The design and synthesis of inhibitors of the enzyme poly (adenosine diphosphate ribose) polymerase and the investigation of the mechanism of action of the suspected ultimate carcinogen chloroacetaldehyde

PhD ThesisThis thesis describes the preparation of inhibitors of the enzyme poly(adenosine diphosphate ribose)polymerase (PARP) and the mechanism of action of the carcinogen vinyl chloride. Initially the enzyme's function, purification and its known inhibitors are discussed. The enzyme's cofactor, nicotinamide adenine diphosphate (NAD+), is described in terms of its function in PARP and other enzymes, and its crystal structure and solution conformation(s). The high field proton and carbon-13 NMR spectra of this compound are fully assigned using two dimensional NMR experiments (COSY and ROESY) and selective spin decoupling. Using similar techniques the high field proton and carbon- 13 NMR spectra of the reduced analogue, NADH, are assigned. This NMR data showed that the conformation of the 1,4- dihydropyridine ring of this molecule was a dynamic equilibrium mixture of two boats. However, it was discovered that these boat conformations were unequally populated and a rationale for this phenomenon is given. The rationale of enzyme inhibitor design with respect to PARP is described and the subsequent preparation and potencies of PARP inhibitors are both described and discussed. These included the preparation of 3-methoxycyclohexane carboxamide which was synthesised to test a hypothesis of the nature of PARP inhibitors. Partially saturated substituted benzamide systems were synthesised to determine the degree of unsaturation necessary for inhibition while similar compounds were synthegised by reducing N-alkylnicotinamides. Direct NAD+ analogues were prepared in which an adenine unit was separated by a methylene chain from a 3-oxy substituted benzamide. By varying the length of the methylene chain 3-(9-(12-dodecyloxy)adenine)benzamide was shown to inhibit the enzyme. Amongst these inhibitors were the unique compounds benzoxazole-4-carboxamides which were found to be the most potent inhibitors to be synthesised during this period. A correlation of the physical properties of literature compounds with their activity is discussed along with the applicability of this technique to PARP and its inhibitors. The metabolic activation and carcinogenesis of vinyl chloride are described. The reaction of chloroacetaldehyde (a metabolite of vinyl chloride) is discussed with a detailed description of its reaction with nucleosides to form the fluorescent etheno compounds, with particular emphasis on the adeninq system. A detailed investigation of the reaction mechanism of chloroacetaldehyde with adenosine was undertaken by determining the intermediates and products of the acid catalysed decomposition of N6-(2,2-dimethoxyethyl)adenosine. An ethenoadenine deuteriation experiment along with the previous experiment gave evidence toward the reaction mechanism of chloroacetaldehyde with adenosine. In summary, the cofactor of PARP NAD+ has been fully characterised by proton and carbon-13 NMR spectroscopy; a unique conformational. property of NADH has been characterised; a new and novel class of potent inhibitors of PARP has been discovered and the reaction mechanism of chloroacetaldehyde with adenosine has been determined.Northern Cancer Research Campaign

Bayesian time perception

Time is an elemental dimension of human perception, cognition and action. Innumerable studies have investigated the perception of time over the last 100 years, but the computational basis for the processing of temporal information remains unknown. This thesis aims to understand the mechanisms underlying the perceived timing of stimuli. We propose a novel Bayesian model of when stimuli are perceived that is consistent with the predictive coding framework – such a perspective to how the brain deals with temporal information forms the core of this thesis. We theorize that that the brain takes prior expectations about when a stimulus might occur in the future (prior distribution) and combines it with current sensory evidence (likelihood function) in order to generate a percept of perceived timing (posterior distribution). In Chapters 2-4, we use human psychophysics to show that the brain may bias perception such that slightly irregularly timed stimuli as reported as more regular. In Chapter 3, we show how an environment of irregularity can cause regularly timed sequences to be perceived as irregular whilst Chapter 4 shows how changes in the reliability of a signal can cause an increased attraction towards expectation

Transportation Planning in a Growing Community

The Town of Red Cross - Past and Present: Red Cross was founded in the late 1700s and for nearly two hundred years life changed very little. Originally called “Red Crossing,” the Town owes its name to the rich red dirt of the Piedmont and the crossroads of what would eventually become N.C. 24/27 and N.C. 205. N.C. 24/27 remained unpaved until 1925, and N.C. 205 was not paved until 1941. Rolling farmland dotted by farmhouses dominated the landscape. A few small-scale farming-related businesses and industries served the area, and by the early 1900s, several small stores operated at the crossroads. Life and land use remained fairly constant until the latter part of the Twentieth Century

Rapid calibration to dynamic temporal contexts

The prediction of future events and the preparation of appropriate behavioural reactions rely on an accurate perception of temporal regularities. In dynamic environments, temporal regularities are subject to slow and sudden changes, and adaptation to these changes is an important requirement for efficient behaviour. Bayesian models have proven a useful tool to understand the processing of temporal regularities in humans; yet an open question pertains to the degree of flexibility of the prior that is required for optimal modelling of behaviour. Here we directly compare dynamic models (with continuously changing prior expectations) and static models (a stable prior for each experimental session) with their ability to describe regression effects in interval timing. Our results show that dynamic Bayesian models are superior when describing the responses to slow, continuous environmental changes, whereas static models are more suitable to describe responses to sudden changes. In time perception research, these results will be informative for the choice of adequate computational models and enhance our understanding of the neuronal computations underlying human timing behaviour

Increased SK3 expression in DM1 lens cells leads to impaired growth through a greater calcium-induced fragility

Although cataract is a characteristic feature of myotonic dystrophy type 1 (DM1), little is known of the underlying mechanisms. We generated four lens epithelial cell lines derived from DM1 cataracts and two from age-matched, non-DM cataracts. Small-pool PCR revealed typical large triplet repeat expansions in the DM1 cells. Furthermore, real-time PCR analysis showed reduced SIX5 expression and increased expression of the Ca2+-activated K+ channel SK3 in the DM1 cells. These cells also exhibited longer population doubling times which did not arise through reduced proliferation, but rather increased cell death as shown by increased release of lactate dehydrogenase (LDH). Using 86Rb+ as a tracer for K+, we found no difference in the resting K+ influx or efflux kinetics. In all cases, the ouabain sensitive component of the influx contributed ~50% of the total. However, stimulating internal Ca2+ by exposure to ionomycin not only caused greater stimulation of K+ (86Rb) efflux in the DM1 cells but also induced a higher rate of cell death (LDH assay). Since both the hyper-stimulation of K+ efflux and cell death were reduced by the highly specific SK inhibitor apamin, we suggest that increased expression of SK3 has a critical role in the increased Ca2+-induced fragility in DM1 cells. The present data, therefore, both help explain the lower epithelial cell density previously observed in DM1 cataracts and provide general insights into mechanisms underlying the fragility of other DM1-affected tissues

EXPRESS: Rapid calibration to dynamic temporal contexts

TThe prediction of future events and the preparation of appropriate behavioural reactions rely on an accurate perception of temporal regularities. In dynamic environments, temporal regularities are subject to slow and sudden changes, and adaptation to these changes is an important requirement for efficient behaviour. Bayesian models have proven a useful tool to understand the processing of temporal regularities in humans; yet an open question pertains to the degree of flexibility of the prior that is required for optimal modelling of behaviour. Here we directly compare dynamic models (with continuously changing prior expectations) and static models (a stable prior for each experimental session) with their ability to describe regression effects in interval timing. Our results show that dynamic Bayesian models are superior when describing the responses to slow, continuous environmental changes, whereas static models are more suitable to describe responses to sudden changes. In time perception research, these results will be informative for the choice of adequate computational models and enhance our understanding of the neuronal computations underlying human timing behaviour

Local and Regional North Carolina Collaborations: Case Studies from the North Carolina Chapter of the American Planning Association

Carolina Planning regularly publishes a feature highlighting projects from members of the North Carolina Chapter of the American Planning Association (APA-NC). This year’s submissions range from multi-county visioning efforts to small-town transportation planning, demonstrating that planning at any scale can benefit from innovative collaboration. Includes the following case studies: Innovative Partnerships (Introduction); Opt-In Project Explores Uncharted Planning Territory in Southwestern NC; Transportation Planning in a Growing Community; Playing Around with Civic Engagement Strategies in the Triad; Simple Changes and Collaboration: NCDOT and the Town of West Jefferson Partner to Revitalize Downtow

Sitting time, fidgeting and all-cause mortality in the UK Women's Cohort Study

Introduction: Sedentary behaviours (including sitting) may increase risk of mortality independently of physical activity level. Little is known about how fidgeting behaviours might modify the association. Methods: Data were drawn from the UK Women’s Cohort Study. In 1999/2002, 12,778 women (age 37 to 78) provided data on average daily sitting time, overall fidgeting (irrespective of posture), and a range of relevant covariates including physical activity, diet, smoking status and alcohol consumption. Participants were followed for mortality over a mean of 12 years. Proportional hazards Cox regression models were used to estimate the relative risk of mortality in the high (vs. low) and medium (vs. low) sitting time groups. Results: Fidgeting modified the risk associated with sitting time (p value for interaction = 0.04), leading us to separate groups for analysis. Adjusting for a range of covariates, sitting for 7+ hours/day (vs. <5 hours/day) was associated with 30% increased risk of all-cause mortality (HR = 1.30, 95% CI 1.02, 1.66) only among women in the low fidgeting group. Among women in the high fidgeting group, sitting for 5/6 (vs. <5 hrs/day) was associated with decreased risk of mortality (HR = 0.63, 95% CI 0.43, 0.91), adjusting for a range of covariates. There was no increased risk of mortality from longer sitting time in the middle and high fidgeting groups. Conclusions: Fidgeting may reduce the risk of all-cause mortality associated with excessive sitting time. More detailed and better validated measures of fidgeting should be identified in other studies in order to replicate these findings and identity mechanisms, particularly measures that distinguish fidgeting in a seated from standing posture

Bridging Alone: Religious Conservatism, Marital Homogamy, and Voluntary Association Membership

This study characterizes social insularity of religiously conservative American married couples by examining patterns of voluntary associationmembership. Constructing a dataset of 3938 marital dyads from the second wave of the National Survey of Families and Households, the author investigates whether conservative religious homogamy encourages membership in religious voluntary groups and discourages membership in secular voluntary groups. Results indicate that couples’ shared affiliation with conservative denominations, paired with beliefs in biblical authority and inerrancy, increases the likelihood of religious group membership for husbands and wives and reduces the likelihood of secular group membership for wives, but not for husbands. The social insularity of conservative religious groups appears to be reinforced by homogamy—particularly by wives who share faith with husbands

Dealing with Alcohol-related problems in the Night-Time Economy: A Study Protocol for Mapping trends in harm and stakeholder views surrounding local community level interventions

<p>Abstract</p> <p>Background</p> <p>This project will provide a comprehensive investigation into the prevalence of alcohol-related harms and community attitudes in the context of community-based interventions being implemented to reduce harm in two regional centres of Australia. While considerable experimentation and innovation to address these harms has occurred in both Geelong and Newcastle, only limited ad-hoc documentation and analysis has been conducted on changes in the prevalence of harm as a consequence, leaving a considerable gap in terms of a systematic, evidence-based analysis of changes in harm over time and the need for further intervention. Similarly, little evidence has been reported regarding the views of key stakeholder groups, industry, government agencies, patrons or community regarding the need for, and the acceptability of, interventions to reduce harms. This project will aim to provide evidence regarding the impact and acceptability of local initiatives aimed at reducing alcohol-related harms.</p> <p>Methods/Design</p> <p>This study will gather existing police data (assault, property damage and drink driving offences), Emergency Department presentations and Ambulance attendance data. Further, the research team will conduct interviews with licensed venue patrons and collect observational data of licensed venues. Key informant interviews will assess expert knowledge from key industry and government stakeholders, and a community survey will assess community experiences and attitudes towards alcohol-related harm and harm-reduction strategies. Overall, the project will assess: the extent of alcohol-related harm in the context of harm-reduction interventions, and the need for and acceptability of further intervention.</p> <p>Discussion</p> <p>These findings will be used to improve evidence-based practice both nationally and internationally.</p> <p>Ethical Approval</p> <p>This project has been approved by Deakin University HREC.</p