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Developing a patient reported experience measure for gastrointestinal procedures (ENDOPREM)
Get PDFM.D. ThesisBackground: Gastrointestinal (GI) endoscopy and computed tomography
colonoscopy (CTC) are important diagnostic and therapeutic tools in the investigation
and management of gastrointestinal diseases. Current measures of patient
satisfaction and experience within GI endoscopy are clinician derived and measured.
This study aims to develop a patient reported experience measure (PREM) specific
to GI procedures, derived from the patient’s perspective.
Methods: The study comprised three phases. Phase 1: This qualitative phase
involved semi-structured interviews with patients who had recently undergone
endoscopy/CTC. Thematic analysis identified important aspects of the patient
experience. Phase 2: A questionnaire bank was developed from the thematic
analysis. An iterative process of review and revision within the wider study team
refined the questions. Rounds of cognitive interviews with patients who had
undergone GI procedures were used to further refine the questionnaire. Phase 3:
The resultant PREM was prospectively administered, for self-completion, to 1652
patients following a GI procedure. IBM® SPSS® 24 was used to investigate the
psychometric properties of the instrument.
Results: Phase 1: 35 participants participated in semi-structured interviews. Six
over-arching themes were identified: anxiety, expectations, information &
communication, embarrassment & dignity, choice &control and comfort. Phase 2:
Areas related to these themes were structured by procedural stage. Ten rounds of
review and revision within the study team were conducted, followed by five rounds of
cognitive interviews (total n=15). Phase 3: 799 participants completed the
questionnaire (response rate= 48.4%). Of the 59 questionnaire items, a ‘ceiling’
effect was present in 24. No questions demonstrated ‘floor’ effects. Individual item
completion rates were high, with only three items having >5% missing. Exploratory
factor analysis identified potential scales within the questionnaire.
Conclusion: The ENDOPREM™ is a tool which assesses all aspects of the GI
procedure experience. Potential future uses include assessing patient experience in
routine care or comparing experience associated with different endoscopic
interventions in trials
Controllable inkjet printing of graphene oxide and hydrophilic nanomaterials
Get PDFPhD ThesisGraphene oxide(GO)ink has been widely researched but problems still exist with regards to its printability and conductivity. The following work aims to combat both these issues, leading to the formulation of a glycol-based GOinkwith optimised rheologyfor use witha Dimatix inkjet system. Results show superior print quality and resolution, with films characterised by robust AFM, Raman, XRD, FTIR and optical studies. Through the application of a two-step reduction process, incorporating both chemical and thermal treatments, the conductivity of films are able to exceed 50,000 S/m. This is the highest value reported to date for an inkjet-printed graphene film yet the corresponding sheet resistance (740 W/sq) and transmittance (29 %) remain poor compared to indium tin oxide films currentlyused in optoelectronic devices.This work also aimed to apply the techniques of printing GO to alternative hydrophilic nanomaterials. The synthesis of cadmium sulphide/DNA nanowires via a wet chemical method allows for the easy formulation of a novel semiconducting ink that is also compatible with an inkjet printing system. The printing of highlyresolved cadmium sulphide/DNA droplet arrays on glass is reported with characterisation by fluorescence microscopy, AFM and Raman. The fabrication of conductive nanowire devices is also accomplished by the printing of CdS/DNA features over interdigitated electrodes and sintering of material. Findings show that inkjet printing could be a successful method of processing these semiconducting nanowires towards labelling and gas sensing applications
The influence of intercellular connections on beta cell functions
Get PDFPh. D. Thesis.Background
Pancreatic beta cells cultured in 2D systems display similar characteristics to dysfunctional beta cells of patients with diabetes. This thesis explores the impact of cell connectivity on functional and metabolic characteristics of insulin secreting beta cells.
Methods
The MIN6 mouse beta cell line was cultured as 2D monolayers and 3D structures named pseudoislets. The insulin secretion response and metabolic function of 2D and 3D MIN6 cultures, along with islets from human donors were compared. Roles for cell-cell interactions in regulating metabolic changes were explored with focus on the gap junctional protein, connexin36, using an inducible knockdown MIN6 cell line.
Results
MIN6 pseudoislets displayed improved functional responses compared to monolayers with 7.4-fold and 1.5-fold increases in glucose-induced insulin secretion respectively. XFe24 seahorse bioanalyser data showed the improved glucose-stimulated pseudoislet response was fuelled by large increases in glycolytic flux and a more moderate increase in oxidative phosphorylation. Basal insulin secretion and basal oxidative phosphorylation were both higher in monolayers but there were no differences in basal glycolysis. Human islets displayed a similar phenotype to pseudoislets with high contributions of glycolysis to glucose-induced ATP production. Pseudoislets indicated some hypoxia through trends towards increased lactate dehydrogenase and phosphoinositide-dependent kinase-1 expression and increased glycolytic activity of phosphfructokinase-1 and glyceraldehyde 3-phosphate
dehydrogenase but superior glucose sensing through decreased hexokinase I and increased GLUT2 expression and more active mitochondrial activities of pyruvate carboxylase, citrate synthase, α-ketoglutarate dehydrogenase, and malate dehydrogenase. Knockdown of connexin36 did not alter glucose-stimulated insulin secretion or metabolic flux. However, there was a trend towards increased basal insulin secretion and basal oxidative phosphorylation indicating a possible role for this connection in regulating basal metabolic flux.
Conclusions
The improved glucose-stimulated secretion conferred by pseudoislet configuration was accompanied by an increase in ATP production suggesting a role for alteration in metabolic flux in the improved functionality. Improved functional responses of beta cells in 3D structures was accompanied by a small increase in oxidative phosphorylation but a large increase in glycolysis that cannot be fully explained by hypoxia. Connexin36 may play a role in regulating the basal response but other connections are involved in regulating the glucose-stimulated response.Medical Research Council (MRC
Live-Cell Imaging of Human Oocytes and Regulation of Cohesin Removal in Meiosis II
Get PDFPh. D. ThesisThe “maternal age effect” describes the striking increase in risk of miscarriage and chromosomally abnormal embryos and children from women older than 35. Studies in mice have shown that the protein complex Cohesin is reduced in an age-dependent manner. This protein complex ensures accurate segregation during both rounds of meiosis, by holding the chromosomes together and providing a counteracting force to spindle microtubules.
Despite a wealth of knowledge generated from human oocytes, there are few live-cell studies, in part due to the paucity of material. This thesis uses human oocytes specifically donated for research to assess the effect of age on alignment of chromosomes at metaphase I and -II, which is a predictor of missegregation. Using high-resolution live-cell microscopy, it is clear that increased age is associated with chromosomes that are misaligned in metaphase I and -II.
At metaphase II, eggs arrest until they are fertilised by sperm. The regulation of how chromosomes separate at this point is poorly understood. While the bulk of Cohesin is removed in anaphase I, a small amount is “protected” by Shugoshin 2 and remains between the centromeres to allow for faithful segregation in meiosis II. Currently, there is poor experimental work to support the hypotheses proposed to explain how the mechanisms that protect Cohesin in meiosis I are removed in meiosis II. One of these hypotheses is the requirement of spindle tension to separate the protector, Shugoshin 2, from Cohesin in meiosis II. Here, I show that spindle tension is not required for deprotection and that alternative models should be considered, such as one which suggests that higher-order regulation around meiosis II resumption is orchestrated by the Anaphase Promoting Complex and its coactivator Cdc20, APC/CCdc20, a protein complex that is active at anaphase onset
Slug Flow-Induced Vibration and Vortex-Induced Vibration of Catenary Risers
Get PDFPhD ThesisSubsea risers are widely used in the offshore oil and gas industry for deep-water productions where hydrocarbon flows of multiple phases may appear in several flow patterns. Characterized by the alternating liquid- and gas-dominant regions, slug flows are practically problematic due to a fluctuation in flow momenta and pressure potentially causing slug flow-induced vibration (SIV). Compared with a single-phase flow-induced vibration, fundamental understanding of SIV phenomena and the associated effects on the dynamics of catenary risers are still lacking.
Therefore, the present thesis aims to investigate SIV of flexible catenary risers subject to external excitations by the vortex-induced vibration (VIV). The literature of multiphase flows (gas-liquid flow patterns, flow maps) and flow-induced vibrations (SIV, VIV) is reviewed. A mechanistic steady-state slug model is considered to provide the slug flow-induced loads with a two-dimensional continuum riser model. A numerical study for fundamental planar dynamics of an inclined curved flexible riser carrying slug gas-liquid flows is conducted. Results demonstrate several SIV features and effects of slug flow characteristics (slug unit length, translational velocity and fluctuation frequencies) on the riser resonant dynamics. These fundamental observations enable a further study with combined VIV-SIV phenomena.
To incorporate the VIV effect, a phenomenological model based on wake oscillators is used and first implemented for an elastically mounted rigid cylinder in uniform flows subject to cross-flow/in-line VIV. The dynamical system described by coupled nonlinear cylinder-wake oscillators is solved by a numerical-analytical approach. Several important VIV characteristics are captured, highlighting abilities of this semi-empirical model in VIV prediction. By applying the distributed van der Pol wake-oscillators, this low-order model is then extended to the analysis of cross-flow-only VIV of a catenary riser under uniform flows perpendicular to its initial curvature plane. Some experimentally observed VIV features of flexible cylinders are numerically predicted. By comparing VIV-only with VIV-SIV responses, VIV is found to prevail in the combined external-internal excitation cases. Slug flows generally result in a significant multi-mode VIV, which becomes more pronounced at higher external flow velocities: this is attributed to the effect of softening stiffness of the pipe caused by internal slug flow mass and pressure leading to a higher-mode VIV.
To validate the numerical model of SIV, a small-scale flexible pipe conveying gas-liquid flows has been investigated experimentally for a catenary configuration. The experiment is carried out in an air-water test loop with a hanging tube section made of silica gel. Both slug flow-induced pipe motions and gas-liquid flow patterns are recorded simultaneously using non-intrusive high-speed cameras. Pressure variations are also measured at the pipe inlet and outlet by two pressure transducers. The flow-transporting system is tested by employing different gas-liquid flow rates. Occurrence of slug flows is captured at the relatively high ratios of the gas to liquid superficial velocities rendering a large-amplitude SIV of the flexible pipe. Under different flow conditions, slug flow characteristics are observed to vary significantly in terms of the travelling velocities, lengths and frequencies of different slug units. These entail the unsteady SIV with modulated amplitudes and frequencies. Some qualitative aspects of slug characteristics and SIV responses from numerical predictions are captured in the laboratory tests. Through comparisons of riser vibration responses and frequencies between numerical and experimental results, the prediction abilities of the mathematical model are recognized.
In summary, this thesis has investigated fundamental and potential SIV effects on catenary risers. A significant role of the slug unit length and the slug translational velocity should be considered individually for determining large-amplitude SIV which is of practical importance from a design viewpoint. Possible amplifications of riser responses due to slug flows are remarked in combined VIV-SIV scenarios. This is meaningful for the riser stress, fatigue and failure assessment. Although the present numerical model is limited to steady-state slug flows, it paves a way for a future development of computationally efficient tools benefiting the screening analysis of subsea risers transporting gas-liquid slug flows
Understanding the Molecular Mechanisms of Mitochondrial Muscle Disease using a Quantitative, Quadruple Immunofluorescent Assay
Get PDFPh. D. ThesisMitochondrial disease can be caused by pathogenic variants in either nuclear DNA or mitochondrial DNA (mtDNA). Affected patients present with a considerable genetic, clinical and biochemical heterogeneity, thereby complicating the diagnostic pathways. For pathogenic mtDNA variants, this is further compounded by heteroplasmy – the mixture of both wild-type and mutated mtDNA existing within a single cell. When the proportion of mutant mtDNA exceeds a critical threshold of mutated mtDNA molecules, the cell becomes biochemically dysfunctional. This dysfunction can be identified using a recently described quadruple immunofluorescent assay through the assessment of Complex I, Complex IV and porin protein expression in individual fibres within 10μm skeletal muscle sections from patients. The assessment of Complex I dysfunction is crucial, as deficiency of this particular respiratory chain complex is the most common form of mitochondrial dysfunction observed in mitochondrial disease. This thesis describes two studies that have utilised this assay with the aim of i) improving the diagnosis of mitochondrial disease and ii) understanding the molecular mechanisms underlying mitochondrial disease and the expression of a biochemical defect.
Given the lack of histochemical tests to interrogate Complex I in skeletal muscle, the viability of the quadruple immunofluorescent assay in detecting Complex I deficiency was assessed in a heterogeneous cohort of patients with genetically-proven pathogenic variants within mtDNA-encoded or nuclear-encoded (structural Complex I subunits or assembly factors) genes. Clear diagnostic potential was evident, particularly for patients with nuclear-encoded gene defects whilst highlighting the necessity of complete mitochondrial genome sequencing in the diagnostic work up of patients.
The quadruple immunofluorescent assay was also used to characterise –at a single cell level - the biochemical status of skeletal muscle fibres from patients harbouring the m.3243A>G mutation, the most prevalent, pathogenic heteroplasmic mtDNA point mutation. Two groups of patients were identified: those showing a predominance of Complex I deficiency and those expressing no biochemical defect in their skeletal muscle. Isolation of single muscle fibres enabled investigation of the relationship between the biochemical deficiency and single fibre m.3243A>G heteroplasmy, identifying differences in the biochemical threshold for Complex I deficiency between patients
Exploring Grandparenthood in the Context of Grandchild Disability: Intergenerational Perspectives
Get PDFPhD ThesisAvailable evidence indicates that grandparents can play a crucial role in the support of their
grandchildren with a disability, their adult children and the entire family unit. Shifts in family
relationships and roles, demographics and broader contexts are placing greater importance on
the role of grandparents. The lived experience of grandparenting in the context of grandchild
disability has received scant attention in research, policy or practice, particularly in the UK, yet
advances in neonatal and paediatric care have resulted in more children surviving with disabling
conditions.
This qualitative study aimed to explore the role of grandparents in the lives of children with a
disability from their perspective and that of the parents, and to understand the impact of ageing
on the dynamics of intergenerational support. In-depth interviews were conducted with
grandparents (n=21) and parents (n=11) of children aged 4-19 years with disabilities including
autism spectrum disorder, cerebral palsy, and congenital disorders.
Three overarching themes were identified: experiences of diagnosis and family adaptation,
maintaining intergenerational harmony, and changes in the dynamics of care with age.
Grandparents were a vital source of support, aiding families to adapt and continually adjust to
the needs of the grandchild. Grandparents, however, also recounted their own hardships, which
included financial challenges, long hours of care, and lack of engagement in third age activities.
Capturing both parent and grandparent perspectives demonstrated extensive intergenerational
negotiation, and at times conflict, around employment, role boundaries, types of support, and
the emotional work in maintaining intergenerational harmony. Intergenerational support shifted
and was continually renegotiated across the lifecourse trajectory. As well as giving voice to a
previously under-researched group, these findings inform key debates, such as how extending
working life policies can impact on families with disabled children
Development of mass spectrometric approaches to advance drug discovery
Get PDFPhD ThesisThe work in this thesis concerns the development of advanced mass spectrometric techniques to study model systems for drug discovery and accelerate the process of high-throughput screening (HTS) in the field of inflammation. This has primarily been achieved using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) to develop screening assays for anti-inflammatory compounds. MALDI-TOF-MS utilises a laser to ablate and consequently ionise biomolecules that are co-crystallised with a matrix. The absorbed laser energy enables the transfer of charge to the analyte molecule from the excited matrix molecules before acceleration into the TOF tube where analytes are then separated based on their mass to charge ratio (m/z) and detected to generate a mass spectrum. With the development of advanced mass spectrometers that have unmatched laser and digitiser speeds these instruments now rival established fluorescent based drug discovery assays.
The first study presented in this thesis is the development of a HTS assay to identify inhibitors of salt inducible kinases (SIKs) in vitro. These kinases are an attractive drug target as they mediate the production and secretion of the cytokine interleukin 10 (IL-10) in immune cells. When secreted, IL-10 dampens the pro-inflammatory response in surrounding macrophages and suppresses a further inflammatory response. Using a peptide-based assay, a proof of concept screen generated a positive correlation with an established fluorescent based assay, thus validating MALDI-TOF-MS as a viable alternative for HTS. Interestingly, several compounds were identified as kinase activators. Inhibitors of SIKs could be particularly beneficial for patients with autoimmune diseases who require immunosuppressive drugs as they may increase the production of IL-10, thus preventing excessive inflammation and subsequent onset of pathogenesis.
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MALDI-TOF-MS was then applied to the development of cellular based assays to screen compounds against cell types that transition into different phenotypes. Rigorous development of a technically reproducible sample preparation technique enabled robust phenotyping of different cell lines. This method was then successfully applied to study mouse embryonic stem cells (mESCs), which transition between naïve and pluripotent phenotypes under pharmacologically controlled conditions. Moreover, a cellular assay to identify inhibitors of inflammation in the presence of a bacterial ligand lipopolysaccharide (LPS) in monocytes was developed. Here, the BCR-ABL inhibitor nilotinib was identified as having anti-inflammatory properties. Interestingly, its first-generation counterpart imatinib was not identified as having this effect, suggesting novel off-target effects for nilotinib. Its anti-inflammatory properties were then validated by cell biology techniques that demonstrated the reduction of the pro-inflammatory cytokine TNF-α in nilotinib-treated cells.
Finally, I utilised advanced high-resolution mass spectrometry to undertake secretomic analysis of L929 fibroblasts, which are commonly used to generate supplemented media that aids the differentiation of bone marrow-derived macrophages (BMDMs). These cells are promising drug targets and the understanding of their differentiation state is paramount to potential drug screening campaigns. A detailed proteomic profile of L929 supplemented media, as well an in-depth proteomic profile of BMDMs differentiated under various conditions were generated.Taken together the work of this thesis contributes valuable knowledge in the field of how mass spectrometry can be utilised to advance HTS campaigns as well as provide valuable insight to the biology of promising drug models
Neurocognitive patterns and progression of mild cognitive impairment with Lewy bodies or Alzheimer’s disease
Get PDFPhD ThesisMild cognitive impairment is a heterogeneous condition; while typically viewed as the transition stage between healthy cognitive function and dementia, the cognitive and clinical patterns of this condition vary, as do its prognosis. The two most common neurodegenerative dementias, dementia with Lewy bodies and Alzheimer’s disease, differ in their clinical features, patterns of cognitive impairment, and prognosis. It is not known whether the respective mild cognitive impairment stages preceding onset of dementia also differ in these characteristics. Two cohorts of people with recent mild cognitive impairment diagnosis were assessed, undergoing annual review of cognition, diagnosis and presence of clinical features of Lewy body disease by an expert panel of old age psychiatrists, and repeated imaging, to reach a consensus diagnosis of either mild cognitive impairment due to Alzheimer’s disease, or with Lewy bodies, in line with current consensus criteria for these. Making use of annually repeated cognitive assessment and clinical diagnostic information, the longitudinal progression of these two conditions was characterised with flexible statistical methods, using the first cohort for model development, and the second for validation: different trajectories of decline in specific cognitive domains were observed in the diagnostic groups, reflecting typical patterns of impairment in their respective dementia syndromes. Mild cognitive impairment with Lewy bodies was also observed to have a worse prognosis, in keeping with its dementia stage, with a greater risk of progressive cognitive decline, and faster onset of dementia. Individuals with neuropsychiatric symptoms (cognitive fluctuations and visual hallucinations) were also at more risk of decline than those with slower-developing features (REM sleep behaviour disorder or parkinsonism) or those with Alzheimer’s disease. Mild cognitive impairment with Lewy bodies may therefore feature early cognitive, clinical, and prognostic differences from Alzheimer’s disease, reflecting its eventual dementia syndrome
Assessment Change in Omani Post-Basic (16- 18) Biology Education: evaluating the journey from policy as written, to policy as enacted and policy as experienced.
Get PDFPh. D. ThesisThis study explored the phenomenon of changes to Biology assessment in Omani secondary
education with a focus on formative assessment (FA). The recent changes in curricula,
following the implementation of the new post-basic education system in Oman (for students
aged 16- 18 years old) in the academic year 2007/ 2008 required new assessment practices.
This research study investigates how the Biology assessment was written by assessment
policy makers, how it was enacted by Biology teachers and how it was experienced by
Biology students using a qualitative dominant case study approach. Data was collected via
two semi-structured interviews with assessment policy makers, four with Biology teachers
and six focus groups with students; a 30-item questionnaire to 96 students; 11 non-participant
lesson observations and assessment document analysis.
The research explored both the benefits and challenges of implementing assessment change
in the Biology classroom. This study provides an insight into the understandings of the
assessment practices of four Biology teachers. The impact of a top-down policy approach on
teachers’ perceptions and the enactment of both formative and summative assessment is
explored. The findings revealed the importance of dialogue with Biology teachers during the
planning or design phase of changes to assessment policy for its implementation to succeed.
The teachers had not experienced any support or further training in assessment practices. The
study also valued the voices of students regarding assessment, which considerably affected
their approaches to study. The findings suggest that participating students could see the
benefits of talking with their peers to understand some topics related to Biology problems. It
is recommended that the Ministry of Education and local educational authority should
support Biology teachers in their assessment practices through dialogue and professional
development activities to ensure the successful implementation of the assessment system.
Keywords: FA, SA, Post-basic education, Assessment policy.Ministry of Higher
Educatio