Gold Nanoparticles as Adjuncts in Electrophoretic Analysis of Biopolymers

This thesis discusses two projects which investigate the analysis and application of biomolecules. Chapter 1 provides the background and context of this work. Chapter 2 discusses the synthesis and characterisation of three types of gold nanoparticles, sodium citrate, cetyltrimethyl ammonium bromide (CTAB) and 4-dimethylamino pyridine (DMAP) protected gold nanoparticles, as well as two types of polyacrylamide gel, TBE and tris-HCl. Successful characterisation of these gold nanoparticles was carried out by dynamic light scattering, ultra-violet visible spectroscopy and transmission emission microscopy. The characterisation of polyacrylamide gels was achieved by investigation by rheology, optical coherence tomography and scanning electron microscopy. Rheological analysis demonstrated that the addition of gold nanoparticles to polyacrylamide reduces the elasticity of the gel but stabilises the linear viscoelastic range, while not changing the core properties of the material. These materials were then used to separate sulfur containing biomolecules by polyacrylamide gel electrophoresis. Chapter 3 discusses the efficacy of the three types of gold nanoparticles to separate sulfur containing biomolecules when added to polyacrylamide gel electrophoresis. Citrate and CTAB gold nanoparticles where unsuccessful when used to analyse phosphorothioated DNA, however, DMAP gold nanoparticles showed promising results. DMAP gold nanoparticles were further tested with the proteins lysozyme, bovine serum albumin (BSA), glutathione-S-transferase (GST) and RNAse A (RNAse). The retention of BSA and GST was successful however there was continued problems with visualising lysozyme and RNAse. This led to the development of ATP polyacrylamide gel electrophoresis. Chapter 4 discusses the theoretical application of the development of catalytic DNA aptamers using in vitro selection. We synthesised a DNA aptamer library, two primers and two biotinylated strands of DNA, to be used in the selection process. However due to poor purity and yields, the project remains in its infancy. We discuss the methods by which we would conduct this research and our justifications for these processes

Juvenile justice: punitive perspectives on children, sex offences and the age of criminal responsibility

There are social and legislative drives to investigate factors that may diminish criminal responsibility within juvenile sexual offenders, and whether they are publicly perceived to hold moral capacity on sexual deviancy. The current study aimed to measure attitudes towards the criminal responsibility of juvenile sexual offenders; in relation to perpetrator age, intellectual disability and type of sexual coercion committed. This study also assessed associations between perceived criminal responsibility, attitudes towards treatment of sexual offenders, and understanding of UK age legislation. A sample of 206 UK adults found juveniles were more likely to be perceived of diminished criminal responsibility when involved in a sexual relationship with a peer of the same developmental age; diagnosed with an intellectual disability; and committing a non-contact offence. Overall attitudes towards the treatment of juvenile sexual offenders were positive, with significant associations between perceptions of treatment and perceptions of criminal responsibility. Understanding of UK age legislation demonstrated that participants had an overall poor understanding of the age of criminal responsibility – presenting no significant associations. Limitations and implications for evidence-based sex offender policies are discussed

Subtle deregulation of the Wnt-signalling pathway through loss of Apc2 reduces the fitness of intestinal stem cells

The importance of the Wnt-signaling pathway on the regulation and maintenance of the intesti- nal stem cell (ISC) population is well recognized. However, our current knowledge base is founded on models using systems of gross deregulation of the Wnt-signaling pathway. Given the importance of this signaling pathway on intestinal homeostasis, there is a need to explore the role of more subtle alterations in Wnt-signaling levels within this tissue. Herein, we have used a model of Apc2 loss to meet this aim. Apc2 is a homolog of Apc which can also form a destruction complex capable of binding b-catenin, albeit less efficiently than Apc. We show that systemic loss of Apc2 results in an increase in the number of cells displaying nuclear b-catenin at the base of the intestinal crypt. This subsequently impacts the expression levels of several ISC markers and the fitness of ISCs as assessed by organoid formation efficiency. This work pro- vides the first evidence that the function and fitness of ISCs can be altered by even minor mis- regulation of the Wnt-signaling pathway. Our data highlights the importance of correct maintenance of this crucial signaling pathway in the maintenance and function of the ISC popu- lation

An all-in-one UniSam vector system for efficient gene activation

AbstractWe have generated a drug-free, all-in-one dCAS9-SAM vector that can activate endogenous gene expression with the potential to modify cell fate. We demonstrate that this strategy can be used in a number of cell lines and avoids exceptionally high levels of gene expression that are observed in standard transgenic approaches. Compared to the multi-plasmid system, this all-in-one vector activates gene expression to a comparable level but the reduced overall DNA content results in significantly higher viability of transfected cells. This allowed us to use the RUNX1C-GFP human embryonic stem cell reporter cell line to monitor gene activation in individual cells and to show that activation could occur at all stages of the cell cycle.</jats:p

Does physical activity mediate the associations between blue space and mental health? : a cross-sectional study in Australia

Background Research has begun to examine whether blue space is beneficial to mental health. While results are promising, it is difficult to know which aspects of mental health or mental ill-health may benefit most. Physical activity has been proposed as one potential mechanism via which blue space may be associated with better mental health. However, very few studies have examined mechanisms. We examined associations between blue space proximity and a range of mental health outcomes and examined which of these associations were mediated by physical activity. Methods 350 participants (M=38.74, SD=14.92, 70% female) self-reported their weekly physical activity and completed measures of depression, anxiety, and psychological wellbeing. We then used GIS software to calculate blue space proximity (i.e., coastal and inland), and structural equation modelling with mediation paths to determine the role of physical activity in the associations between bluespace and mental health. Results Physical activity partially mediated the associations between coastal proximity and depression (β=0.02, 95% CI=0.001, 0.05), anxiety (β=0.03, 95% CI=0.01, 0.06), and wellbeing (β = −0.03, 95% CI=−0.08, −0.01), and fully mediated the associations between inland water proximity and depression (β=0.02, 95% CI=0.003, 0.05), anxiety (β=0.03, 95% CI=0.01, 0.07), and wellbeing (β = −0.03, 95% CI=−0.07, −0.01). Conclusion While physical activity appears to explain associations between inland blue space and mental health outcomes, it only partially explains the association between coastal blue space and mental health, suggesting other mechanisms may play a role and even inactive exposure may be beneficial

Neural responses to a modified Stroop paradigm in patients with complex chronic musculoskeletal pain compared to matched controls: an experimental functional magnetic resonance imaging study

Background: Chronic musculoskeletal pain (CMSKP) is attentionally demanding, complex and multi-factorial; neuroimaging research in the population seen in pain clinics is sparse. A better understanding of the neural activity underlying attentional processes to pain related information compared to healthy controls may help inform diagnosis and management in the future. Methods: Blood oxygenation level dependent functional magnetic resonance imaging (BOLD fMRI) compared brain responses in patients with CMSKP (n=15) and healthy controls (n=14) while completing a modified Stroop task using pain-related, positive-emotional, and neutral control words. Results: Response times in the Stroop task were no different for CMSKP patients compared with controls, but patients were less accurate in their responses to all word types. BOLD fMRI responses during presentation of pain-related words suggested increases in neural activation in patients compared to controls in regions previously reported as being involved in pain perception and emotion: the anterior cingulate cortex, insula and primary and secondary somatosensory cortex. No fMRI differences were seen between groups in response to positive or control words. Conclusions: Using this modified Stroop tasks, specific differences were identified in brain activity between CMSKP patients and controls in response to pain-related information using fMRI. This provided evidence of differences in the way that pain-related information is processed in those with chronic complex musculoskeletal pain that were not detectable using the behavioural measures of speed and accuracy. The study may be helpful in gaining new insights into the impact of attention in those living with chronic pai

“Nobody knows, or seems to know how rheumatology and breastfeeding works”: Women's experiences of breastfeeding whilst managing a long-term limiting condition – A qualitative visual methods study

Background Only around 1% of babies in the UK are breastfed exclusively until six months of age as recommended by the World Health Organisation. One in ten women who have recently given birth in the UK have a long-term illness and they are at increased risk of stopping breastfeeding early. We considered women with autoimmune rheumatic diseases as an exemplar group of long term illnesses, to explore the barriers and enablers to breastfeeding Aim To understand the experiences of infant feeding among women with autoimmune rheumatic diseases and to identify potential barriers and enablers. Design Qualitative visual timeline-facilitated interviews. Participants and setting 128 women with autoimmune rheumatic diseases who were considering pregnancy, pregnant, or had young children took part in an online survey as part of the STAR Family Study. Of these, 13 women who had children were purposefully sampled to be interviewed. Interviews took place in person or on the telephone. Timeline-facilitated interviews were used to focus on lived experiences and topics important to the women, including early parenting. We conducted a focused thematic analysis of women's lived experiences of infant feeding. Results Three main themes were identified in relation to breastfeeding: lack of information about medication safety, lack of support in decision-making and maintaining breastfeeding, and maternal guilt. Conclusions Women with autoimmune rheumatic diseases found it difficult to access the information they needed about medications to make informed decisions about breastfeeding. They often also felt pressurised into breastfeeding and experienced feelings of guilt if they were unable, or did not wish to breastfeed. Tailored interventions are required that adopt a non-judgmental and person-centred approach to support decision-making in regard to infant feeding, providing women with information that can best enable them to make infant feeding choices

The outcome of pregnancy in women with cystic fibrosis : a UK population based descriptive study.

Peer reviewedPostprin

Prognostic factors for a change in eye health or vision: A rapid review

The general public are advised to have regular routine eye examinations to check their vision and ocular health; however current UK guidance on how often to have eye examinations is not evidence-based and was issued in 2002.This Rapid Review aims to provide an evidence base that stakeholders can use to form updated guidance for Wales by asking the question ‘What are the prognostic factors for a change in ocular status in the general population attending routine eye examinations?’The review included evidence available from January 2009 up until August 2023. Evidence was included from 2011 up until 2023. 19 studies were included: two systematic reviews; nine prospective cohort studies; three retrospective cohort studies; two longitudinal studies; two case-control studies; and one cross-sectional study were included.<br/

Pregnancy rates and outcomes amongst women with cystic fibrosis in the UK : comparisons with the general population before and after the introduction of disease modifying treatment, 2003-17

Acknowledgements We thank the UK CF Registry team and the UK CF centres and clinics for submitting data to the Registry. Special thanks to the people with cystic fibrosis and their families who have agreed for their UK CF Registry data to be used for research. Funding The study was funded by a Welsh Government Research for Patient and Public Benefit grant. The funder was not involved in the study design, data collection, data analysis, data interpretation or the writing of the report. DT-R is funded by the MRC on a Clinician Scientist Fellowship (MR/P008577/1).Peer reviewedPublisher PD