. The resulting NO loaded material released the radical gas in the presence of water and/or UV-light. By incorporating the CHx into the MOF and NO loading this complex the duration and release of NO was greatly enhanced over that of either of the components alone. On formulating the CHx loaded 3D frameworks into pellets, or even into a polyurethane polymer film, their ability to release the antiseptic under simulated topical conditions was maintained. The NO-CHx-CPO-27 composite film that has been prepared has proven to be able to simultaneously store and release both NO and CHx. Each component of the complex has more than one function and the quantity and duration of release of NO is again higher and longer than either of the two moieties alone. The release of these two antibacterial agents from a MOF is novel and is very exciting as it opens up the possibility of engineering products with multiple actions to fight infection. Owing to their high stability and shape persistence properties, the CC3 cage series (CC3, RCC3, FT-RCC3 and AT-RCC3) was chosen as the basis of an investigation into the potential use of porous organic cages as delivery systems for nitric oxide gas. NO has been stored in these porous materials through coordination to amine groups forming Nnitrosamine groups. Release of NO from these types of compounds can be triggered by various mechanisms including water and UV-light, the amine group being regenerated after the release of NO. The release performance significantly increased when the materials were exposed to UV-light and/or suspended in water. As a result of this investigation, these covalent organic molecular cages can now be added to the existing list of NO-based therapies available to medical professionals.

In this dissertation, density functional theory (DFT) calculations were used to investigate (1)NO2 adsorption on BaO in NOx Storage Reduction (NSR) catalyst affected by the morphology of BaO and the γ-Al2O3 support, (2) energy barrier of H2 dissociative adsorption over Mg clusters affected by its electronic structure, and (3) comparison of the activities of CeO2 clusters affected by two different supports--monoclinic ZrO2 and non-spinel γ-Al2O3. Our results showed that the electronic effect caused by the non-stoichiometry of the bare BaO clusters and surfaces improves their reactivities toward NO2 adsorption greatly, whereas the geometric structure of the catalyst has only minor effect on the activity; we also found that the γ-Al2O3 substrate improves the reactivities of the supported BaO clusters and at the same time the interface between BaO and γ-Al2O3 provided a unique and highly reactive environment for NO2 adsorption. Hydrogen dissociation barrier over pure Mg clusters is greatly affected by the electronic structure of the clusters--closed shell clusters such as Mg10 and Mg92- have higher energy barrier toward H2 dissociation; however, H2 dissociation over clusters that are two electrons shy from the closed electronic shell are relatively easier. As substrates, neither ZrO2(111) nor γ-Al2O3(100) affects the reactivity of the supported Ce2O4 toward CO2 adsorption and CO physisorption significantly; whereas the reactivity of Ce2O4 toward CO reactive adsorption were found to be affected by the two substrates very differently.

Three systems of electrospun composite membranes with piezoelectric polymer polyvinylidene fluoride (PVDF) as matrix incorporating: 1) Carbon based fillers: carbon nanotube (CNT) and graphene oxide (GO); 2) Ceramic based fillers-barium titanate (BT), zinc oxide (ZnO) and nanoclays (halloysite and bentonite); 3) Cellulosic fillers: microcrystalline cellulose (MCC) and nanocrystalline cellulose (NCC) at different loadings were prepared by electrospinning process. Influence of filler type and loading on total PVDF crystallinity (Xc), relative fraction of β phase (piezoelectric phase) in total crystalline PVDF (Fβ), volume fraction of β phase in the samples (vβ) and piezoelectric coefficient d33 were characterised and analysed. Correlation between vβ and piezoelectric performance (d33) will be focused by this work. A common situation was observed for the composites-d33 increased while vβ is reduced by the fillers, so it can be concluded that d33 of the composites is not totally up to their vβ, there are other factors that need to be taken into account. For example, for carbon based filler like CNT, it increased electric conductivity of sample during and after electrospinning process, making it easier for charges produced by β crystals from inside of sample to be transferred to surfaces of the sample, and possibly promoting orientation of β crystals in d33 direction, therefore enhanced d33 of the composites though β phase formation was significantly hindered by inclusion of CNT; For piezoelectric ceramic fillers like BT and ZnO, a possible combined piezoelectricity from filler and β phase PVDF might enhanced d33 though less β phase was formed; And for non-piezoelectric and non-conductive fillers, enhancement in orientation of β crystals might play a major role in promotion of d33. Keywords: electrospinning; polyvinylidene fluoride (PVDF); nanocomposites; piezoelectric coefficient d33; energy harvesting.

Abstract The aim of the study was to evaluate pulmonary endogenous and inhaled nitric oxide (NO) in neonates with severe respiratory failure. Infant autopsy documents were reviewed for fulminant early-onset bacterial pneumonia. 12 infants with the onset at &lt; 72 h of age and three control groups were identified. Immunohistochemistry revealed that 11 of the infants with early-onset pneumonia (92%) had no or faint inducible nitric oxide synthase (NOS2) staining in their alveolar macrophages (AM). All control infants, regardless of their postnatal age, had NOS2-positive AM. The marker of NO-toxicity, nitrotyrosine, was low in all specimens. To confirm this finding, airway specimens of 21 newborns requiring mechanical ventilation were examined. Seven of them had fulminant early-onset pneumonia with maternal ascending intra-uterine infection (IUI). The controls had no infection at birth despite IUI or neither infection nor IUI. In early-onset pneumonia, NOS2 and nitrotyrosine immunoreactivity were low at birth and increased during the recovery phase (p &lt; 0.05). Analyses of interleukin-1 and surfactant protein A showed the same pattern of age-dependent change. Of the autopsied infants, 12 had received inhaled NO (iNO) before death. Each case was paired with a matched control. Additional five infants without respiratory failure prior to death were also studied. The iNO-treated ones tended to have more intensive NOS2 staining in the bronchiolar epithelium and adjacent tissue than the controls. No differences in other NOS isoforms or nitrotyrosine were detected. A novel method for exhaled NO measurements of intubated infants was developed. Six preterm and six term newborns were prospectively recruited for expired and nasal NO measurements. During the first week of life, the preterm infants showed a different pattern of exhaled NO excretion compared to the term infants. For the pilot intervention study on very early iNO, the eligible patients had a birth weight &lt; 1500 g and progressive, therapy-resistant respiratory failure before five hours of age. Five infants received iNO, showed immediately improved oxygenation and survived without deleterious side effects. Deficient production of NO in small premature infants is associated with severe infection and respiratory failure. Very early iNO therapy may be exceptionally effective in a select group of infants, and did not appear to cause oxidation lung injury.

Magister Scientiae - MSc / It is well known that a lead pencil is made of graphite, a naturally form of carbon, this is important but not very exciting. The exciting part is that graphite contains stacked layers of graphene and each and every layer is one atom thick. Scientists believed that these graphene layers could not be isolated from graphite because they were thought to be thermodynamically unstable on their own and taking them out from the parent graphite crystal will lead them to collapse and not forming a layer. The question arose, how thin one could make graphite. Two scientists from University of Manchester answered this question by peeling layers from a graphite crystal by using sticky tape and then rubbing them onto a silicon dioxide surface. They managed to isolate just one atom thick layer from graphite for the first time using a method called micromechanical cleavage or scotch tape. In this thesis chemical method also known as Hummers method has been used to fabricate graphene oxide (GO) and reduced graphene oxide. GO was synthesized through the oxidation of graphite to graphene oxide in the presence of concentrated sulphuric acid, hydrochloric acid and potassium permanganate. A strong reducing agent known as hydrazine hydrate has also been used to reduce GO to rGO by removing oxygen functional groups, but unfortunately not all oxygen functional groups have been removed, that is why the final product is named rGO. GO and rGO solutions were then deposited on silicon substrates separately. Several characterization techniques in this work have been used to investigate the optical properties, the morphology, crystallography and vibrational properties of GO and rGO.

Metal oxides exhibit a wide range of chemical and electronic properties, making them an extremely interesting subject for numerous applications in modern electronics. The primary goal of this research is to develop metal-oxide-based electronic devices, including thin-film transistors (TFTs), resistance random-access memory (RRAM) and planar nano-devices. This research requires different processing techniques, novel device design concepts and optimisation of materials and devices. The first experiments were carried out to optimise the properties of zinc oxide (ZnO) semiconductors, in particular the carrier concentration, which determines the threshold voltage of the TFTs. Thermal annealing is one common method to affect carrier concentration and most work in the literature reports performing this process in a single-gas environment. In this work, however, annealing was carried out in a combination of air and nitrogen, and it was found that the threshold voltage could be tuned over a wide range of pre-determined values.Further experiments were undertaken to enhance the carrier mobility of ZnO TFTs, which is the most important material quality parameter. By optimising deposition conditions and incorporating a high-k gate dielectric layer, the devices showed saturation mobility values over 50 cm2/Vs at a low operating voltage of 4 V. This is, to our knowledge, one of the highest field-effect mobility values achieved in ZnO-based TFTs by room temperature sputtering. As an important type of metal-oxide-based novel memory devices, which have been studied intensively in the last few years, RRAM devices were also explored. New materials, such as tin oxide (SnOx), were tested, exhibiting bipolar-switching operations and a relatively large resistance ratio. As a novel process variation, anodisation was performed, which yielded less impressive results than SnOx, but with a potential for ultra-low-cost manufacturing. Finally, novel planar nano-devices were explored, which have much simpler structures than conventional multi-layered transistors and diodes. Three types of ZnO-based nano-devices (a side-gated transistor, a self-switching diode and a planar inverter) were fabricated using both e-beam lithography and chemical wet etching. After optimisation of the challenging wet etching procedure at nanometre scale, ZnO nano-devices with good reproducibility and reliability have been demonstrated.

Current and potential distribution in electrolyte cells. A cell has been built which has two Luggin capillaries mounted on micrometer drives so that the current and potential distribution around a working electrode can be measured. The current and potential distributions for a variety of electrode geometries have been measured. Experiments have been performed which show the effect of electrode geometry, current density and electrolyte conductivity on the current and potential distribution. Experimentally measured current distributions are compared with those calculated from existing theory.^8 Kinetic measurements of the electrolytic formation of argentous oxides in potassium hydroxide. Cyclic current step measurements during the Ag-Ag_2O oxidation plateau have been made. The results of measurements are very similar to those obtained from systems (Ag-Ag(NH_3)_2^+, (Fe^+2-Fe^+3) where the concentration of the electroactive species is diffusion controlled. The exchange current density, i_0, calculated from the relationship developed by Wijnen and Smit^49 decreased from 1.44 ma/cm^2 to 0.75 ma/cm^2 along the Ag-Ag_2O plateau. The changes in the ohmic resistance of the silver oxide film formed in 0.0543, 0.109, 1.09, 5.43, and 10.9 F KOH have been measured. The maximum resistance of the oxide measured in these electrolytes was 0.31, 0.28, 2.44, 5.15, and 0.16ohm-cm^2. Evidence for a proposed explanation of the changes which occurs in the oxide film resistance is presented.

Catecholamines are elaborated in stress responses to mediate vasoconstriction, and elevate systemic vascular resistance and blood pressure. They are elaborated in disorders such as sepsis, cocaine abuse, and cardiovascular disease. The aim of the study was to determine whether catecholamines affect nitric oxide (NO) production, as NO is a vasodilator and counteracts the harmful effects of catecholamines. RAW264.7 macrophage cells were cultured with lipopolysaccharide (LPS)±epinephrine, norepinephrine, and dopamine at 5×10-6M concentrations for 24h. Supernatants were harvested for measuring NO by spectrophotometry using the Greiss reagent and cells were harvested for detecting inducible NO synthase (iNOS) by Western blot. NO production in RAW 264.7 macrophages was increased significantly by addition of LPS (0.5-10ng/ml) in a dose-dependent fashion. The NO production induced by LPS was further enhanced by epinephrine and norepinephrine, and to a lesser extent by dopamine. These increases in NO correlated with expression of iNOS protein in these cells. The enhancing effect of iNOS synthesis by epinephrine and norepinephrine on LPS-induced macrophages was down regulated by β-adrenoceptor antagonist, propranolol, and dexamethasone. The results suggest that catecholamines have a synergic effect on LPS in induction of iNOS synthesis and NO production, and this may mediate some of the vascular effects of infection. These data support a novel role for catecholamines in disorders such as septic shock and cocaine use, and indicate that β-adrenoceptor antagonists and glucocorticoids may be used therapeutically for modulation of the catecholamine-NO axis in disease states.