Osteopontin (OPN) is a matricellular cytokine found in most tissues and body fluids. It is involved in a variety of cell processes by binding to integrins and CD44 receptors, and it modulates immune responses. To investigate the functions of OPN during colitis the DSS acute colitis model in OPN-/- and WT control mice was utilized. OPN-/- mice were more susceptible to DSS-induced colitis than the DSS-treated WT control mice. The increased susceptibility of the OPN-/- mice was characterized by greater intestinal crypt destruction; high myeloperoxidase activity of infiltrating neutrophils; lack of differentiation of inflammatory cells such as lymphocytes subsets (CD4+, CD8+) and macrophages (F4/80); reduced production of certain cytokines, especially TNF-alpha; and non-programmed cell death of enterocytes. It is postulated that the hyperactivity of neutrophils may explain the increased tissue destruction during experimental colitis in the absence of OPN. Analysis of OPN’s impact on neutrophil function showed that while OPN may be important for the recruitment and migration of neutrophils, the expression of OPN by neutrophils is not required for manifestation of their destructive capabilities. This would suggest that OPN administration may protect the intestines from the adverse effects of colitis. Exogenous bovine milk OPN (20 μg/ml), administered for 8 days dissolved in the drinking water, ameliorated DSS-induced colitis. It diminished signs of disease, with a greater impact in the WT than the OPN-/- mice. It reduced the levels of neutrophils, macrophages, and pro-inflammatory mediators in the colon tissue. Recombinant OPN failed to reproduce the beneficial effects of milk OPN, which suggests that post-translational modifications of OPN are crucial to ameliorate experimental colitis. Collectively, these studies demonstrate that OPN has a protective effect during experimental colitis and that the oral administration of bovine milk OPN (20 μg/ml) ameliorates acute DSS-induced colitis. The results of this study also imply that the protective effect probably depends on a post-translationally modified form of OPN, and may require intracellular-OPN as a cofactor for significant attenuation of colitis. Future research could concentrate on more detailed investigation of these latter findings to determine OPN’s mechanism of action.

In this thesis, the discharge products formed at the cathode and the performance and cell chemistry of sodium-oxygen batteries have been studied. This was carried out using different NaOTf salt concentrations. The influence of different salt concentrations on sodium-oxygen batteries was investigated since it has been shown that increasing the salt concentration beyond conventional concentrations could result in advantages such as increased stability of the electrolytes towards decomposition, higher thermal stability and lower volatility. An increase in salt concentration has also been shown to influence the electrochemical potential window. The solubility of NaOTf was investigated in two different solvents, DME and diglyme. NaOTf was found to be more soluble in DME compared to diglyme but due to the volatile nature of DME, three different concentrations of NaOTf were prepared with diglyme as solvent. Experimentation involved discharging the batteries to either maximum or limited capacity. The discharge products were examined and characterized using XRD and SEM. The main discharge product was identified as sodium superoxide although sodium peroxide dihydrate was also identified in one battery. A trend of higher capacity and voltage plateaus with higher salt concentration was also found. The influence of trace amounts of water was suggested as one explanation as it works as a catalyst, promoting sodium superoxide cube growth due to improved transportation of superoxide. Another or contributing explanation could be a possible change in donor number with increased salt concentration, resulting in higher solubility and longer lifetime of superoxide, promoting the growth of sodium superoxide cubes.

Objectives: To explore salt content in bread and to evaluate the feasibility of reducing salt contained in ‘pan francés’ bread. Design: The study had two phases. Phase 1, an exploratory phase, involved the estimation of salt contained in bread as well as a triangle taste test to establish the amount of salt to be reduced in ‘pan francés’ bread without detection by consumers. In Phase 2, a quasi-experimental, pre–post intervention study assessed the effects of the introduction of low-salt bread on bakery sales. Setting: A municipal bakery in Miraflores, Lima, Peru. Subjects: Sixty-five clients of the bakery in Phase 1 of the study; sales to usual costumers in Phase 2. Results: On average, there was 1·25 g of salt per 100 g of bread. Sixty-five consumers were enrolled in the triangle taste test: fifty-four (83·1 %) females, mean age 58·9 (SD 13·7) years. Based on taste, bread samples prepared with salt reductions of 10 % (P = 0·82) and 20 % (P =0·37) were not discernible from regular bread. The introduction of bread with 20 % of salt reduction, which contained 1 g of salt per 100 g of bread, did not change sales of ‘pan francés’ (P=0·70) or other types of bread (P =0·36). Results were consistent when using different statistical techniques. Conclusions: The introduction of bread with a 20 % reduction in salt is feasible without affecting taste or bakery sales. Results suggest that these interventions are easily implementable, with the potential to contribute to larger sodium reduction strategies impacting the population’s cardiovascular health.

The purpose of this study was to see if any soil bacteria were able to use pentachlorophenol or sodium pentachlorophenate either aerobically or anaerobically as a sole carbon source, to see if any soil bacteria could survive in high concentrations of sodium pentachlorophenate, to determine the maximum concentration of sodium pentachlorophenate which permitted the growth of some soil bacteria, to see the effects of varying concentrations of sodium pentachlorophenate on the growth curves of soil bacteria capable of growing in its presence, and to see if any soil bacteria could degrade sodium pentachlorophenate.

The emergence of pyrethroid resistant Anopheles funestus (a major African vector) in malaria affected parts of KwaZuluNatal, South Africa was correlated with the malaria epidemic of 1996 2000. This finding prompted the necessity of incorporating insecticide resistance management strategies into formal malaria control policy in South Africa. Resistance management strategies often rely on the assumption of reduced fitness associated with insecticide resistance and are based on the principle that resistance genes will tend to drift out of vector populations in the absence of insecticide selection pressure. This study aimed to determine whether a fitness cost is associated with pyrethroid resistance as well as to determine the stability and mode of inheritance of the resistance genes in a pyrethroid resistant (FUMOZR) strain of An. funestus. It also aimed to sequence and analyze a segment of the sodium channel gene for any kdrtype mutation(s) that may be associated with pyrethroid resistance. The final aim was to determine the resistance mechanisms involved in a Ghanaian field population of An. funestus resistant to DDT and pyrethroids. Results obtained suggest that pyrethroid resistance in southern African An. funestus did not incur any loss of fitness. FUMOZR had a reproductive advantage over a pyrethroid susceptible An. funestus strain (FANG) in terms of higher fertility, proportion of females laying eggs and eggtoadult survivorship, and a lower sterility Click to buy NOW! PDFXCHANGE www.docutrack. com Click to buy NOW! PDFXCHANGE www.docutrack. com iv rate. However, FUMOZR had a slower developmental time from egg hatch to adult emergence than FANG. Results of crosses and backcrosses carried out between FUMOZR and FANG were consistent with a monofactorial and autosomal mode of inheritance in which the resistant genes presented as incompletely dominant. The resistant gene was found to be stable over several generations in the absence of insecticide selection pressure. Analysis of the genomic and mRNA sequences of the IIS5 IIS6 segment of the sodium channel gene showed a high sequence identity between FUMOZR and FANG suggesting that the two strains are genetically similar. The kdrtype mutation was absent from this region supporting previous evidence that the resistance mechanism is primarily metabolic. Bioassay data showed that a Ghanaian field population of An. funestus from Obuasi, Ghana, was resistant to DDT and pyrethroids. Molecular analysis of the IIS5 IIS6 segment of the sodium channel gene showed an absence of kdrtype mutations previously associated with insecticide resistance. Biochemical analysis suggests that resistance is metabolically mediated primarily by elevated levels of and esterases with monooxygenases and GSTs playing a lesser role. The presence of an altered acetylcholinesterase conferring carbamate resistance was also evident in the population. These results have implications for the management of resistance in malaria control programmes in Africa.

Many strains of Gram-negative bacteria, such as Pseudomonas, are able to utilize a variety of unusual chemicals, including a wide range or aromatic hydrocarbons and their derivatives for growth. Bacteria with the potential to degrade sodium benzoate were isolated, identified and grown as biofilms on sodium benzoate in a laboratory-scale fluidized bed biofilm bioreactor. Four Pseudomonas strains identified as P. aeruginosa (BDS2) P. putida (BDS1 and GR1) and Burkholderia cepecia (GR3FAR) were used in a laboratory-scale FBBR together with two Bacillus strains - Bacillus macroides (SBSY4) and Bacillus simplex (MAR). Sodium benzoate biodegradation capacities of these species were compared under batch and continuous operations. Biofilm and planktonic bacterial growth dynamics were monitored by plate counts, and optical density measurements (230nm) determined benzoate biodegradation. Overall, higher attached and planktonic bacterial counts were determined under batch compared to continuous mode. In addition to this, the ability of attached cells to use sodium benzoate as their sole carbon source was compared to their suspended counterparts in a batch system. There were more attached counts compared to suspended cells and attached cells apparently degraded sodium benzoate better than planktonic cells. Similarly, higher rates of benzoate depletion were found to occur under batch compared to the continuous system. It thus appeared that more cell growth implied more substrate consumption. SEM showed attached cells and microcolonies of all the isolates on GAC, indicating their biofilmforming abilities.

Para atingir o regime de condensação de Bose-Einstein (CBE) em gases de metais alcalinos são necessárias várias etapas: feixe desacelerado, aprisionamento magneto-óptico, aprisionamento magnético e por fim o resfriamento evaporativo. Como estamos interessados em atingir o regime de CBE precisamos nos preocupar com as várias etapas intermediárias. Neste trabalho apresentaremos a construção e caracterização de uma armadilha magnética para um gás de átomos neutros de sódio. O sistema optado por nós foi o \"folha de trevo\", com o qual conseguimos a seguinte configuração de campos magnéticos: 140 gauss de campo de fundo na direção axial, 117 gauss/cm de gradiente radial e 106 gauss/cm 2 de curvatura na direção axial. Para gerarmos esta configuração de campo e desligarmos estes campos em um tempo menor que um milisegundo foi necessária a construção de um sistema de chaveamento e controle que será descrito e caracterizado neste trabalho. Com este sistema em funcionamento observamos aproximadamente 10 8 átomos aprisionados e um tempo de 1 segundo. Além disso, observamos os átomos adaptando-se a diferentes formas de potenciais de aprisionamento / In order to obtain Bose- Einstein condensation ( BEC ) in alkali gases several steps are needed : slowing beam , magneto- optical trapping , trapping magnetic and finally evaporative cooling . Since, we are interested in achieving BEC regime we need to consider about the various intermediate steps . In this work, we present the construction and characterization of a magnetic trap for a gas neutral atom of sodium. The system we chose was the \" clover leaf \" , with which we got the following configuration of magnetic fields : 140 gauss field background in the axial direction , 117 gauss / cm radial gradient and 106 gauss / cm 2 of curvature axial direction . For generating this field configuration and disconnecting these fields in a time of less than one millisecond required the construction of a switching system and control that will be described and characterized in this work. With this system, noted in about 10 8 trapped atoms and a time of 1 second. Besides, we observe the atoms adapting to different forms of potential imprisonment

by Paul Li Wai Ching. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1990. / Includes bibliographical references. / Acknowledgements --- p.i / Summary --- p.ii / Index to Figures --- p.V / Index to Tables --- p.vii / Abbreviations --- p.viii / CONTENTS / Chapter Chapter 1 --- INTRODUCTION --- p.1 / Chapter Chapter 2 --- LITERATURE REVIEW : SALT AND HYPERTENSION / Chapter 2.1. --- Summary of evidence linking salt and hypertension --- p.4 / Chapter 2.1.1. --- Epidemiological studies --- p.4 / Chapter 2.1.2. --- Dietary intervention studies --- p.7 / Chapter 2.1.3. --- Experimental studies --- p.9 / Chapter 2.2. --- Cellular sodium transport --- p.10 / Chapter 2.2.1. --- The Sodium Pump --- p.10 / Chapter 2.2.2. --- Defects in sodium transport in hypertension --- p.14 / Chapter 2.3. --- Hypothesis linking salt to the pathogenesis of hypertension --- p.15 / Chapter 2.4. --- Evidence for the presence of natriuretic Hormone --- p.18 / Chapter 2.4.1. --- Indirect evidence --- p.18 / Chapter 2.4.2. --- Direct evidence --- p.18 / Chapter 2.4.3. --- The source and properties of natriuretic hormone --- p.20 / Chapter 2.4.4. --- Other natriuretic factors --- p.21 / Chapter Chapter 3 --- REGULATION OF THE SODIUM PUMP / Chapter 3.1. --- General introduction --- p.24 / Chapter 3.2. --- Regulation of the sodium pump by intracellular sodium --- p.24 / Chapter 3.3. --- "Effects of ethanol on Na+,K+-ATPase activity" --- p.26 / Chapter 3.4. --- "Effects of potassium depletion on Na+,K+-ATPase activity" --- p.27 / Chapter 3.4.1. --- In vivo studies of sodium pump regulation by potassium --- p.27 / Chapter 3.4.2. --- In vitro studies of sodium pump regulation by potassium --- p.29 / Chapter 3.5. --- Effects of cardiac glycosides on the sodium pump --- p.30 / Chapter 3.5.1. --- In vivo studies of sodium pump regulation by cardiac glycosides --- p.31 / Chapter 3.5.2. --- In vitro studies of sodium pump regulation by cardiac glycosides --- p.33 / Chapter 3.6. --- Effects of dietary salt on the sodium pump --- p.35 / Chapter 3.6.1. --- Acute effects of salt-loading --- p.35 / Chapter 3.6.2. --- Chronic effects of salt-loading --- p.36 / Chapter Chapter 4 --- AIMS OF THE STUDY --- p.39 / Chapter Chapter 5 --- MEASUREMENT OF THE SODIUM PUMP ACTIVITY / Chapter 5.1. --- General introduction --- p.41 / Chapter 5.2. --- The measurement of sodium pump activity --- p.42 / Chapter 5.2.1. --- The sodium pump transport activity --- p.42 / Chapter 5.2.2. --- Quantitation of the number of sodium pump sites --- p.45 / Chapter 5.2.3. --- The measurement of enzyme activity --- p.47 / Chapter (a) --- Introduction --- p.47 / Chapter (b) --- Preparation of tissues and detergent treatment --- p.48 / Chapter (c) --- Measurement of ATPase activity by measuring the rate of release of inorganic phosphate --- p.49 / Chapter (d) --- The coupled-enzyme assay --- p.53 / Chapter (e) --- The K+-stimulated 3-0-MFPase assay --- p.54 / Chapter Chapter 6 --- METHODS - ESTABLISHMENT AND EVALUATION / Chapter 6.1. --- Chemicals --- p.57 / Chapter 6.2. --- "Measurement of Na+,K+-ATPase activity by the rate of release of inorganic phosphate" --- p.58 / Chapter 6.3. --- "Automated coupled-enzyme assay of Na+,K+-ATPase activity" --- p.62 / Chapter 6.4. --- "The measurement of Na+,K+-ATPase activity by the potassium-stimulated 3-0-MFPase assay" --- p.67 / Chapter 6.5. --- Determination of protein concentration --- p.70 / Chapter 6.6. --- Statistical analysis --- p.73 / Chapter 6.7. --- Results --- p.73 / Chapter 6.7.1. --- Evaluation of the inorganic phosphate release method --- p.73 / Chapter 6.7.2. --- Evaluation of the coupled-enzyme method --- p.78 / Chapter 6.7.3. --- Evaluation of the K+-stimulated 3-0-MFPase method --- p.89 / Chapter 6.7.4. --- Evaluation of the protein determination method --- p.94 / Chapter 6.8. --- Discussion --- p.96 / Chapter Chapter 7 --- "THE EFFECTS OF DIGOXIN TREATMENT ON Na+,K+-ATPase ACTIVITY OF DIFFERENT TISSUES" / Chapter 7.1. --- Introduction --- p.101 / Chapter 7.2. --- Materials and Methods --- p.103 / Chapter 7.2.1. --- Animals and diets --- p.103 / Chapter 7.2.2. --- Drugs --- p.103 / Chapter 7.2.3. --- Pharmacokinetics of digoxin --- p.103 / Chapter 7.2.4. --- The digoxin regimes --- p.104 / Chapter 7.2.5. --- Preparation and deoxycholate treatment of tissue homogenates --- p.105 / Chapter 7.2.6. --- "Measurement of Na+,K+-ATPase activity" --- p.107 / Chapter 7.2.7. --- Digoxin radioimmunoassay --- p.107 / Chapter 7.2.8. --- Measurement of plasma electrolytes --- p.109 / Chapter 7.3. --- Results --- p.110 / Chapter 7.3.1. --- The pharmacokinetics of digoxin in the rat --- p.110 / Chapter 7.3.2. --- Plasma digoxin levels --- p.110 / Chapter 7.3.3. --- Effects of digoxin treatment on body weight --- p.113 / Chapter 7.3.4. --- Effects of digoxin treatment on plasma electrolytes --- p.117 / Chapter 7.3.5. --- "Effects of digoxin treatment on tissue Na+,K+-ATPase activity" --- p.119 / Chapter 7.4. --- Discussion --- p.129 / Chapter Chapter 8 --- THE SALT-LOADING EXPERIMENT / Chapter 8.1. --- Introduction --- p.140 / Chapter 8.2. --- Materials and Methods --- p.142 / Chapter 8.2.1. --- Animals --- p.142 / Chapter 8.2.2. --- The salt-loading protocol --- p.142 / Chapter 8.2.3. --- Preparation of crude tissue homogenates --- p.143 / Chapter 8.2.4. --- "Measurement of Na+,K+-ATPase activity" --- p.144 / Chapter 8.2.5. --- Analysis of urinary electrolytes --- p.144 / Chapter 8.2.6. --- Measurements of body weight and wet weight of kidney --- p.145 / Chapter 8.3. --- Results --- p.145 / Chapter 8.3.1. --- Effects of salt loading on the body weight --- p.145 / Chapter 8.3.2. --- Effects of salt loading on 24-hour urinary sodium excretion --- p.148 / Chapter 8.3.3. --- Effects of salt loading on the wet weight of kidney --- p.152 / Chapter 8.3.4. --- "Effects of salt loading on tissue Na+,K+-ATPase activity" --- p.152 / Chapter 8.4. --- Discussion --- p.163 / Chapter Chapter 9 --- CONCLUSIONS AND FUTURE WORK --- p.175 / REFERENCES --- p.183

Genetic mutations perturbing inactivation of human cardiac voltage-gated sodium channels (VGSCs), specifically Nav1.5, can cause long QT syndrome type 3 (LQT3). LQT3 is a cardiac disorder in which patients experience syncope and ventricular tachyarrhythmia, and are thus predisposed to sudden cardiac death. Deeper understanding of the structural dynamics of VGSC inactivation is needed to inform treatment of and drug design for potentially life-threatening arrhythmias. A well supported hypothesis is that the VGSC inactivated state is stabilized by hydrophobic interactions between the inactivation gate and an unknown binding site potentially involving the underside of the channel pore, C-terminus (C-T), and auxiliary proteins. Despite advances in biophysical and structural characterization of VGSCs, the specific molecular components and timing of their interactions within the inactivation complex remain unclear. Fluorescence imaging approaches that connect conformational change with channel function in mammalian cells could provide much needed mechanistic insight on the structural dynamics of the VGSC inactivation complex. This thesis describes the development of a site-specific fluorescent unnatural amino acid (UAA) labeling and spectral imaging methodology to probe the cardiac VGSC, Nav1.5, inactivation complex in live mammalian cells. First, UAA mutagenesis experiments were performed to validate orthogonal synthetase-tRNA (aaRS-tRNA) technology for fluorescent labeling of intracellular and membrane proteins in mammalian cells. Next, towards investigating conformational dynamics and intramolecular interactions related to inactivation, the Nav1.5 inactivation gate was labeled with a single environmentally sensitive fluorescent UAA L-anap. While the function of L-anap labeled channels was altered, their function remained within pathophysiological range. Then, imaging of L-anap labeled Nav1.5 in mammalian cells afforded characterization of unique L-anap spectra at different sites in the inactivation gate. Finally, using potassium-depolarization (K-depolarization) as rough means of voltage control, L-anap spectral shifts demonstrated conformational changes between the closed and open-inactivated states, which depended on the presence of the distal C-T (DCT). Site-specific L-anap labeling of the inactivation gate combined with spectral imaging and K-depolarization affords a general imaging assay to directly monitor conformational rearrangements of the Nav1.5 inactivation gate in channels expressed in live mammalian cells. While interactions with the DCT are specifically probed, this general assay provides an opportunity to bring necessary unification of ideas about VGSC inactivation, as well as insight on outstanding questions of VGSC regulation.

Proper functioning of the voltage gated sodium channel, NaV1.5, is essential for maintenance of normal cardiac electrophysiological properties. Changes to the biophysical properties of sodium channels can take many forms and can affect the peak component of current carried during phase zero of the action potential; the “persistent” or “late” current component conducted during the repolarizing phases of the action potential; the availability of the channel as seen by changes in window current; and the kinetics of channel transitions between closed, opened and inactivated states. Mutations in NaV1.5 that alter these parameters of channel function are linked to a number of cardiac diseases including arrhythmias such as atrial fibrillation. In addition, mutations in many of the auxiliary proteins that form part of the sodium channel macromolecular complex have likewise been associated with diseases of the heart. Mutations in regions of the sodium channel responsible for interactions with these auxiliary proteins have also been linked to various dysfunctional cardiac states. Indeed, a large number of disease causing mutations are localized to the C-terminal domain of NaV1.5, a hotspot for interacting proteins. Using a transgenic mouse model, we show that expression of a mutant sodium channel with gain-of-function properties conferring increased persistent current, is sufficient to cause both structural and electrophysiological abnormalities in the heart driving the development of spontaneous and prolonged episodes of atrial fibrillation. The sustained and spontaneous atrial arrhythmias, an unusual if not unique phenotype in mice, enabled explorations of mechanisms of atrial fibrillation using in vivo (telemetry), ex vivo (optical voltage mapping), and in vitro (cellular electrophysiology) techniques. Since persistent sodium current was the driver of the structural and electrophysiological abnormalities leading to atrial fibrillation, we subsequently pursued studies exploring the mechanisms of persistent sodium current. Prior work of heterologously expressed sodium channels identified calmodulin as a regulator of persistent current. Mutation of the calmodulin binding site in the C-terminus of the cardiac sodium channel caused increased persistent current when the channel was expressed heterologously. The role of calmodulin in the regulation of the sodium channel in cardiomyocytes has not been definitively determined. We created transgenic mice expressing human sodium channels harboring a mutation of the calmodulin binding site. Using whole cell patch clamping, we found, in contrast to previously reported findings, that ablation of the calmodulin binding site did not induce increased persistent sodium current. Instead, loss of calmodulin binding stabilized the inactivated state by shifting the V50 for steady-state inactivation in the hyperpolarizing direction. Furthermore, loss of calmodulin binding sped up the transition to the inactivated state demonstrated by a significantly shortened tau of inactivation. In contrast to studies performed in heterologous expression systems, our findings thus suggest that in heart cells, calmodulin binding increases availability, similar to its role in regulating NaV1.4 channels. The studies were then expanded to explore the role of other interacting proteins, fibroblast growth factor (FGF) homologous factors (FHF), in the presence and absence of calmodulin binding. Using whole cell patch clamping, we found that a mutation (H1849R) of the sodium channel causing decreased FHF binding affinity leads to a rightward shift in steady-state inactivation and a slowed rate of inactivation of INa. A third mutant channel, with concurrent decreased FHF and calmodulin binding affinity similarly results in a rightward shift in steady-state inactivation suggesting a dominant effect of the H1849R mutation. Persistent current was not elevated in either of these mutant channels. Importantly, the methodology that we report enables us and other groups to carry out studies of human sodium channels in the native environment of NaV1.5. Our investigation into calmodulin’s role, which yielded conclusions distinct from prior findings in heterologous expression systems, demonstrates the value of this approach.