Paediatric ependymomas are the third most common childhood brain cancer and represent a prognostic and therapeutic challenge. Previous evidence suggests that telomerase, a ribonucleoprotein critical in permitting limitless growth potential, may serve as both a prognostic marker and therapeutic target. Immunohistochemical analysis (n=198) and enzymatic detection (n=25) of telomerase was performed to determine prevalence and prognostic potential. The telomerase inhibitor Imetelstat was used to study telomerase inhibition in paediatric ependymoma cell lines, tumour initiating cells (TICs) and both subcutaneous and intracranial xenografts. Telomerase activity was detected in 76% of primary ependymomas and was associated with a reduced five-year progression-free survival (30% vs 75%). Telomerase inhibition in vitro resulted in shortened telomeres, increased senescence, growth inhibition and reduced self-renewal capacity. In vivo, Imetelstat shortened telomeres and reduced subcutaneous tumour volume by 40% compared to control mice. Therefore, telomerase may serve as an ideal prognostic marker and therapeutic target in paediatric ependymoma.

The most common evidence for inhibition of return (IOR) is the robust finding of increased response times to targets that appear at previously cued locations following a cue-target interval exceeding ~ 300 ms. In a variation on this paradigm, Abrams and Dobkin (1994a) observed that IOR was greater when a saccadic response was made to a peripheral than to a central arrow, leading to the conclusion that saccadic responses to peripheral targets comprise motoric and perceptual components (the two components theory for IOR) whereas saccadic responses to a central target comprise a single motoric component. In contrast to the foregoing findings, Taylor and Klein (2000) discovered that IOR for saccadic responses was equivalent for randomly intermixed central and peripheral targets, suggesting a single motoric flavor under these conditions. To resolve the apparent discrepancy, a strict replication of Abrams and Dobkin was conducted in which central and peripheral targets were either blocked or mixed. In the blocked design, peripheral targets resulted in more IOR than central targets, while in the mixed design, replicating Taylor and Klein (2000), target type had no bearing on the magnitude of IOR (i.e., equivalent IOR was obtained for both target types). This pattern of results suggests that the confound inherent in Abrams and Dobkin's blocked design generated a pattern of results that "masqueraded" as two components of IOR.

Prompted by oculomotor theories of attention, the present experiments explore the role of saccade activation in the generation of two cueing effects: exogenous capture (Experiment 1) and inhibition of return (IOR; Experiment 2). Exogenous capture is shortlived and marked by faster responding toward recently stimulated locations, whereas the longer-lasting IOR manifests as slower responding toward those locations. Within each experiment, Group A performed in a dual-task in which on most trials a peripheral target had to be identified but infrequently a central arrow probe called for an eye movement instead, while for Group B the tasks were the same except saccade trials were frequent and target identification trials were infrequent. In Experiment 1, for group A uninformative cues captured attention as measured by faster digit identification at the cued location, an effect not accompanied by saccade activation. For group B, cues generated saccade activation without capturing attention. Thus saccade activation need not accompany exogenous covert capture, and covert capture need not accompany saccade activation. In Experiment 2, group A exhibited IOR which slowed digit identification, but did not affect saccadic responding, while Group B exhibited no IOR in either digit identification or eye movement trials. This finding provides converging evidence that IOR can be dichotomized into two forms; one which delays motor production itself (Evidenced amply elsewhere, e.g., Taylor & Klein, 2000) and another which delays responding by applying inhibition at a perceptual-motor interface which can operate in independence from its motoric cousin.

Executive function, the cognitive processes that allow the voluntary control of goal-directed behaviour, can be studied through the examination of inhibition of action. The countermanding paradigm has been shown to be a powerful tool to examine a subject’s ability to withhold responses to a go stimulus when a stop signal is presented occasionally. Logan and Cowan (1984) developed a race model to account for countermanding performance in humans, proposing that independent go and stop process initiated by the go and stop signals respectively, race toward a finish line whereby the first process to cross its finish line determines the behavioural outcome. The model allows estimation of the stop signal response time, a variable that is not directly observable. The race model has yet to be validated for countermanding performance in rats. Using a new rodent countermanding task inspired directly from human studies, male Wistar rats were trained to respond to a visual stimulus (go signal) by pressing a lever below an illuminated light for food reward, but to countermand lever the press (25% of trials) subsequent to an auditory tone (stop signal) presented after a variable delay. The ability to cancel a response decreased as stop signal delay increased. The stop signal response time for rats was estimated to be 157 ms, a value within the range of human estimates. Predictions of countermanding performance made by the race model were generally respected. Response times of movements that escape inhibition: 1) were shorter than those of movements made in the absence of a stop signal; 2) gradually lengthened with increasing stop signal delay; and most importantly, 3) were predicted by the race model. These findings demonstrate that the countermanding performance of rodents can be accounted for by a simple race model, which has been applied successfully in human studies and nonhuman primate models. This new animal model will permit complementary invasive investigations of brain mechanisms underlying inhibitory control and refine the existing rodent models of neurological disease and impulsivity. / Thesis (Master, Neuroscience Studies) -- Queen's University, 2008-09-30 11:35:44.713

Paediatric ependymomas are the third most common childhood brain cancer and represent a prognostic and therapeutic challenge. Previous evidence suggests that telomerase, a ribonucleoprotein critical in permitting limitless growth potential, may serve as both a prognostic marker and therapeutic target. Immunohistochemical analysis (n=198) and enzymatic detection (n=25) of telomerase was performed to determine prevalence and prognostic potential. The telomerase inhibitor Imetelstat was used to study telomerase inhibition in paediatric ependymoma cell lines, tumour initiating cells (TICs) and both subcutaneous and intracranial xenografts. Telomerase activity was detected in 76% of primary ependymomas and was associated with a reduced five-year progression-free survival (30% vs 75%). Telomerase inhibition in vitro resulted in shortened telomeres, increased senescence, growth inhibition and reduced self-renewal capacity. In vivo, Imetelstat shortened telomeres and reduced subcutaneous tumour volume by 40% compared to control mice. Therefore, telomerase may serve as an ideal prognostic marker and therapeutic target in paediatric ependymoma.

It has been a practice to use seawater for preparing acid in offshore operations where fresh water is relatively expensive or logistically impossible to use. However, hydrochloric acid will release calcium ion into solution, which will combine with sulfate ion in seawater (greater than 3000 ppm) and calcium sulfate will precipitate once it exceeds its critical scaling tendency. A few studies have provided evidence for this problem and how to address this problem has not been fully examined. Core flood tests were conducted using Austin Chalks cores (1.5 in. x 6 in. and 1.5 in. x 20 in.) with permeability 5 md to investigate the effectiveness of scale inhibitor. A synthetic seawater was prepared according to the composition of seawater in the Arabian Gulf. Calcium, sulfate ions, and scale inhibitor concentrations were analyzed in the core effluent samples. Solids collected in the core effluent samples were analyzed using X-ray photoelectron spectroscopy (XPS) technique and thermodynamic calculation using OLI Analyzer software were conducted to identify the critical scaling tendency of calcium sulfate at different temperatures. Results showed that calcium sulfate precipitation occurred when seawater was used in any stage during matrix acidizing including preflush, post-flush, or in the main stage. Injection rate was the most important parameter that affected calcium sulfate precipitation; permeability reduction was significant at low flow rates, while at high rates wormhole breakthrough reduced the severity of the problem. More CaSO4 precipitated at high temperatures, accounting for more significant permeability reduction in the cores. The values of critical scaling tendency at various temperatures calculated by OLI ScaleChem 4.0.3 were believed to be 2.1, 2.0, and 1.2 respectively. A scale inhibitor (a sulfonated terpolymer) was found to be compatible with hydrochloric acid systems and can tolerate high concentration of calcium (30,000 mg/l). Analysis of core effluent indicated that the new treatment successfully eliminated calcium sulfate scale deposition. The concentration of scale inhibitor ranged from 20 to 250 ppm, depending on the scaling tendencies of calcium sulfate. This work confirms the damaging effect of preparing hydrochloric acid solutions using seawater on the permeability of carbonate cores. Therefore, it is recommended to use fresh water instead of seawater to prepare HCl acids whenever possible. If fresh water is not available, then a proper scale inhibitor should be added to the acids to avoid calcium sulfate precipitation.

The enzyme 3-deoxy D-arabino-heptulosonate 7-phosphate (DAH7P) synthase catalyses the first step of the shikimate pathway. This pathway lies at the heart of bacterial metabolism, and is responsible for the synthesis of a variety of compounds essential to the chemistry of life; from the aromatic amino acids phenylalanine, tyrosine and tryptophan, to a number of aromatic and non-aromatic natural products. This thesis describes the design, synthesis and evaluation of inhibitors of DAH7P synthase. These inhibitors exploit a variety of strategies to interrupt the activity of DAH7P synthase, ranging from simple substrate mimicry to inhibitors that mimic unstable reaction intermediates; inhibitors that exploit metal coordination and entropic effects, and inhibitors that gain improved potency by interacting with multiple sites. In Chapter Two, the synthesis of a mimic for a proposed unstable reaction intermediate is described, and its interaction with DAH7P synthase characterised. The compound was prepared in twelve steps from D-arabinose, and was found to be a slow-tight binding inhibitor of Escherichia coli DAH7P synthase. In Chapter Three, a number of compounds are prepared that were designed to bind to the phosphoenolpyruvate subsite of the DAH7P synthase active site. The binding of these compounds to the enzyme is investigated in order to gain an understanding of the factors involved in DAH7P synthase inhibition. The enantiomeric phospholactates were prepared, and the extent of inhibition of E. coli DAH7P synthase was shown to be dependent on compound chirality. Several other phosphoenolpyruvate-like molecules were prepared, and were also shown to be effective DAH7P synthase inhibitors. In Chapter Four extended compounds are designed that will bind the enzyme by multiple interactions at both substrate binding sites. Four compounds were prepared, and an increase in inhibitory potency was observed. In Chapter Five computational techniques are explored to aid the interpretation of the inhibition of DAH7P synthase by the compounds prepared in these studies. Several approaches for more potent inhibition of this enzyme are outlined and discussed.

Moderate doses of alcohol impair response inhibition activation. Recent work has shown that, during a single dose, response inhibition recovers from the impairing effects of alcohol more slowly than response activation. Evidence for a lag in tolerance development to inhibitory versus activational mechanisms suggests that, as blood alcohol declines, drinkers’ response inhibition might continue to be impaired, despite the recovery of response activation. However, this has not been studied across repeated doses. This study examined how cross-session tolerance to alcohol develops differentially between response activation and inhibition. Thirty-two healthy adults performed a cued go/no-go task that measured response activation and inhibition. The study tested the degree to which response activation and inhibition developed acute and cross-session tolerance to a moderate dose of alcohol (0.65 g/kg) administered twice. Alcohol impaired response activation and inhibition during both administrations. Response activation displayed acute tolerance to alcohol during both administrations and cross-session tolerance from the first to second administration. Response inhibition was impaired by each alcohol administration but showed no acute or cross-session tolerance. Evidence of biased recovery of response activation over inhibition during a single dose and as doses are repeated could contribute to some of the impulsive behavior commonly observed under alcohol.

Due to the depletion of petroleum reserves and environmental concerns, bioethanol has been identified as an alternative fuel to petrol. Bioethanol is a fuel of bio-origin derived from renewable biomass. Starch and sugar containing materials are the primary sources of carbon for bioethanol production. Starch is firstly hydrolysed into simple sugars which are later fermented to bioethanol using Saccharomyces cerevisiae (S. cerevisiae). The fermentation of sugars to bioethanol is however limited by inhibition of S. cerevisiae by the major product of the process, bioethanol. The challenge is thus in keeping the bioethanol concentration at levels which are not harmful to the fermenting organism. Keeping bioethanol concentration low in the broth will provide a suitable environment for yeast to grow and thus increase the overall production. Currently bioethanol producers use high water dilution rates to keep the bioethanol concentrations in the broth low enough so that yeast is not harmed. This excess water has to be removed in the downstream process, which is expensive. The use of excessive amounts of water in the fermentation can be avoided by continual removal of bioethanol from the broth. During this investigation the experimental conditions for the hydrolysis process were determined. A pH of 5.5 was determined as the best pH for Termamyl SC at 95°C with a pH of 5.0 for Spirizyme Fuel at 55°C during the liquefaction and the saccharification step, respectively. During the fermentation process the influence of yeast concentration on bioethanol production was investigated by varying the yeast concentration between 2 g.L-1 and 7 g.L-1. A yeast concentration of 5 g.L-1 produced the highest bioethanol yield of 0.48 g.g-1 after 48 hours of fermentation using S. cerevisiae. Later during the investigation a coupled fermentation/pervaporation system was employed in a batch system for continual removal of bioethanol in the fermentation broth in a process called simultaneous fermentation and separation (SFS). Through the continuous removal of bioethanol from the fermentation broth, the bioethanol concentration in the broth was kept low enough so that it was not harmful to the fermenting organism but the overall fermentation yield was not improved. Pervaporation is a membrane separation process used to separate azeotropic mixtures such as bioethanol and water. It is highly efficient, cost effective and uses less energy than distillation. During the SFS process a bioethanol yield of 0.22 g.g-1 was obtained. The SFS process yield for bioethanol was low compared to 0.45 g.g-1 of the traditional batch fermentation process. The lower overall bioethanol yield obtained in the SFS process could be attributed to only the supernatant being used in the SFS process and not the entire fermentation broth as in the traditional process. The results from this study proved that the SFS process was less efficient compared to the traditional batch fermentation process with respect to the bioethanol yield, but that the fermentation could be carried out without the necessity for additional process water. / Thesis (M.Sc. Engineering Sciences (Chemical and Minerals Engineering))--North-West University, Potchefstroom Campus, 2010.

Substances of natural and synthetic origin were studied using analytical, bioassay guided isolation, metabolomics and medicinal chemistry techniques. In a section focused on the plant family Marcgraviaceae, a validated method for the quantification of six pentacyclic triterpenes (α and β Amyrin lupeol, ursolic acid, betulin and betulinic acid) in the Souroubea spp was developed. Quantification of the triterpenes in the crude extracts was achieved using HPLC-APCI mass selective detection. The calibration curves for the five triterpenes evaluated were highly linear (r2 >0.993) and percentage recovery from spiked samples were greater than 94% for all compounds. The LOD for betulinic acid was 0.01 µg for betulinic acid on column and LOQ was 0.03 µg. The method was successfully applied to 41 crude extracts from leaf and stem of Souroubea spp, from two locations in Costa Rica. The method is suitable for quality control of raw materials used in the manufacture of natural health products. The use of modern metabolomic techniques, UHPLC-QTOF allowed the identification of five putative makers that can potentially be used in distinguishing between the two Souroubea species. The validated method was used in the quantification of the above triterpenes in a total of thirteen Marcgraviaceae species collected in Costa Rica. It was established that betulinic acid and β- Amyrin could be used as makers for this family of tropical vines. These same thirteen plants extracts were evaluated in antifungal and quorum sensing inhibition bioassays. Marcgravia nervosa was the only species that showed significant activity in both bioassays. Bioassay guided fractionation of the crude ethanolic extract of M. nervosa led to the identification of 2-methoxynaphthoquinone as the bioactive compound responsible for the bioactivity. The crude leaf ethanolic extract from M. nervosa showed a significant inhibition of QS comparable or somewhat better than D. pulchra extracts with the M. nervosa extract showing stronger inhibiting QS with a halo of 21.8mm, more than D. pulcra extracts which generated a halo of 15.9mm. The active quinone has a MIC of 85 µM against Saccharomyces cerevisiaBY4741 (haploid) and 100 µM against Saccharomyces cerevisiae BY4743 (diploid) compared to berberine (positive control) with a MIC 600 µM for both strains. This quinone is not present in any of the other twelve species of Marcgraviaceae available to us. In work focusing on organic synthesis, a total of 57 semi-synthetic derivatives of dillapiol, safrol and piperonal were prepared and evaluated for their inhibitory activity in a CYP 3A4 bioassay to assess their potential use as pesticide synergists. The synergistic activity of dillapiol has been improved 45 fold; analog 31 has an IC50 = 0.2 µM compared with dillapiol IC50= 9.18 µM. A number of other compounds structurally related to 31 showed similar levels of activity. A screening of a compound library identified the amino sulfoxide 3 as a potential lead for the design of a selective connexin blocker with potential application in the treatment of spinal cord injuries. The use of X-ray crystallography permitted the correction of the original structure assigned to 3. Once the structure was corrected a total of 6 analogs were prepared. Compound 3 has the highest inhibition of GJIC whereas compound 8 and compound 2, reduced anionic hemi-channel activity. Compound 2 also reduced the cationic activity of the hemi-channels.