The Role of Glutathione Transferases in the Detoxification of TNT

Manufacture, use, storage and improper disposal of the explosive 2,4,6-trinitrotoluene (TNT) have led to widespread, global contamination of soil and groundwater. TNT is highly toxic and recalcitrant to degradation resulting in environmental build-up with far reaching ecological and health implications. It is therefore a priority to remove contaminating TNT from the environment. Phytoremediation is a promising solution; suitable plants possess some natural ability to transform TNT, high biomass, deep root systems, requirement for minimal nutrient input and ability to reduce contamination spread by wind or water erosion; making them an attractive remediation system. Key genes involved in the detoxification of TNT by plants have been recently identified by expression studies and the encoded enzymes characterised. This has lead to the thorough investigation of the enzymes in the pathway of TNT detoxification; Phase I transformation includes oxophytodienoate reductases, with uridine diphosphate (UDP) glycosyltransferases (UGTs) playing a role in the Phase II conjugation step. The expression studies identified additional enzymes also likely to be involved in these phases including glutathione transferases (GSTs). GSTs are known to detoxify compounds by conjugation to glutathione (GSH), and like UGTs are Phase II detoxification enzymes. This thesis presents an investigation into whether plant GSTs play a role in the detoxification of TNT. In vitro analysis of recombinant GSTs was performed to elucidate the activity of GSTs towards TNT. Seven GSTs were cloned, expressed and purified from Escherichia coli. TNT assays performed with pure enzyme indicated that at least two of the GSTs were able to transform TNT. Analysis of the reaction product by mass spectrometry showed that TNT was conjugated to glutathione through substitution of a nitro-group, a highly desirable reaction as the removal of a nitro group from TNT is likely to increase the likelihood of subsequent mineralisation of the pollutant. This is the first identification of enzymes capable of this transformation. The two GSTs which exhibited activity towards TNT were overexpressed in Arabidopsis to clarify if the conjugating activity observed in vitro was able to confer increased tolerance to TNT to the transformed plants. Transgenic lines showed no enhanced growth compared to wild type plants on TNT amended media, root lengths appeared slightly shorter while TNT uptake and biomass were reduced. The role of GSTs in the detoxification of TNT remains unresolved however it is likely that GSTs do not play an integral role in TNT detoxification in plants. Nonetheless, the two GSTs characterised in the project are the first examples of plant enzymes which are able to catalyse the removal of nitro groups from TNT. Engineering these GSTs to improve their ability to transform TNT could offer an opportunity for effective environmental remediation

Factors influencing job satisfaction at Toyota SA Motors.

Thesis (MBA)-University of KwaZulu-Natal, Westville, 2010.The South African Motor industry is evolving through challenging times, placing tremendous pressure on Toyota SA Motors (TSAM) to remain efficient. The organisation has identified human capital as one of its most valuable resources contributing to the achievement of this goal. It is therefore imperative that the organisation retain staff members through focusing on the influences of job satisfaction, an impoliant aspect affecting staffretention at TSAM. The objective of the study was to examine the factors influencing job satisfaction at TSAM with the aim of detennining if new age job benefits influence job satisfaction, the impact of new age benefits on the business, if job satisfaction would result in staff retention and to provide recommendations to TSAM on how to stimulate job satisfaction levels in order to increase staff retention. This was to be achieved through the brcakdown and analysis of the relevant components of job satisfaction, along with theory comparison and the aid of quantitative research. The objective was achieved by sampling 235 employees of TSAM. A response rate of 66% was achieved using an online questionnaire to collect data. The data analysis process was completed using the Statistical Package for the Social Sciences (SPSS). Overall, new age benefits were found to have a positive influence on job satisfaction with the highest preferences being flexitime, flexible workspace, a free canteen and a gym facility. The other new age benefits proved to be less popular amongst respondents. A large percentage of respondents believed that TSAM should revise the benefits offered and that the company has the capacity to do so. It was found that the majority of respondents would accept a job offer elsewhere if benefits were good and remuneration was less then expected. Employees believed that new age benefits would have a positive impact on the business as job satisfaction results in staff retention within Toyota SA Motors and that staff turnover is a criteria to judge the company by

Postural Changes in Blood Pressure Associated with Interactions between Candidate Genes for Chronic Respiratory Diseases and Exposure to Particulate Matter

BACKGROUND. Fine particulate matter [aerodynamic diameter ≤ 2.5 μm (PM2.5)] has been associated with autonomic dysregulation. OBJECTIVE. We hypothesized that PM2.5 influences postural changes in systolic blood pressure (ΔSBP) and in diastolic blood pressure (ΔDBP) and that this effect is modified by genes thought to be related to chronic lung disease. METHODS. We measured blood pressure in participants every 3-5 years. ΔSBP and ΔDBP were calculated as sitting minus standing SBP and DBP. We averaged PM2.5 over 48 hr before study visits and analyzed 202 single nucleotide polymorphisms (SNPs) in 25 genes. To address multiple comparisons, data were stratified into a split sample. In the discovery cohort, the effects of SNP x PM2.5 interactions on ΔSBP and ΔDBP were analyzed using mixed models with subject-specific random intercepts. We defined positive outcomes as p < 0.1 for the interaction; we analyzed only these SNPs in the replicate cohort and confirmed them if p < 0.025 with the same sign. Confirmed associations were analyzed within the full cohort in models adjusted for anthropometric and lifestyle factors. RESULTS. Nine hundred forty-five participants were included in our analysis. One interaction with rs9568232 in PHD finger protein 11 (PHF11) was associated with greater ΔDBP. Interactions with rs1144393 in matrix metalloprotease 1 (MMP1) and rs16930692, rs7955200, and rs10771283 in inositol 1,4,5-triphosphate receptor, type 2 (ITPR2) were associated with significantly greater ΔSBP. Because SNPs associated with ΔSBP in our analysis are in genes along the renin-angiotensin pathway, we then examined medications affecting that pathway and observed significant interactions for angiotensin receptor blockers but not angiotensin-converting enzyme inhibitors with PM2.5. CONCLUSIONS. PM2.5 influences blood pressure and autonomic function. This effect is modified by genes and drugs that also act along this pathway.National Institute of Environmental Health Sciences (T32 ES07069, ES0002, ES015172-01, ES014663, P01 ES09825); United States Environmental Protection Agency (R827353, R832416); National Institutes of Health/National Institute of Aging (AG027014); United States Department of Veterans Affairs; Massachusetts Veterans Epidemiology Research and Information Cente

Black Carbon Exposure, Oxidative Stress Genes, and Blood Pressure in a Repeated-measures Study

Background: Particulate matter (PM) air pollution has been associated with cardiovascular morbidity and mortality, and elevated blood pressure (BP) is a known risk factor for cardiovascular disease. A small number of studies have investigated the relationship between PM and BP and found mixed results. Evidence suggests that traffic-related air pollution contributes significantly to PM-related cardiovascular effects. Objectives: We hypothesized that black carbon (BC), a traffic-related combustion by-product, would be more strongly associated with BP than would fine PM [aerodynamic diameter ≤ 2.5 μm (PM$_{2.5}$)], a heterogeneous PM mixture, and that these effects would be larger among participants with genetic variants associated with impaired antioxidative defense. Methods: We performed a repeated-measures analysis in elderly men to analyze associations between PM$_{2.5}$ and BC exposure and BP using mixed-effects models with random intercepts, adjusting for potential confounders. We also examined statistical interaction between BC and genetic variants related to oxidative stress defense: GSTM1, GSTP1, GSTT1, NQO1, catalase, and HMOX-1. Results: A 1-SD increase in BC concentration was associated with a 1.5-mmHg increase in systolic BP [95% confidence interval (CI), 0.1–2.8] and a 0.9-mmHg increase in diastolic BP (95% CI, 0.2–1.6). We observed no evidence of statistical interaction between BC and any of the genetic variants examined and found no association between PM$_{2.5}$ and BP. Conclusions: We observed positive associations between BP and BC, but not between BP and PM$_{2.5}$, and found no evidence of effect modification of the association between BC and BP by gene variants related to antioxidative defense

Medium-Term Exposure to Traffic-Related Air Pollution and Markers of Inflammation and Endothelial Function

Bac k g r o u n d: Exposure to traffic-related air pollution (TRAP) contributes to increased cardiovascular risk. Land-use regression models can improve exposure assessment for TRAP. Objectives: We examined the association between medium-term concentrations of black carbon (BC) estimated by land-use regression and levels of soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1), both markers of inflammatory and endothelial response. Me t h o d s: We studied 642 elderly men participating in the Veterans Administration (VA) Normative Aging Study with repeated measurements of sICAM‑1 and sVCAM‑1 during 1999–2008. Daily estimates of BC exposure at each geocoded participant address were derived using a validated spatiotemporal model and averaged to form 4-, 8-, and 12-week exposures. We used linear mixed models to estimate associations, controlling for confounders. We examined effect modification by statin use, obesity, and diabetes. Re s u l t s: We found statistically significant positive associations between BC and sICAM‑1 for averages of 4, 8, and 12 weeks. An interquartile-range increase in 8-week BC exposure (0.30 μg/m3) was associated with a 1.58 % increase in sICAM‑1 (95 % confidence interval, 0.18–3.00%). Overall association

Student autonomy of feedback format in higher education and perceived functional behaviours for academic development

In the current context of promoting active learning and raising student engagement within Higher Education, an increasing amount of research has looked at pedagogical-based design and factors that contribute to functional behaviours surrounding the interaction and use of academic assessment feedback. However, few studies have considered the perceived influence of student autonomy over feedback format and whether this promotes engagement and academic development. In this study, we recruited level 5 and 6 students (N = 38) on an undergraduate Education Programme (that has consistently implemented student feedback choice) to participate in initial self-reporting and subsequent focus groups ("soft triangulation‟). The findings revealed three core themes: [1] Personalisation – (a) sense of autonomy/involvement, (b) engagement and (c) motivation, [2] Clarity – (d) depth and detail, and [3] Areas for development. Overall, these findings suggest that feedback type – and the inherent option to choose – has a functional impact on academic engagement and development. We discuss these findings in relation to a sense of being valued that was associated with autonomy of choice, a divergence in how and when students engage with feedback, as well as the requirement for academic clarity and provision of formats that support academic development.University of Derby Undergraduate Research Scholarship Scheme (URSS

Particulate Air Pollution, Oxidative Stress Genes, and Heart Rate Variability in an Elderly Cohort

Background and Objectives: We have previously shown that reduced defenses against oxidative stress due to glutathione S-transferase M1 (GSTM1) deletion modify the effects of PM[2.5] (fine-particulate air pollution of < 2.5 μm in aerodynamic diameter) on heart rate variability (HRV) in a cross-sectional analysis of the Normative Aging Study, an elderly cohort. We have extended this to include a longitudinal analysis with more subjects and examination of the GT short tandem repeat polymorphism in the heme oxygenase-1 (HMOX-1) promoter. Methods: HRV measurements were taken on 539 subjects. Linear mixed effects models were fit for the logarithm of HRV metrics—including standard deviation of normal-to-normal intervals (SDNN), high frequency (HF), and low frequency (LF)—and PM2.5 concentrations in the 48 hr preceding HRV measurement, controlling for confounders and a random subject effect. Results: PM2.5 was significantly associated with SDNN (p = 0.04) and HF (p = 0.03) in all subjects. There was no association in subjects with GSTM1, whereas there was a significant association with SDNN, HF, and LF in subjects with the deletion. Similarly, there was no association with any HRV measure in subjects with the short repeat variant of HMOX-1, and significant associations in subjects with any long repeat. We found a significant three-way interaction of PM[2.5] with GSTM1 and HMOX-1 determining SDNN (p = 0.008), HF (p = 0.01) and LF (p = 0.04). In subjects with the GSTM1 deletion and the HMOX-1 long repeat, SDNN decreased by 13% [95% confidence interval (CI), −21% to −4%], HF decreased by 28% (95% CI, −43% to −9%), and LF decreased by 20% (95% CI, −35% to −3%) per 10 μg/m3 increase in PM. Conclusions: Oxidative stress is an important pathway for the autonomic effects of particles

Postural Changes in Blood Pressure Associated with Interactions between Candidate Genes for Chronic Respiratory Diseases and Exposure to Particulate Matter

Background: Fine particulate matter [aerodynamic diameter ≤ 2.5 μm (PM2.5)] has been associated with autonomic dysregulation.Objective We hypothesized that PM2.5 influences postural changes in systolic blood pressure (ΔSBP) and in diastolic blood pressure (ΔDBP) and that this effect is modified by genes thought to be related to chronic lung disease. Methods: We measured blood pressure in participants every 3–5 years. ΔSBP and ΔDBP were calculated as sitting minus standing SBP and DBP. We averaged PM2.5 over 48 hr before study visits and analyzed 202 single nucleotide polymorphisms (SNPs) in 25 genes. To address multiple comparisons, data were stratified into a split sample. In the discovery cohort, the effects of SNP × PM2.5 interactions on ΔSBP and ΔDBP were analyzed using mixed models with subject-specific random intercepts. We defined positive outcomes as p < 0.1 for the interaction; we analyzed only these SNPs in the replicate cohort and confirmed them if p < 0.025 with the same sign. Confirmed associations were analyzed within the full cohort in models adjusted for anthropometric and lifestyle factors. Results: Nine hundred forty-five participants were included in our analysis. One interaction with rs9568232 in PHD finger protein 11 (PHF11) was associated with greater ΔDBP. Interactions with rs1144393 in matrix metalloprotease 1 (MMP1) and rs16930692, rs7955200, and rs10771283 in inositol 1,4,5-triphosphate receptor, type 2 (ITPR2) were associated with significantly greater ΔSBP. Because SNPs associated with ΔSBP in our analysis are in genes along the renin–angiotensin pathway, we then examined medications affecting that pathway and observed significant interactions for angiotensin receptor blockers but not angiotensin-converting enzyme inhibitors with PM2.5. Conclusions: PM2.5 influences blood pressure and autonomic function. This effect is modified by genes and drugs that also act along this pathway

Black Carbon Exposure, Oxidative Stress Genes, and Blood Pressure in a Repeated-Measures Study

BACKGROUND. Particulate matter (PM) air pollution has been associated with cardiovascular morbidity and mortality, and elevated blood pressure (BP) is a known risk factor for cardiovascular disease. A small number of studies have investigated the relationship between PM and BP and found mixed results. Evidence suggests that traffic-related air pollution contributes significantly to PM-related cardiovascular effects. OBJECTIVES. We hypothesized that black carbon (BC), a traffic-related combustion by-product, would be more strongly associated with BP than would fine PM [aerodynamic diameter ≤ 2.5 μm (PM2.5)], a heterogeneous PM mixture, and that these effects would be larger among participants with genetic variants associated with impaired antioxidative defense. METHODS. We performed a repeated-measures analysis in elderly men to analyze associations between PM2.5 and BC exposure and BP using mixed-effects models with random intercepts, adjusting for potential confounders. We also examined statistical interaction between BC and genetic variants related to oxidative stress defense: GSTM1, GSTP1, GSTT1, NQO1, catalase, and HMOX-1. RESULTS. A 1-SD increase in BC concentration was associated with a 1.5-mmHg increase in systolic BP [95% confidence interval (CI), 0.1-2.8] and a 0.9-mmHg increase in diastolic BP (95% CI, 0.2-1.6). We observed no evidence of statistical interaction between BC and any of the genetic variants examined and found no association between PM2.5 and BP. CONCLUSIONS. We observed positive associations between BP and BC, but not between BP and PM2.5, and found no evidence of effect modification of the association between BC and BP by gene variants related to antioxidative defense.National Institute of Environmental Health Sciences (ES015172, ES014663); National Cancer Institute (2-T32-CA009330); United States Environmental Protection Agency (R832416); United States Deparment of Veterans Affairs; Massachusetts Veterans Epidemiology Research and Information Cente