Microneedles (MNs) are minimally invasive drug delivery devices that can be used for transdermal drug delivery (TDD). They have the potential to enhance drug delivery for therapeutic agents which are currently difficult to deliver, costly, or cause a wide range of side effects. The therapeutic agent must normally overcome the highly hydrophobic barrier of the skin’s outermost layer, the stratum corneum (s.c.), in order to be delivered transdermally. MNs have a height of 50 m to 1500 m and overcome the barrier by penetrating the s.c. and creating aqueous pores. They can be fabricated from different materials, such as silicon, metal, glass, sugars, and polymers. Dissolving polymeric MNs are used to penetrate the s.c. and release the drug incorporated in the polymeric matrix into the epidermis when they dissolve by taking up skin interstitial fluid (ISF). This means that when nanoparticles (NPs) are delivered using dissolving MNs, the amount delivered is dependent upon the amount localized in the needle portion of the array. The present study developed and characterized dissolving MNs using a novel two-step process which improves the loading distribution of the therapeutic agent in the needle portion of the array. The two-step process fabricates the needles and the baseplate individually, allowing both to be fabricated from different for- mulations and optimized separately. The MNs were loaded with a model protein, ovalbumin (OVA), and poly(lactic-co-glycolide acid) (PLGA) NPs. When the loading distribution was measured for the NPs, it was found that about 80% of the NPs were localized in the needles. This is almost double the localization compared to arrays using the standard fabrication method. The MNs loaded with NPs were used in an in vivo mouse model. The NPs were successfully delivered into the skin of the mice and were detected in the draining lymph nodes. This showed that the developed TDD device can be used for targeted delivery of the lymphatic system. This study was overall successful in engineering a MN-mediated intradermal NP delivery device that can be used to target the lymphatic system.

Background: Hyperinsulinaemic hypoglycaemia (HH) is characterized by unregulated secretion of insulin in the presence of hypoglycaemia. Mutations in nine different genes (ABCC8, KCNJ11, GLUD1, HNF4A, HNF1A, GCK, HADH, SLC16A1 and UCP2) have so far been identified as a cause of HH. Mutations in ABCC8 and KCNJ11, which encode the sulfonylurea receptor 1 (SUR1) and potassium inward-rectifying 6.2 (Kir6.2) subunits of ATP-sensitive potassium channel (KATP channel), are the most common cause of HH. At least two (possibly more) well-described histological subtypes are associated with HH: a focal form and a diffuse form. Diffuse HH are most commonly due to recessive and dominant mutations in ABCC8 and KCNJ11. Focal HH results due to somatic loss of the maternal 11p allele (11p15.1 to 11p15.5) involving the ABCC8 and KCNJ11 region in patients with paternally inherited mutation in ABCC8 or KCNJ11. HH can be transient and resolve within few weeks. Transient HH is seen in association with intrauterine growth restriction, maternal diabetes mellitus, perinatal asphyxia, erythroblastosis fetalis, maternal administration of sulfonylureas, and intravenous glucose infusions during labour. In large series of HH patients, the underlying genetic cause was not identified in approximately 50% of the patients. Whole-exome sequencing is a powerful and cost-effective tool for identifying genetic basis of diseases. It involves sequencing the protein coding regions of the human genome. Aim: To identify novel genetic mechanisms of HH. To functionally characterize two novel KATP channel mutations associated with a unique clinical phenotype. Patients: Four patients had protein-sensitive HH with normal serum ammonia. Two families (one consanguineous and one non-consanguineous) had two affected siblings with HH. All these patients had negative molecular genetics for ABCC8, KCNJ11, GLUD1 and HADH. One patient had a unique phenotype of HH and cardiac arrhythmias. In addition, three patients had two novel KATP channel mutations associated with a unique clinical phenotype (transient HH and combined focal/diffuse HH). Methods: Whole-exome sequencing (WES) was performed on nine patients (from seven families), their parents and unaffected siblings to identify novel, deleterious gene variants. Based on the available biological information, two novel gene variants (p.F108del K2P17 and p.A422V Kv6.2) were considered potential disease causing. The mutations were created in plasmid constructs containing the WT cDNA sequence for KCNK17 (K2P17) and KCNG2 (Kv6.2) using site-directed mutagenesis. These constructs were transfected into HEK293 cells, which were studied by whole-cell patch clamping. Two novel KATP channel mutations (p.T1516M SUR1 and p.*391Rext*94 Kir6.2) were also studied. For studying the p.*391Rext*94 Kir6.2 mutation, the WT human cDNA and 3′UTR sequence of KCNJ11 was cloned into pcDNA 3.1 vector. The KATP channel mutations were created in plasmid construct containing the WT cDNA sequence for ABCC8/KCNJ11 using site-directed mutagenesis. These constructs were transfected into HEK293 cells, which were then studied by whole-cell patch clamping. The p.*391Rext*94 Kir6.2 is a non-stop mutation and the transcripts can undergo degradation by non-stop decay phenomenon. The presence of p. *391Rext*94 Kir6.2 mutant transcript in the pancreatic tissue of the affected patient was studied by cDNA sequencing and Western blotting on the patient’s pancreatic tissue extracted RNA and protein fraction respectively. Results: WES identified two potential disease-causing heterozygous gene variants (p.F108del K2P17 and p.A422V Kv6.2) in a family with a phenotype of HH and cardiac arrhythmia, which were confirmed by Sanger sequencing. Whole-cell patch clamping experiments on HEK293 cells proved both variants to be pathogenic under heterozygous expression. Whole-cell patch clamping experiments on the two novel KATP channel mutations (p.T1516M SUR1 and p.*391Rext*94 Kir6.2) was indicative of pathogenic nature of the mutation. Analysis of the pancreatic tissue, obtained at surgery from the patient with non-stop KCNJ11 mutation (p.*391Rext*94 Kir6.2), by cDNA sequencing and Western blotting established the presence of transcript and protein with non-stop mutation. Conclusions: Mutations in KCNK17 (K2P17) and KCNG2 (Kv6.2) are potential novel genetic mechanisms for HH and cardiac arrhythmias. More patients with similar phenotype need to be screened for mutations in these two genes. A novel mechanism for HH (combined focal and diffuse HH phenotype) and molecular basis for a transient type of HH was identified.

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Aims of Thesis • Section 1 – To ascertain the incidence of non alcoholic fatty liver disease in patients with growth hormone deficiency and the impact of hormone replacement. • Section 2 – Validation of the glucagon stimulation test in healthy subjects and hypopituitary patients. • Section 3 – A prospective longitudinal study of pituitary function in survivors of Subarachnoid Haemorrhage and evaluation of changes in Quality of life following growth hormone replacement. • Section 4 – An analysis of the Pfizer KIMS database to evaluate changes in Quality of Life and Body Composition following Growth Hormone Replacement in patients with Traumatic Brain Injury.

The research for this thesis examined the effects of diabetic peripheral neuropathy (DPN) on biomechanical factors related to the risk of falling during locomotor tasks, and the effects of a 16-week multi-factorial intervention on the identified impairments. The speed of ankle and knee strength generation, and muscular activations were measured during stair ascent and descent; and minimum toe clearance, stepping accuracy, and visual gaze parameters were measured during level ground walking. Patients with DPN, diabetes patients with no neuropathy and non-diabetic controls were measured before and after a 16-week intervention consisting of high-load resistance exercises and a visual gaze training task. Patients with DPN displayed slower ankle and knee strength generation during stair ascent and descent than healthy controls (p<0.05). Post-intervention, strength was generated faster at the ankle and knee during both tasks (p<0.05), which is expected to improve stability during the weight acceptance phase. During level ground walking, patients with DPN displayed a higher minimum toe clearance (p<0.05), which is expected to reduce the risk of tripping on smaller, less observable hazards; but displayed a decreased stepping accuracy (p<0.05), which may reduce the ability to avoid tripping hazards. Stepping accuracy was improved as a result of the intervention (p<0.05), which may originate from improvements in visual gaze strategy and motor control, contributing to reduce the risk of tripping in patients with DPN. Biomechanical impairments during locomotion were observed in patients with DPN; however, the intervention improved these aspects and may reduce the risk of falling in this population.

Diabetes mellitus is considered to be a global epidemic and a growing burden on public health. It is associated with significant morbidity and mortality rates. For that reason, along with the advancement in technology, healthcare providers are moving towards telemonitoring to identify self-management approaches as a means to address healthcare needs where patients live and shift the power and responsibility to them. This dissertation investigates the impact of remote monitoring on the management of diabetes mellitus. The primary research question was: does remote monitoring help achieve self-management of diabetes mellitus and improve the clinical outcome. A systematic review of the literature showed that remote monitoring is a promising tool yet implementation needs to be supported with more research and evidence. A prospective study was conducted to explore three different remote monitoring solutions to evaluate feasibility and establish the benefits of utilizing remote monitoring for self-managing type-2 diabetes mellitus. Furthermore, a retrospective evaluation of a large-scale telemonitoring solution was conducted to support the hypothesis and help addressing the question of this thesis. A theoretical underpinning of innovation adoption and diffusion and health behaviour change theories was developed to fully understand technology acceptance amongst users in relation to this thesis. This dissertation pragmatically discusses the methodological challenges a researcher would face during a clinical trial in a diabetes clinic.

The thesis comprises two pieces of research: A systematic literature review which synthesises the literature examining the psychosocial variables related to glycaemic control in adolescents with type 1 diabetes, and an empirical paper which explores the lived experience of adolescents who take methotrexate to manage their Juvenile Idiopathic Arthritis (JIA). The systematic review comprised searches of three electronic databases (Cumulative Index to Nursing & Allied Health Literature [CINAHL], Psychinfo and Medline) which yielded 1136 separate records. Eligibility assessment criteria were applied, resulting in fifty-five articles which were assessed for quality prior to data extraction. A meta-analysis and narrative synthesis of the eight most frequently occurring psychosocial variables related to metabolic control is presented. A vast number of disparate psychosocial variables were presented within the literature. A scan of the correlation coefficients in the extracted data indicated no variables showed strong associations with glycaemic control. Narrative and meta-analysis of the eight variables most frequently measured in the literature showed weak to moderate associations with metabolic control in adolescents with type 1 diabetes, however heterogeneity within results suggests issues with study quality. Tentative conclusions suggest that psychosocial variables are largely unrelated to glycaemic control with the exception of internalising behaviours; suggesting psychosocial interventions seeking to improve self-management through glycaemic control should focus on reducing the internal distress of the adolescent. The review highlighted the need to consistently utilise validated, reliable assessment measures in future research. The empirical paper used qualitative methodology to explore the in-depth, lived experience of teenagers who take methotrexate to manage their symptoms of Juvenile Idiopathic Arthritis. A topic guide was used to facilitate individual interviews with six female participants aged 13-17 who were taking or had taken methotrexate within the last year. Analysis of interview transcripts using Interpretative Phenomenological Analysis yielded three master themes: 'Who am I, and who am I to you?: Relationships with self and others', 'Methotrexate: Friend, foe or forgotten', and 'Surviving on the battlefield: Attack, retreat and defeat'. Themes are discussed in relation to existing literature in JIA and chronic illness in childhood, with inconsistencies and new content highlighted. The clinical implications of the research are discussed.

Research thesis encompassing two papers: Psychological factors associated with diabetes self-management among adolescents with Type 1 diabetes: a systematic review The purpose of the review was to determine what psychological factors are associated with diabetes self-management. Twenty-one articles were determined to be eligible for this review. Numerous psychological factors were found to be associated with self management; however, correlations were typically small to moderate. Study validity was variable and there was little overlap between psychological factors examined. Variables are presented in a narrative synthesis. The strongest associations were found between social anxiety and diet (among boys); greater intrinsic motivation, conscientiousness and diet; and extraversion and exercise. Evidence exists for relationships between psychological factors and diabetes self-management but due to the individual nature of the studies, firm conclusions cannot be drawn. Future research needs to attempt replication and utilise validated measures to provide a stronger evidence base from which to develop theory for this population. The relationships between diabetes distress, illness perceptions and glycaemic control in adults with Type 2 diabetes This study aimed to investigate whether illness perceptions moderate the relationship between diabetes distress and glycaemic control. Participants with Type 2 DM attending diabetes outpatient clinics (n = 82) completed the Diabetes Distress Scale 17, Brief Illness Perceptions Questionnaire and the Patient Health Questionnaire 9 as well as providing demographic and clinical information. Most recent HbA 1 c and BMI were collected from medical records. Personal Control was the only significant contributor in the final regression model predicting HbA 1 c. The relationship between regimen-related distress and HbA 1 c was mediated by personal control. Moderating effects were non-significant. Personal control has an important role in explaining the link between diabetes distress and HbA 1 c. Psychological interventions seeking to improve HbA 1 c need to address individuals' perceptions of personal control.

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